United States May 2006
EPW600
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EPA/600/R-05/144bB
May 2006
Air Quality Criteria for Lead
Volume II
National Center for Environmental Assessment-RTF Office
Office of Research and Development
U.S. Environmental Protection Agency
Research Triangle Park, NC
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DISCLAIMER
This document is a second external review draft being released for review purposes only
and does not constitute U.S. Environmental Protection Agency policy. Mention of trade names
or commercial products does not constitute endorsement or recommendation for use.
PREFACE
National Ambient Air Quality Standards (NAAQS) are promulgated by the United States
Environmental Protection Agency (EPA) to meet requirements set forth in Sections 108 and 109
of the U.S. Clean Air Act. Those two Clean Air Act sections require the EPA Administrator
(1) to list widespread air pollutants that reasonably may be expected to endanger public health or
welfare; (2) to issue air quality criteria for them that assess the latest available scientific
information on nature and effects of ambient exposure to them; (3) to set "primary" NAAQS to
protect human health with adequate margin of safety and to set "secondary" NAAQS to protect
against welfare effects (e.g., effects on vegetation, ecosystems, visibility, climate, manmade
materials, etc); and (5) to periodically review and revise, as appropriate, the criteria and NAAQS
for a given listed pollutant or class of pollutants.
Lead was first listed in the mid-1970's as a "criteria air pollutant" requiring NAAQS
regulation. The scientific information pertinent to Lead NAAQS development available at the
time was assessed in the EPA document Air Quality Criteria for Lead; published in 1977. Based
on the scientific assessments contained in that 1977 lead air quality criteria document (1977 Lead
AQCD), EPA established a 1.5 |ig/m3 (90-day average) Lead NAAQS in 1978.
To meet Clean Air Act requirements noted above for periodic review of criteria and
NAAQS, new scientific information published since the 1977 Lead AQCD was later assessed in
a revised Lead AQCD and Addendum published in 1986 and in a Supplement to the 1986
AQCD/Addendum published by EPA in 1990. A 1990 Lead Staff Paper, prepared by EPA's
Office of Air Quality Planning and Standards (OPQPS), drew upon key findings and conclusions
from the 1986 Lead AQCD/Addendum and 1990 Supplement (as well as other OAQPS-
sponsored lead exposure/risk analyses) in posing options for the EPA Administrator to consider
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with regard to possible revision of the Lead NAAQS. However, EPA decided not to revise the
lead NAAQS at that time.
The purpose of this revised Lead AQCD is to critically evaluate and assess the latest
scientific information that has become available since the literature assessed in the above 1986
Lead AQCD/Addendum and 1990 Supplement, with the main focus being on pertinent new
information useful in evaluating health and environmental effects of ambient air lead exposures.
This includes discussion in this document of information regarding: the nature, sources,
distribution, measurement, and concentrations of lead in the environment; multimedia lead
exposure (via air, food, water, etc.) and biokinetic modeling of contributions of such exposures
to concentrations of lead in brain, kidney, and other tissues (e.g., blood and bone concentrations,
as key indices of lead exposure).; characterization of lead health effects and associated exposure-
response relationships; and delineation of environmental (ecological) effects of lead. This
Second External Review Draft of the revised Lead AQCD mainly assesses pertinent literature
published or accepted for publication through June, 2004.
The First External Review Draft (dated December 2005) of the revised Lead AQCD
underwent public comment and was reviewed by the Clean Air Scientific Advisory Committee
(CASAC) at a public meeting held in Durham, NC on February 28-March, 2006. The public
comments received and CASAC recommendations were taken into account in making
appropriate revisions to this document and incorporating them into this Second External Review
Draft (dated May, 2006) which is being released for further public comment and CASAC review
at a public meeting to be held June 28-29, 2006. Public comments and CASAC advice received
on these Second External Review Draft materials will be taken into account in incorporating
further revisions into the final version of this Lead AQCD, which must be completed and issued
by October 1, 2006. Evaluations contained in the present document will be drawn on to provide
inputs to an associated Lead Staff Paper prepared by EPA's Office of Air Quality Planning and
Standards (OAQPS), which will pose options for consideration by the EPA Administrator with
regard to proposal and, ultimately, promulgation of decisions on potential retention or revision,
as appropriate, of the current Lead NAAQS.
Preparation of this document has been coordinated by staff of EPA's National Center
for Environmental Assessment in Research Triangle Park (NCEA-RTP). NCEA-RTP scientific
staff, together with experts from academia, contributed to writing of document chapters.
Il-iv
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Earlier drafts of document materials were reviewed by scientists from other EPA units
and by non-EPA experts in several public peer consultation workshops held by EPA in
July/August 2005.
NCEA acknowledges the valuable contributions provided by authors, contributors, and
reviewers and the diligence of its staff and contractors in the preparation of this draft document.
II-v
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Air Quality Criteria for Lead
(Second External Review Draft)
VOLUME I
EXECUTIVE SUMMARY E-l
1. INTRODUCTION 1-1
2. CHEMISTRY, SOURCES, AND TRANSPORT OF LEAD 2-1
3. ROUTES OF HUMAN EXPOSURE TO LEAD AND OBSERVED
ENVIRONMENTAL CONCENTRATIONS 3-1
4. LEAD TOXICOKINETICS AND MEASUREMENT/MODELING OF HUMAN
EXPOSURE IMPACTS ON INTERNAL TISSUE DISTRIBUTION OF LEAD 4-1
5. TOXICOLOGICAL EFFECTS OF LEAD IN LABORATORY ANIMALS,
HUMANS, AND IN VITRO TEST SYSTEMS 5-1
6. EPIDEMIOLOGIC STUDIES OF HUMAN HEALTH EFFECTS
ASSOCIATED WITH LEAD EXPOSURE 6-1
7. INTEGRATIVE SYNTHESIS OF LEAD EXPOSURE/HEALTH
EFFECTS INFORMATION 7-1
8. ENVIRONMENTAL EFFECTS OF LEAD 8-1
VOLUME II
CHAPTER 5 ANNEX (TOXICOLOGICAL EFFECTS OF LEAD IN
LABORATORY ANIMALS, HUMANS, AND IN VITRO TEST SYSTEMS) AX5-1
CHAPTER 6 ANNEX (EPIDEMIOLOGIC STUDIES OF HUMAN HEALTH
EFFECTS ASSOCIATED WITH LEAD EXPOSURE) AX6-1
CHAPTER 8 ANNEX (ENVIRONMENTAL EFFECTS OF LEAD) AX8-1
Il-vi
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Table of Contents
Page
List of Tables Il-xi
List of Figures II-xx
Authors, Contributors, and Reviewers II-xxii
U.S. Environmental Protection Agency Project Team for Development of Air Quality
Criteria for Lead II-xxviii
U.S. Environmental Protection Agency Science Advisory Board (SAB) Staff Office
Clean Air Scientific Advisory Committee (CASAC) II-xxx
Abbreviations and Acronyms II-xxxii
AX5. CHAPTER 5 ANNEX AX5-1
ANNEX TABLES AX5-2 AX5-1
ANNEX TABLES AX5-3 AX5-15
ANNEX TABLES AX5-4 AX5-37
ANNEX TABLES AX5-5 AX5-60
ANNEX TABLES AX5-6 AX5-75
ANNEX TABLES AX5-7 AX5-111
ANNEX TABLES AX5-8 AX5-121
ANNEX TABLES AX5-9 AX5-147
ANNEX TABLES AX5-10 AX5-157
ANNEX TABLES AX5-11 AX5-187
AX6. CHAPTER 6 ANNEX AX6-1
ANNEX TABLES AX6-2 AX6-2
ANNEX TABLES AX6-3 AX6-28
ANNEX TABLES AX6-4 AX6-68
ANNEX TABLES AX6-5 AX6-117
ANNEX TABLES AX6-6 AX6-153
ANNEX TABLES AX6-7 AX6-161
ANNEX TABLES AX6-8 AX6-181
ANNEX TABLES AX6-9 AX6-195
ANNEX SECTION AX6-10 AX6-236
AX8. CHAPTER 8 ANNEX - ENVIRONMENTAL EFFECTS OF LEAD AX8-1
AX8.1 TERRESTRIAL ECOSYSTEMS AX8-1
AX8.1.1 Methodologies Used in Terrestrial Ecosystems
Research AX8-1
AX8.1.1.1 Lead Isotopes and Apportionment AX8-1
AX8.1.1.2 Speciation in Assessing Lead
Bioavailability in the Terrestrial
Environment AX8-3
AX8.1.1.3 Tools for Bulk Lead Quantification
and Speciation AX8-9
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Table of Contents
(cont'd)
AX8.1.1.4 Biotic Ligand Model AX8-18
AX8.1.1.5 Soil Amendments AX8-20
AX8.1.1.6 Future Needs AX8-23
AX8.1.2 Distribution of Atmospherically Delivered Lead in
Terrestrial Ecosystems AX8-23
AX8.1.2.1 Speciation of Atmospherically-
Delivered Lead in Terrestrial
Ecosystems AX8-26
AX8.1.2.2 Tracing the Fate of Atmospherically
Delivered Lead in Terrestrial
Ecosystems AX8-33
AX8.1.2.3 Inputs/Outputs of Atmospherically
Delivered Lead in Terrestrial
Ecosystems AX8-35
AX8.1.3 Terrestrial Species Response/Mode of Action AX8-39
AX8.1.3.1 Lead Uptake AX8-39
AX8.1.3.2 Resistance Mechanisms AX8-45
AX8.1.3.3 Physiological Effects of Lead AX8-47
AX8.1.3.4 Factors that Modify Organism
Response AX8-49
AX8.1.3.5 Summary AX8-55
AX8.1.4 Exposure-Response of Terrestrial Species AX8-58
AX8.1.4.1 Summary of Conclusions from the
1986 Lead Criteria Document AX8-59
AX8.1.4.2 Recent Studies on the Effects of
Lead on Primary Producers AX8-61
AX8.1.4.3 Recent Studies on the Effects of
Lead on Consumers AX8-62
AX8.1.4.4 Recent Studies on the Effects of
Lead on Decomposers AX8-81
AX8.1.4.5 Summary AX8-86
AX8.1.5 Effects of Lead on Natural Terrestrial Ecosystems AX8-88
AX8.1.5.1 Effects of Terrestrial Ecosystem
Stresses
on Lead Cycling AX8-89
AX8.1.5.2 Effects of Lead Exposure on
Natural Ecosystem Structure
and Function AX8-94
AX8.1.5.3 Effects of Lead on Energy Flows
and Biogeochemical Cycling AX8-99
AX8.1.5.4 Summary AX8-105
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Table of Contents
(cont'd)
AX8.2 AQUATIC ECOSYSTEMS AX8-106
AX8.2.1 Methodologies Used in Aquatic Ecosystem
Research AX8-106
AX8.2.1.1 Analytical Methods AX8-106
AX8.2.1.2 Ambient Water Quality Criteria:
Development AX8-108
AX8.2.1.3 Ambient Water Quality Criteria:
Bioavailability Issues AX8-110
AX8.2.1.4 Sediment Quality Criteria:
Development and Bioavailability
Issues AX8-112
AX8.2.1.5 Metal Mixtures AX8-115
AX8.2.1.6 Background Lead AX8-116
AX8.2.2 Distribution of Lead in Aquatic Ecosystems AX8-116
AX8.2.2.1 Speciation of Lead in Aquatic
Ecosystems AX8-117
AX8.2.2.2 Spatial Distribution of Lead in
Aquatic Ecosystems AX8-121
AX8.2.2.3 Tracing the Fate and Transport
of Lead in Aquatic Ecosystems AX8-138
AX8.2.2.4 Summary AX8-143
AX8.2.3 Aquatic Species Response/Mode of Action AX8-143
AX8.2.3.1 Lead Uptake AX8-144
AX8.2.3.2 Resistance Mechanisms AX8-150
AX8.2.3.3 Physiological Effects of Lead AX8-157
AX8.2.3.4 Factors That Modify Organism
Response to Lead AX8-160
AX8.2.3.5 Factors Associated with Global
Climate Change AX8-172
AX8.2.3.6 Summary AX8-173
AX8.2.4 Exposure/Response of Aquatic Species AX8-173
AX8.2.4.1 Summary of Conclusions From
the Previous Criteria Document AX8-173
AX8.2.4.2 Recent Studies on Effects of Lead
on Primary Producers AX8-175
AX8.2.4.3 Recent Studies on Effects of Lead
on Consumers AX8-181
AX8.2.4.4 Recent Studies on Effects of Lead
on Decomposers AX8-191
AX8.2.4.5 Summary AX8-191
Il-ix
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Table of Contents
(cont'd)
AX8.2.5 Effects of Lead on Natural Aquatic Ecosystems AX8-192
AX8.2.5.1 Case Study: Coeur d'Alene
River Watershed AX8-193
AX8.2.5.2 Biotic Condition AX8-195
AX8.2.5.3 Summary AX8-206
REFERENCES AX8-208
II-x
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List of Tables
Number Page
AX5-2.1 Effect of Lead on Erythrocyte Morphology, Mobility, and Other
Miscellaneous Parameters AX5-2
AX5-2.2 Lead, Erythrocyte Heme Enzymes, and Other Parameters AX5-6
AX5-2.3 Lead Binding and Transport in Human Erythrocytes AX5-9
AX5-2.4 Lead Effects on Hematological Parameters AX5-10
AX5-2.5 Lead Interactions with Calcium Potassium in Erythrocytes AX5-12
AX5-2.6 Lead, Heme and Cytochrome P-450 AX5-13
AX5-2.7 Lead, Erythrocyte Lipid Peroxidation, and Antioxidant Defense AX5-14
AX5-3.1 Summary of Key Studies on Neurochemical Alterations AX5-16
AX5-3.2 Summary of Key Studies on Neurophysiological Assessments AX5-20
AX5-3.3 Summary of Key Studies on Changes in Sensory Function AX5-21
AX5-3.4 Summary of Key Studies on Neurobehavioral Toxicity AX5-22
AX5-3.5 Summary of Key Studies on Cell Morphology and Metal Disposition AX5-31
AX5-3.6 Key Studies Evaluating Chelation of Pb in Brain AX5-33
AX5-4.1 Effect of Lead on Reproduction and Development in Mammals AX5-3 8
AX5-4.2 Effect of Lead on Reproduction and Development in Mammals AX5-45
AX5-4.3 Effect of Lead on Reproduction and Development in Mammals AX5-55
AX5-5.1 In Vivo and In Vitro Studies of the Effects of Lead Exposure on
Production and Metabolism of Reactive Oxygen Species (ROS),
Nitric Oxide (NO), and Soluble Guanylate Cyclease (sCG) AX5-61
AX5-5.2 Studies of the Effects of Lead Exposure on PKC Activity,
Activation, and Apoptosis AX5-66
AX5-5.3 Studies of the Effects of Lead Exposure on Blood Pressure and
Adrenergic System AX5-67
Il-xi
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List of Tables
(cont'd)
Number
AX5-5.4 Studies of the Effects of Lead Exposure on Renin-angiotensin System,
Kallikrein-Kinin System, Prostaglandins, Endothelin, and Atrial
Natriuretic Peptide (ANP) AX5-69
AX5-5.5 Studies of Effect of Lead on Vascular Contractility AX5-70
AX5-5.6 Effects of Lead on Cultured Endothelial Cell Proliferation,
Angiogenesis, and Production of Heparan Sulfate Proteoglycans
andtPA AX5-71
AX5-5.7 Studies of the Effect of Lead on Cultured Vascular Smooth
Muscle Cells AX5-74
AX5-6.1 Genotoxic/Carcinogenic Effects of Lead - Laboratory Animal Studies AX5-76
AX5-6.2 Genotoxic/Carcinogenic Effects of Lead - Human Cell Cultures AX5-78
AX5-6.3 Genotoxic/Carcinogenic Effects of Lead - Carcinogenesis Animal
Cell Cultures AX5-79
AX5-6.4 Genotoxic/Carcinogenic Effects of Lead - Genotoxicity Laboratory
Animal Studies AX5-81
AX5-6.5 Genotoxic/Carcinogenic Effects of Lead - Genotoxicity Human Cell
Cultures Mutagenesis AX5-85
AX5-6.6 Genotoxic/Carcinogenic Effects of Lead - Genotoxicity Human Cell
Cultures Clastogenicity AX5-86
AX5-6.7 Genotoxic/Carcinogenic Effects of Lead - Genotoxicity Human Cell
Cultures DNA Damage AX5-88
AX5-6.8 Genotoxic/Carcinogenic Effects of Lead - Genotoxicity Animal Cell
Cultures Mutagenicity AX5-90
AX5-6.9 Genotoxic/Carcinogenic Effects of Lead - Genotoxicity Animal Cell
Cultures Clastogenicity AX5-92
AX5-6.10 Genotoxic/Carcinogenic Effects of Lead - Genotoxicity Animal Cell
Cultures DNA Damage AX5-95
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List of Tables
(cont'd)
Number
AX5-6.11 Genotoxic/Carcinogenic Effects of Lead - Genotoxicity
Non-Mammalian Cultures AX5-97
AX5-6.12 Genotoxic/Carcinogenic Effects of Lead - Genotoxicity as it Pertains
to Potential Developmental Effects AX5-98
AX5-6.13 Genotoxic/Carcinogenic Effects of Lead - Genotoxicity as it Pertains
to Potential Developmental Effects - Children AX5-99
AX5-6.14 Genotoxic/Carcinogenic Effects of Lead - Epigenetic Effects and
Mixture Interactions - Animal AX5-100
AX5-6.15 Genotoxic/Carcinogenic Effects of Lead - Epigenetic Effects and
Mixture Interactions -Human AX5-101
AX5-6.16 Genotoxic/Carcinogenic Effects of Lead - Epigenetic Effects and
Mixture Interactions - DNA Repair - Human AX5-102
AX5-6.17 Genotoxic/Carcinogenic Effects of Lead - Epigenetic Effects and
Mixture Interactions - DNA Repair - Animal AX5-103
AX5-6.18 Genotoxic/Carcinogenic Effects of Lead - Mitogenesis - Animal AX5-104
AX5-6.19 Genotoxic/Carcinogenic Effects of Lead - Mitogenesis Human
and Animal Cell Culture Studies AX5-107
AX5-6.20 Genotoxic/Carcinogenic Effects of Lead-Mitogenesis Other AX5-110
AX5-7.1 Light Microscopic, Ultrastructural, and Functional Changes AX5-112
AX5-7.2 Lead and Free Radicals AX5-114
AX5-7.3 Chelation with DMSA AX5-117
AX5-7.4 Effect of Chelator Combinations AX5-118
AX5-7.5 Effect of Other Metals on Lead AX5-119
AX5-8.1 Bone Growth in Lead-exposed Animals AX5-122
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List of Tables
(cont'd)
Number
AX5-8.2 Regulation of Bone Cell Function in Animals - Systemic Effects
of Lead AX5-126
AX5-8.3 Bone Cell Cultures Utilized to Test Effects of Lead AX5-129
AX5-8.4 Bone Lead as a Potential Source of Toxicity in Altered
Metabolic Conditions AX5-137
AX5-8.5 Uptake of Lead by Teeth AX5-143
AX5-8.6 Effects of Lead on Enamel and Dentin Formation AX5-144
AX5-8.7 Effects of Lead on Dental Pulp Cells AX5-145
AX5-8.8 Effects of Lead on Teeth-Dental Caries AX5-146
AX5-9.1 Studies on Lead Exposure and Immune Effects in Humans AX5-148
AX5-9.2 Effect of Lead on Antibody Forming Cells (AFC) AX5-151
AX5-9.3 Studies Reporting Lead-Induced Suppression of Delayed Type
Hypersensitivity and Related Responses AX5-152
AX5-9.4 Effect of Lead on Allogeneic and Syngeneic Mixed Lymphocyte
Responses (MLR) AX5-153
AX5-9.5 Effect of Lead on Mitogen-Induced Lymphoid Proliferation AX5-154
AX5.9.6 Pattern of Lead-Induced Macrophage Immunotoxicity AX5-156
AX5-10.1 Hepatic Drug Metabolism AX5-158
AX5-10.2 Biochemical and Molecular Perturbations in Lead-induced Liver
Tissue AX5-163
AX5-10.3 Effect of Lead Exposure on Hepatic Cholesterol Metabolism AX5-166
AX5-10.4 Lead, Oxidative Stress, and Chelation Therapy AX5-167
AX5-10.5 Lead-induced Liver Hyperplasia: Mediators and Molecular
Mechanisms AX5-172
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List of Tables
(cont'd)
Number
AX5-10.6 Effect of Lead Exposure on Liver Heme Synthesis AX5-178
AX5-10.7 Lead and In Vitro Cytotoxicity in Intestinal Cells AX5-181
AX5-10.8 Lead and Intestinal Uptake - Effect on Ultrastructure, Motility,
Transport, and Miscellaneous AX5-182
AX5-10.9 Lead, Calcium, and Vitamin D Interactions, and Intestinal Enzymes AX5-185
AX5-11.1 Lead-Binding Proteins AX5-188
AX6-2.1 Prospective Longitudinal Cohort Studies of Neurocognitive
Ability in Children AX6-3
AX6-2.2 Meta- and Pooled-Analyses of Neurocognitive Ability in Children AX6-9
AX6-2.3 Cross-sectional Studies of Neurocognitive Ability in Children AX6-11
AX6-2.4 Effects of Lead on Academic Achievement in Children AX6-14
AX6-2.5 Effects of Lead on Specific Cognitive Abilities in Children —
Attention/Executive Functions, Learning, and Visual-Spatial Skills AX6-17
AX6-2.6 Effects of Lead on Disturbances in Behavior, Mood, and Social
Conduct in Children AX6-19
AX6-2.7 Effects of Lead on Sensory Acuities in Children AX6-22
AX6-2.8 Effects of Lead on Neuromotor Function in Children AX6-23
AX6-2.9 Effects of Lead on Direct Measures of Brain Anatomical
Development and Activity in Children AX6-24
AX6-2.10 Effects of Lead on Reversibility of Lead-Related Deficits in
Children AX6-26
AX6-3.1 Neurobehavioral Effects Associated with Environmental Lead
Exposure in Adults AX6-29
II-xv
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List of Tables
(cont'd)
Number
AX6-3.2 Symptoms Associated with Occupational Lead Exposure in Adults AX6-32
AX6-3.3 Neurobehavioral Effects Associated with Occupational Lead
Exposure in Adults AX6-35
AX6-3.4 Meta-analyses of Neurobehavioral Effects with Occupational Lead
Exposure in Adults AX6-48
AX6-3.5 Neurophysiological Function and Occupational Lead Exposure
in Adults AX6-50
AX6-3.6 Evoked Potentials and Occupational Lead Exposure in Adults AX6-55
AX6-3.7 Postural Stability, Autonomic Testing, Electroencephalogram,
Hearing Thresholds, and Occupational Lead Exposure in Adults AX6-58
AX6-3.8 Occupational Exposure to Organolead and Inorganic Lead
in Adults AX6-62
AX6-3.9 Other Neurological Outcomes Associated with Lead Exposure
in Adults AX6-65
AX6-4.1 Renal Effects of Lead-General Population AX6-69
AX6-4.2 Renal Effects of Lead-Occupational Population AX6-80
AX6-4.3 Renal Effects of Lead-Patient Population AX6-99
AX6-4.4 Renal Effects of Lead-Mortality AX6-111
AX6-4.5 Renal Effects of Lead-Children AX6-112
AX6-5.1 Cardiovascular Effects of Lead AX6-118
AX6-5.2 Cardiovascular Morbidity Effects of Lead AX6-147
AX6-5.3 Cardiovascular Mortality Effects of Lead AX6-150
AX6-6.1 Placental Transfer of Lead from Mother to Fetus, Human Studies AX6-154
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List of Tables
(cont'd)
Number
AX6-6.2 Lead Exposure and Male Reproduction: Semen Quality,
Human Studies AX6-156
AX6-6.3 Lead Exposure and Male Reproduction: Time to Pregnancy,
Human Studies AX6-158
AX6-6.4 Lead Exposure and Male Reproduction: Reproductive History,
Human Studies AX6-160
AX6-7.1 Recent Studies of Lead Exposure and Genotoxicity AX6-162
AX6-7.2 Key Occupational Studies of Lead Exposure and Cancer AX6-164
AX6-7.3 Key Studies of Lead Exposure and Cancer in the General Population AX6-170
AX6-7.4 Other Studies of Lead Exposure and Cancer AX6-171
AX6-8.1 Effects of Lead on Immune Function in Children AX6-182
AX6-8.2 Effects of Lead on Immune Function in Adults AX6-186
AX6-9.1 Effects of Lead on Biochemical Effects in Children AX6-196
AX6-9.2 Effects of Lead on Biochemical Effects in Adults AX6-198
AX6-9.3 Effects of Lead on Hematopoietic System in Children AX6-206
AX6-9.4 Effects of Lead on Hematopoietic System in Adults AX6-209
AX6-9.5 Effects of Lead on the Endocrine System in Children AX6-216
AX6-9.6 Effects of Lead on the Endocrine System in Adults AX6-218
AX6-9.7 Effects of Lead on the Hepatic System in Children and Adults AX6-226
AX6-9.8 Effects of Lead on the Gastrointestinal System AX6-228
AX6-9.9 Effects of Lead on the Respiratory Tract in Adults AX6-230
AX6-9.10 Effects of Lead on Bone and Teeth in Children and Adults AX6-231
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List of Tables
(cont'd)
Number
AX6-9.11 Effects of Lead on Ocular Health in Children and Adults AX6-234
AX6-10.1 Average Estimated Slopes for Linear and Log-linear Models in the
Presence of Heteroscedasticity AX6-237
AX8-1.1.1 Relative Standard Deviation (RSD) for Lead Isotope Ratios on
Selected Mass Spectrometers AX8-2
AX8-1.1.2 National Institute of Standards and Technology Lead SRMs AX8-10
AX8-1.1.3 Characteristics for Direct Speciation Techniques AX8-17
AX8-1.1.4 Affinity Constants for Lead AX8-19
AX8-1.3.1 Tissue Lead Levels in Birds Causing Effects AX8-44
AX8-1.4.1 Plant Toxicity Data Used to Develop the Eco-SSL AX8-62
AX8-1.4.2 Plant Toxicity Data Not Used to Develop the Eco-SSL AX8-63
AX8-1.4.3 Avian Toxicity Data Used to Develop the Eco-SSL AX8-66
AX8-1.4.4 Mammalian Toxicity Data Used to Develop the Eco-SSL AX8-72
AX8-1.4.5 Invertebrate Toxicity Data Used to Develop the Eco-SSL AX8-82
AX8-1.4.6 Invertebrate Toxicity Data Not Used to Develop the Eco-SSL AX8-84
AX8-2.1.1 Common Analytical Methods for Measuring Lead in Water,
Sediment, and Tissue AX8-107
AX8-2.1.2 Development of Current Acute Freshwater Criteria for Lead AX8-109
AX8-2.1.3 Recommended Sediment Quality Guidelines for Lead AX8-114
AX8-2.2.1 NAWQA Land Use Categories and Natural/Ambient
Classification AX8-124
AX8-2.2.2 Summary Statistics of Ambient and Natural Levels of Dissolved
Lead in Surface Water AX8-125
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List of Tables
(cont'd)
Number
AX8-2.2.3 Summary Statistics of Ambient and Natural Levels of Total Lead
in<63 |im Bulk Sediment AX8-126
AX8-2.2.4 Summary Statistics of Ambient and Natural Levels of Lead in
Whole Organism and Liver Tissues AX8-135
AX8-2.2.5 Comparison of NCBP and NAWQA Ambient Lead Levels in
Whole Organism Tissues AX8-137
AX8-2.3.1 Bioconcentration Factors for Aquatic Plants AX8-149
AX8-2.3.2 Bioconcentration Factors for Aquatic Invertebrates AX8-149
AX8-2.4.1 Effects of Lead to Freshwater and Marine Invertebrates AX8-183
AX8-2.4.2 Effects of Pb to Freshwater and Marine Fish AX8-188
AX8-2.4.3 Nonlethal Effects in Amphibians AX8-190
AX8-2.5.1 Ecological Attributed Studies by Maret et al. (2003) in the Coeur
d'Alene Watershed AX8-194
AX8-2.5.2 Essential Ecological Attributes for Natural Aquatic Ecosystems
Affected by Lead AX8-197
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List of Figures
Number Page
AX8-1.1.1 Relationship of bioaccessibility (low, medium, high) versus speciation
as shown with scanning electron micrographs of various Pb-bearing
materials AX8-5
AX8-1.1.2 Variation of bioavailability with particle size AX8-7
AX8-1.1.3 Illustration of particle lability and bioavailability at two different sites
with similar total Pb concentrations andPb forms AX8-8
AX8-1.1.4 Scanning electron micrograph of a large native Pb particle showing
outer ring of highly bioavailable Pb-chloride andPb-oxide AX8-8
AX8-1.1.5 Bulk lead versus single species modality AX8-12
AX8-1.4.1 Avian reproduction and growth toxicity data considered in development
oftheEco-SSL AX8-69
AX8-1.4.2 Mammalian reproduction and growth toxicity data considered in
development of the Eco-SSL AX8-81
AX8-2.2.1 Distribution of aqueous lead species as a function of pH based on
a concentration of 1 |igPb/L AX8-119
AX8-2.2.2 Lead speciation versus chloride content (Fernando, 1995) AX8-120
AX8-2.2.3 Spatial distribution of natural and ambient surface water/sediment
sites (Surface water: natural N = 430, ambient N = 3445; Sediment:
natural N = 258, ambient N= 1466) AX8-127
AX8-2.2.4 Spatial distribution of natural and ambient liver tissue sample sites
(Natural N = 83, Ambient N = 559) AX8-128
AX8-2.2.5 Spatial distribution of natural and ambient whole organism tissue
sample sites (Natural N = 93, Ambient N = 332) AX8-129
AX8-2.2.6 Frequency distribution of ambient and natural levels of surface water
dissolved lead (|ig/L) AX8-130
AX8-2.2.7 Spatial distribution of dissolved lead in surface water (N = 3445) AX8-131
II-xx
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List of Figures
(cont'd)
Number
AX8-2.2.8 Frequency distribution of ambient and natural levels of bulk sediment
<63 |im total Pb (|ig/g) AX8-133
AX8-2.2.9 Spatial distribution of total lead in bulk sediment <63 urn (N = 1466) AX8-134
AX8-2.2.10 Frequency distribution of ambient and natural levels of lead in liver
tissue (ng/g dry weight) AX8-136
AX8-2.2.11 Frequency distribution of ambient and natural levels of lead in whole
organism tissue (|ig/g dry weight) AX8-136
AX8-2.2.12 Spatial distribution of lead in liver tissues (N= 559) AX8-139
AX8-2.2.13 Spatial distribution of lead in whole organism tissues (N = 332) AX8-140
AX8-2.2.14 Lead cycle in an aquatic ecosystem AX8-141
-------
Authors, Contributors, and Reviewers
CHAPTER 5 ANNEX (TOXICOLOGICAL EFFECTS OF LEAD IN HUMANS
AND LABORATORY ANIMALS)
Chapter Managers/Editors
Dr. Anuradha Mudipalli—National Center for Environmental Assessment (B243-01),
U.S. Environmental Protection Agency, Research Triangle Park, NC 27711
Dr. Srikanth Nadadur—National Center for Environmental Assessment (B243-01),
U.S. Environmental Protection Agency, Research Triangle Park, NC 27711
Dr. Lori White—National Center for Environmental Assessment (B243-01),
U.S. Environmental Protection Agency, Research Triangle Park, NC 27711
Principal Authors
Dr. Anuradha Mudipalli—National Center for Environmental Assessment (B243-01),
U.S. Environmental Protection Agency, Research Triangle Park, NC 27711 (Sections 5-2, 5-10)
Dr. Stephen Lasley—Dept. of Biomedical and Therapeutic Sciences, Univ. of Illinois College of
Medicine, PO. Box 1649, Peoria, IL 61656 (Section 5.3.1)
Dr. Lori White—National Center for Environmental Assessment (B243-01),
U.S. Environmental Protection Agency, Research Triangle Park, NC 27711 (Section 5.3.1)
Dr. John Rosen—Division of Environmental Sciences, The Children's Hospital at Montefiore,
The Albert Einstein College of Medicine, 111 E. 210th St. Room 401, Bronx, NY 10467
(Section 5.3.2)
Dr. Gary Diamond—Syracuse Research Corporation, 8191 Cedar Street, Akron, NY 14001
(Section 5.4)
Dr. N.D. Vaziri—Division of Nephrology and Hypertension, University of California - Irvine
Medical Center, 101, The City Drive, Bldg 53, Room #125. Orange, CA 92868 (Section 5.5)
Dr. John Pierce Wise, Sr.—Ph.D., Maine Center for Toxicology and Environmental Health,
Department of Applied Medical Sciences, 96 Falmouth Street, PO Box 9300, Portland, ME
04104-9300 (Section 5.6)
Dr. Harvey C. Gonick—David Geffen School of Medicine, University of California at
Los Angeles, CA (201 Tavistock Ave, Los Angeles, CA 90049) (Sections 5.7, 5.11)
-------
Authors, Contributors, and Reviewers
(cont'd)
Principal Authors
(cont'd)
Dr. Gene E. Watson— University of Rochester Medical Center, Box 705, Rochester, NY 14642
(Section 5.8)
Dr. Rodney Dietert— Institute for Comparative and Environmental Toxicology, College of
Veterinary Medicine, Cornell University, Ithaca, NY 14853 (Section 5.9)
Contributors and Reviewers
Dr. Lester D. Grant—National Center for Environmental Assessment (B243-01),
U.S. Environmental Protection Agency, Research Triangle Park, NC 27711
Dr. Paul Reinhart—National Center for Environmental Assessment (B243-01),
U.S. Environmental Protection Agency, Research Triangle Park, NC 27711
Dr. David A. Lawrence—Dept of Environmental and Clinical Immunology, Empire State Plaza
P.O. Box 509, Albany, NY 12201
Dr Michael J. McCabe, Jr.—Dept of Environmental Medicine, University of Rochester,
575 Elmwood Avenue, Rochester, NY 14642
Dr. Theodore I. Lidsky— New York State Institute for Basic Research, 1050 Forest RD,
Staten Island, NY 10314
Dr. Mark H. Follansbee- Syracuse Research Corporation, 8191 Cedar Street, Akron, NY 14001
Ms. Beth Hassett-Sipple—Office of Air Quality Planning and Standards, U.S. Environmental
Protection Agency, Research Triangle Park, NC 27711
Dr. Zachary Pekar—Office of Air Quality Planning and Standards, U.S. Environmental
Protection Agency, Research Triangle Park, NC 27711
-------
Authors, Contributors, and Reviewers
(cont'd)
CHAPTER 6 ANNEX (EPIDEMIOLOGICAL STUDIES OF AMBIENT LEAD
EXPOSURE EFFECTS)
Chapter Managers/Editors
Dr. Jee Young Kim—National Center for Environmental Assessment (B243-01),
U.S. Environmental Protection Agency, Research Triangle Park, NC 27711
Dr. Dennis Kotchmar—National Center for Environmental Assessment (B243-01),
U.S. Environmental Protection Agency, Research Triangle Park, NC 27711
Dr. David Svendsgaard—National Center for Environmental Assessment (B243-01),
U.S. Environmental Protection Agency, Research Triangle Park, NC 27711
Principal Authors
Dr. David Bellinger—Children's Hospital, Farley Basement, Box 127, 300 Longwood Avenue,
Boston, MA 02115 (Section 6.10)
Dr. Margit Bleecker—Center for Occupational and Environmental Neurology, 3901
Greenspring Ave., Suite 101, Baltimore, MD 21211 (Section 6.3)
Dr. Gary Diamond— Syracuse Research Corporation, 8191 Cedar Street, Akron, NY 14001
(Section 6.8, 6.9)
Dr. Kim Dietrich—University of Cincinnati College of Medicine, 3223 Eden Avenue,
Kettering Laboratory, Room G31, Cincinnati, OH 45267 (Section 6.2)
Dr. Pam Factor-Litvak—Columbia University Mailman School of Public Health, 722 West
168th Street, Room 1614, New York, NY 10032 (Section 6.6)
Dr. Vic Hasselblad—Duke University Medical Center, Durham, NC 27713 (Section 6.10)
Dr. Steve Rothenberg—CINVESTAV-IPN, Merida, Yucatan, Mexico & National Institute of
Public Health, Cuernavaca, Morelos, Mexico (Section 6.5)
Dr. Neal Simonsen—Louisiana State University Health Sciences Center, School of Public
Health & Stanley S Scott Cancer Center, 1600 Canal Street, Suite 800, New Orleans, LA 70112
(Section 6.7)
II-xxiv
-------
Authors, Contributors, and Reviewers
(cont'd)
Principal Authors
(cont'd)
Dr. Kyle Steenland—Rollins School of Public Health, Emory University, 1518 Clifton Road,
Room 268, Atlanta, GA 30322 (Section 6.7)
Dr. Virginia Weaver—Johns Hopkins Bloomberg School of Public Health, 615 North Wolfe
Street, Room 7041, Baltimore, MD 21205 (Section 6.4)
Dr. Harvey C. Gonick— David Geffen School of Medicine, University of California at Los
Angeles, CA (201 Tavistock Ave, Los Angeles, CA 90049) (Sections 6.4)
Contributors and Reviewers
Dr. J. Michael Davis—National Center for Environmental Assessment (B243-01),
U.S. Environmental Protection Agency, Research Triangle Park, NC 27711
Dr. Lester D. Grant—National Center for Environmental Assessment (B243-01),
U.S. Environmental Protection Agency, Research Triangle Park, NC 27711
Dr. Kaz Ito—School of Medicine, New York University, Tuxedo, NY 10917
Dr. Kathryn Mahaffey—Office of Prevention, Pesticides and Toxic Substances,
U.S. Environmental Protection Agency, Washington, DC 20460
Dr. Karen Martin—Office of Air Quality Planning and Standards, U.S. Environmental Protection
Agency, Research Triangle Park, NC 27711
Ms. Beth Hassett-Sipple—Office of Air Quality Planning and Standards, U.S. Environmental
Protection Agency, Research Triangle Park, NC 27711
Dr. Zachary Pekar—Office of Air Quality Planning and Standards, U.S. Environmental
Protection Agency, Research Triangle Park, NC 27711
II-XXV
-------
Authors, Contributors, and Reviewers
(cont'd)
CHAPTER 8 - ENVIRONMENTAL EFFECTS OF LEAD
Chapter Manager/Editor
Dr. Timothy Lewis—National Center for Environmental Assessment (B243-01),
U.S. Environmental Protection Agency, Research Triangle Park, NC 27711
Principal Authors
Dr. Ruth Hull—Cantox Environmental Inc., 1900 Minnesota Court, Suite 130, Mississauga, Ontario,
L5N 3C9 Canada (Section 8.1)
Dr. James Kaste—Department of Earth Sciences, Dartmouth College, 352 Main Street, Hanover,
NH 03755 (Section 8.1)
Dr. John Drexler—Department of Geological Sciences, University of Colorado, 1216 Gillespie
Drive, Boulder, CO 80305 (Section 8.1)
Dr. Chris Johnson—Department of Civil and Environmental Engineering, Syracuse University,
365 Link Hall, Syracuse, NY 13244 (Section 8.1)
Dr. William Stubblefield—Parametrix, Inc. 33972 Texas St. SW, Albany, OR 97321 (Section 8.2)
Dr. Dwayne Moore—Cantox Environmental, Inc., 1550A Laperriere Avenue, Suite 103, Ottawa,
Ontario, K1Z 7T2 Canada (Section 8.2)
Dr. David Mayfield—Parametrix, Inc., 411 108th Ave NE, Suite 1800, Bellevue, WA 98004 (Section
8.2)
Dr. Barbara Southworth—Menzie-Cura & Associates, Inc., 8 Winchester Place, Suite 202,
Winchester, MA 01890 (Section 8.3)
Dr. Katherine Von Stackleberg—Menzie-Cura & Associates, Inc., 8 Winchester Place, Suite 202,
Winchester, MA 01890 (Section 8.3)
II-xxvi
-------
Authors, Contributors, and Reviewers
(cont'd)
Contributors and Reviewers
Dr. Jerome Nriagu—Department of Environmental Health Sciences, 109 South Observatory,
University of Michigan, Ann Arbor, MI 48109
Dr. Judith Weis—Department of Biology, Rutgers University, Newark, NJ 07102
Dr. Sharon Harper—National Exposure Research Laboratory (D205-05), U.S. Environmental
Protection Agency, Research Triangle Park, NC 27711
Dr. Karen Bradham—National Research Exposure Laboratory (D205-05), U.S. Environmental
Protection Agency, Research Triangle Park, NC 27711
Dr. Ginger Tennant—Office of Air Quality Planning and Standards, U.S. Environmental
Protection Agency, Research Triangle Park, NC 27711
Ms. Gail Lacey—Office of Air Quality Planning and Standards, U.S. Environmental Protection
Agency, Research Triangle Park, NC 27711
[Note: Any inadvertently omitted names of authors/reviewers will be inserted in the final version
of this LeadAQCD, as will more complete addresses for all authors/reviewers.]
II-xxvii
-------
U.S. Environmental Protection Agency Project Team
for Development of Air Quality Criteria for Lead
Executive Direction
Dr. Lester D. Grant (Director)—National Center for Environmental Assessment-RTF Division,
(B243-01), U.S. Environmental Protection Agency, Research Triangle Park, NC 27711
Scientific Staff
Dr. Lori White (Lead Team Leader)—National Center for Environmental Assessment
(B243-01), U.S. Environmental Protection Agency, Research Triangle Park, NC 27711
Dr. James S. Brown—National Center for Environmental Assessment (B243-01),
U.S. Environmental Protection Agency, Research Triangle Park, NC 27711
Dr. Robert Elias—National Center for Environmental Assessment (B243-01),
U.S. Environmental Protection Agency, Research Triangle Park, NC 27711 (Retired)
Dr. Brooke Hemming—National Center for Environmental Assessment (B243-01), U.S.
Environmental Protection Agency, Research Triangle Park, NC 27711
Dr. Jee Young Kim—National Center for Environmental Assessment (B243-01),
U.S. Environmental Protection Agency, Research Triangle Park, NC 27711
Dr. Dennis Kotchmar—National Center for Environmental Assessment (B243-01),
U.S. Environmental Protection Agency, Research Triangle Park, NC 27711
Dr. Timothy Lewis—National Center for Environmental Assessment (B243-01),
U.S. Environmental Protection Agency, Research Triangle Park, NC 27711
Dr. Anuradha Muldipalli—National Center for Environmental Assessment (B243-01),
U.S. Environmental Protection Agency, Research Triangle Park, NC 27711
Dr. Srikanth Nadadur—National Center for Environmental Assessment (B243-01),
U.S. Environmental Protection Agency, Research Triangle Park, NC 27711
Dr. Paul Reinhart—National Center for Environmental Assessment (B243-01),
U.S. Environmental Protection Agency, Research Triangle Park, NC 27711
Dr. Mary Ross— National Center for Environmental Assessment (B243-01),
U.S. Environmental Protection Agency, Research Triangle Park, NC 27711
Dr. David Svendsgaard—National Center for Environmental Assessment (B243-01),
U.S. Environmental Protection Agency, Research Triangle Park, NC 27711
-------
U.S. Environmental Protection Agency Project Team
for Development of Air Quality Criteria for Lead
(cont'd)
Technical Support Staff
Mr. Douglas B. Fennell—Technical Information Specialist, National Center for Environmental
Assessment (B243-01), U.S. Environmental Protection Agency, Research Triangle Park, NC
27711
Ms. Emily R. Lee—Management Analyst, National Center for Environmental Assessment
(B243-01), U.S. Environmental Protection Agency, Research Triangle Park, NC 27711
Ms. Diane H. Ray—Program Specialist, National Center for Environmental Assessment
(B243-01), U.S. Environmental Protection Agency, Research Triangle Park, NC 27711
Ms. Donna Wicker—Administrative Officer, National Center for Environmental Assessment
(B243-01), U.S. Environmental Protection Agency, Research Triangle Park, NC 27711 (Retired)
Mr. Richard Wilson—Clerk, National Center for Environmental Assessment (B243-01),
U.S. Environmental Protection Agency, Research Triangle Park, NC 27711
Document Production Staff
Ms. Carolyn T. Perry—Task Order Manager, Computer Sciences Corporation, 2803 Slater Road,
Suite 220, Morrisville, NC 27560
Mr. John A. Bennett—Technical Information Specialist, Library Associates of Maryland,
11820 Parklawn Drive, Suite 400, Rockville, MD 20852
Mr. William Ellis—Records Management Technician, InfoPro, Inc., 8200 Greensboro Drive,
Suite 1450, McLean, VA 22102
Ms. Sandra L. Hughey—Technical Information Specialist, Library Associates of Maryland,
11820 Parklawn Drive, Suite 400, Rockville, MD 20852
Dr. Barbara Liljequist—Technical Editor, Computer Sciences Corporation, 2803 Slater Road,
Suite 220, Morrisville, NC 27560
Ms. Michelle Partridge-Doerr—Publications/Graphics Specialist, TEK Systems, 1201 Edwards
Mill Road, Suite 201, Raleigh, NC 27607
Mr. Carlton Witherspoon—Graphic Artist, Computer Sciences Corporation, 2803 Slater Road,
Suite 220, Morrisville, NC 27560
II-xxix
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U.S. Environmental Protection Agency
Science Advisory Board (SAB) Staff Office
Clean Air Scientific Advisory Committee (CASAC)
Chair
Dr. Rogene Henderson*—Scientist Emeritus, Lovelace Respiratory Research Institute,
Albuquerque, NM
Members
Dr. Joshua Cohen—Faculty, Center for the Evaluation of Value and Risk, Institute for Clinical
Research and Health Policy Studies, Tufts New England Medical Center, Boston, MA
Dr. Deborah Cory-Slechta—Director, University of Medicine and Dentistry of New Jersey and
Rutgers State University, Piscataway, NJ
Dr. Ellis Cowling*—University Distinguished Professor-at-Large, North Carolina State
University, Colleges of Natural Resources and Agriculture and Life Sciences, North Carolina
State University, Raleigh, NC
Dr. James D. Crapo [M.D.]*—Professor, Department of Medicine, National Jewish Medical
and Research Center, Denver, CO
Dr. Bruce Fowler—Assistant Director for Science, Division of Toxicology and Environmental
Medicine, Office of the Director, Agency for Toxic Substances and Disease Registry, U.S.
Centers for Disease Control and Prevention (ATSDR/CDC), Chamblee, GA
Dr. Andrew Friedland—Professor and Chair, Environmental Studies Program, Dartmouth
College, Hanover, NH
Dr. Robert Goyer [M.D.]—Emeritus Professor of Pathology, Faculty of Medicine, University
of Western Ontario (Canada), Chapel Hill, NC
Mr. Sean Hays—President, Summit Toxicology, Allenspark, CO
Dr. Bruce Lanphear [M.D.]—Sloan Professor of Children's Environmental Health, and the
Director of the Cincinnati Children's Environmental Health Center at Cincinnati Children's
Hospital Medical Center and the University of Cincinnati, Cincinnati, OH
Dr. Samuel Luoma—Senior Research Hydrologist, U.S. Geological Survey (USGS),
Menlo Park, CA
II-XXX
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U.S. Environmental Protection Agency
Science Advisory Board (SAB) Staff Office
Clean Air Scientific Advisory Committee (CASAC)
(cont'd)
Members
(cont'd)
Dr. Frederick J. Miller*—Consultant, Gary, NC
Dr. Paul Mushak—Principal, PB Associates, and Visiting Professor, Albert Einstein College of
Medicine (New York, NY), Durham, NC
Dr. Michael Newman—Professor of Marine Science, School of Marine Sciences, Virginia
Institute of Marine Science, College of William & Mary, Gloucester Point, VA
Mr. Richard L. Poirot*—Environmental Analyst, Air Pollution Control Division, Department
of Environmental Conservation, Vermont Agency of Natural Resources, Waterbury, VT
Dr. Michael Rabinowitz—Geochemist, Marine Biological Laboratory, Woods Hole, MA
Dr. Joel Schwartz—Professor, Environmental Health, Harvard University School of Public
Health, Boston, MA
Dr. Frank Speizer [M.D.]*—Edward Kass Professor of Medicine, Channing Laboratory,
Harvard Medical School, Boston, MA
Dr. Ian von Lindern—Senior Scientist, TerraGraphics Environmental Engineering, Inc.,
Moscow, ID
Dr. Barbara Zielinska*—Research Professor, Division of Atmospheric Science, Desert
Research Institute, Reno, NV
SCIENCE ADVISORY BOARD STAFF
Mr. Fred Butterfield—CASAC Designated Federal Officer, 1200 Pennsylvania Avenue, N.W.,
Washington, DC, 20460, Phone: 202-343-9994, Fax: 202-233-0643 (butterfield.fred@epa.gov)
(Physical/Courier/FedEx Address: Fred A. Butterfield, III, EPA Science Advisory Board Staff
Office (Mail Code 1400F), Woodies Building, 1025 F Street, N.W., Room 3604, Washington,
DC 20004, Telephone: 202-343-9994)
*Members of the statutory Clean Air Scientific Advisory Committee (CASAC) appointed by the
EPA Administrator
II-xxxi
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Abbreviations and Acronyms
aFGF
AA
AAL
AAS
ABA
ACBP
ACE
ACh
AChE
ACR
AD
ADC
ADP
AE
AEA
AFC
2-AG
A horizon
AHR
AI
ALA
ALAD
ALAS
ALAU
ALD
ALS
ALT
ALWT
AMEM
AMP
ANCOVA
ANF
Angll
ANOVA
a-fibroblast growth factor
arachidonic acid
active avoidance learning
atomic absorption spectroscopy
p-aminoisobutyric acid
Achenbach Child Behavior Profile
angiotensin converting enzyme
acetylcholine
acetylcholine esterase
acute-chronic ratio
adult
analog digital converter
adenosine dinucleotide phosphate
anion exchange
TV-arachi dony 1 ethanol amine
antibody forming cells
2-arachidonylglycerol
uppermost layer of soil (litter and humus)
aryl hydrocarbon receptor
angiotensin I
5-aminolevulinic acid
5-aminolevulinic acid dehydratase
aminolevulinic acid synthetase
urinary 5-aminolevulinic acid
aldosterone
amyotrophic lateral sclerosis
alanine aminotransferase
albumin weight
Alpha Minimal Essential Medium
adenosine monophosphate
analysis of covariance
atrial natriuretic factor
angiotensin II
analysis of variance
II-xxxii
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ANP
AP
AP-1
ApoE
AQCD
Arg
AS52
ASGP-R
AST
ASV
3-AT
ATP
ATP1A2
ATPase
ATSDR
AVCD
AVS
AWQC
P
PFGF
17P-HS
3P-HSD
17P-HSDH
6p-OH-cortisol
B
BAEP
BAER
BAF
Bcell
BCFs
BCS
BDNF
BDWT
BEI
BFU-E
atrial natriuretic peptide
alkaline phosphatase
activated protein-1
apolipoprotein E
Air Quality Criteria Document
arginine
cells derived from the CHO cell line
aceyl glycoprotein receptor
aspartate aminotransferase
anode stripping voltammetry
3-aminotriazole; 3-amino triazide
adenosine triphosphate
sodium-potassium adenosine triphosphase a2
adenosine triphosphatase
Agency for Toxic Substances and Disease Research
atrioventricular conduction deficit
acid volatile sulfide
ambient water quality criteria
beta-coefficient; slope of an equation
P-fibroblast growth factor
17p-hydroxysteriod
3p-hydroxysteriod dehydrogenase
17p-hydroxysteriod dehydrogenase
6-p-hydroxycortisol
both
brainstem auditory-evoked potentials
brainstem auditory-evoked responses
bioaccumulation factor
B lymphocyte
bioconcentration factors
bovine calf serum
brain derived neurotrophic factor
body weight changes
biological exposure index
blood erythroid progenitor
-------
BLL
BLM
BM
BMI
BNDF
BOTMP
BP
BPb
BSA
BSI
BTQ
BUN
bw, b. wt, BW
C3H10T/12
C3, C4
CA
CAS
45Ca
Ca-ATP
Ca-ATPase
CaCO3
CaEDTA
CAL
CaM
Ca-Mg-ATPase
cAMP
CaNa2 EDTA
CANTAB
CAT
CBCL
CBCL-T
CBL
CBLI
CCB
CCD
blood lead level
biotic ligand model
basement membrane
body mass index
brain-derived neurotrophic growth factor
Bruinicks-Oseretsky Test of Motor Proficiency
blood pressure
blood lead concentration
bovine serum albumin
Brief Symptom Inventory
Boston Teacher Questionnaire
blood urea nitrogen
body weight
mouse embryo cell line
complement proteins
chromosome aberration
cornu ammonis 3 region of hippocampus
calcium-45 radionuclide
calcium-dependent adenosine triphosphate
calcium-dependent adenosine triphosphatase
calcium carbonate
calcium disodium ethylenediaminetetraacetic acid
calcitonin
calmodulin
calcium-magnesium-dependent adenosine triphosphatase
cyclic adenosinemonophosphate
calcium disodium ethylenediaminetetraacetic acid
Cambridge Neuropsychological Testing Automated Battery
catalase; Cognitive Abilities Test
Achenbach Child Behavior Checklist
Total Behavior Problem Score
cumulative blood lead
cumulative blood lead index
cytochalasin B
charge-coupled device
II-xxxiv
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CCE
CCL
CCS
109Cd
CdU
CEC
CESD, CES-D
GFAP
CFU-E
CFU-GEMM
CFU-GM
cGMP
ChAT
CHD
CHO
CI
CLE-SV
CLRTAP
CLS
CMC
CMI
CNS
COH
ConA
COR
CoTx
COX-2
CP
CPT
cr
CRAC
CREB
CRF
CRT
Coordination Center for Effects
carbon tetrachloride
cosmic calf serum
coefficient of component variance of respiratory sinus arrhythmia
cadmium-109 radionuclide
urinary cadmium
cation exchange capacity
Center for Epidemiologic Studies Depression (scale)
glial fibrillary acidic protein
colony forming unit blood-erythroid progenitor (cell count)
colony forming unit blood-pluripotent progenitor (cell count)
blood granulocyte/macrophage progenitor (cell count)
cyclic guanosine-3',5'-monophosphate
choline acetyltransferase
coronary heart disease
Chinese hamster ovary cell line
confidence interval
competitive ligand-exchange/stripping voltammetry
Convention on Long-Range Transboundary of Air Pollution
Cincinnati Lead Study
criterion maximum concentration
cell-mediated immunity
central nervous system
cation-osmotic hemolysis
concanavalin A
cortisol
cotreatment
cyclooxygenase-2
coproporphryn
current perception threshold
creatinine
calcium release activated calcium reflux
cyclic AMP-response element binding protein
chronic renal failure
chronic renal insufficiency
II-XXXV
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CSF
CuZn-SOD
CV
CVLT
CVR.R
CYP
CYpSall
D
DA
dbcAMP
DCV
DEAE
DET
DEYO
DPS
dfs
DG
DOT
DL
DMEM
DMEM/F12
DMFS
DMPS
DMSA
DMT
DMTU
DNA
DO
DOC
DOM
DOPAc
DPASV
dp/dt
DPPD
DR
cerebrospinal fluid
copper and zinc-dependent superoxide dismutase
conduction velocity
California Verbal Learning Test
coefficient of variation of the R-R interval
cytochrome (e.g., CYP1A, CYP-2A6, CYP3A4, CYP450)
cytochrome P450 Sal 1
D-statistic
dopamine; dopaminergic
dibutyryl cyclic adenosine-3',5'-monophosphate
distribution of conduction velocities
diethylaminoethyl (chromatography)
diffusive equilibrium thin films
death of young
decayed or filled surfaces, permanent teeth
covariate-adjusted number of caries
dentate gyrus
diffusive gradient thin films
DL-statistic
Dulbecco's Minimal Essential Medium
Dulbecco's Minimal Essential Medium/Ham's F12
decayed, missing, or filled surfaces, permanent teeth
2,3-dimercaptopropane 1-sulfonate
2,3-dimercaptosuccinic acid
Donnan membrane technique
dimethylthiourea
deoxyribonucleic acid
distraction osteogenesis
dissolved organic carbon
dissolved organic carbon
3,4-dihydroxyphenylacetic acid
differential pulse anodic stripping voltammetry
rate of left ventricular isovolumetric pressure
TV-TV-diphenyl-p-phynylene-diamine
drinking water
II-xxxvi
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DSA
DTC
DTK
DTPA
DTT
dw
E
E2
EBE
EBV
EC
ECso
eCB
ECG
Eco-SSL
EDS
EDTA
EEDQ
EEG
EG
EGF
EGG
EGPN
EKG
electro
EM/CM
EMEM
eNOS
EP
EPA
Epi
EPMA
EPO
EPSC
delayed spatial alternation
diethyl dithiocarbomate complex
delayed type hypersensitivity
diethylenetriaminepentaacetic acid
dithiothreitol
dry weight
embryonic day
estradiol
early biological effect
Epstein-Barr virus
European Community
effect concentration for 50% of test population
endocannabinoid
electrocardiogram
ecological soil screening level
energy dispersive spectrometers
ethylenediaminetetraacetic acid
7V-ethoxycarbonyl-2-ethoxy-1,2-dihydroquinone
el ectroencephal ogram
egg
epidermal growth factor
effects on eggs
egg production
electrocardiogram
electrophysiological stimulation
experimental medium-to-control medium (ratio)
Eagle's Minimal Essential Medium
endothelial nitric oxide synthase
erythrocyte protoporphyrin
U.S. Environmental Protection Agency
epinephrine
electron probe microanalysis
erythropoietin
excitatory postsynaptic currents
II-xxxvii
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EPT
ERG
ERL
ERM
EROD
ESCA
ESRD
EST
ESTH
ET
ETOH
EXAFS
EXANES
F
F344
FAV
FBS
FCS
FCV
FD
FEF
FEP
FERT
FEVi
FGF
FI
FIAM
FMLP
fMRI
FR
FSH
FT3
FT4
FTES
macroinvertebrates from the Ephemeroptera (mayflies),
Plecoptera (stoneflies), and Trichoptera (caddisflies) group
electroretinogram; electroretinographic
effects range - low
effects range - median
ethoxyresorufin-0-deethylase
electron spectroscopy for chemical analysis
end-stage renal disease
estradiol
eggshell thinning
endothelein; essential tremor
ethyl alcohol
extended X-ray absorption fine structure
extended X-ray absorption near edge spectroscopy
F-statistic
Fischer 344 (rat)
final acute value
fetal bovine serum
fetal calf serum
final chronic value
food
forced expiratory flow
free erythrocyte protoporphyrin
fertility
forced expiratory volume in one second
fibroblast growth factor (e.g., PFGF, aFGF)
fixed interval (operant conditioning)
free ion activity model
N-formy 1 -L-methi ony 1 -L-l eucy 1 -L-pheny 1 al anine
functional magnetic resonance imaging
fixed-ratio operant conditioning
follicle stimulating hormone
free triiodothyronine
free thyroxine
free testosterone
II-xxxviii
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FTII
FTPLM
FURA-2
FVC
Y-GT
G
GABA
GAG
G12 CHV79
GCI
GD
GDP
GEE
GFAAS
GFR
GGT
GH
GI
GIME-VIP
GIS
GLU
GMAV
GMCV
GMP
GMPH
GnRH
GOT
GP
G6PD, G6PDH
GPEI
gp91phox
GPT
GPx
GRO
Fagan Test of Infant Intelligence
flow-through permeation liquid membranes
l-[6-amino-2-(5-carboxy-2-oxazolyl)-5-benzofuranyloxy]-2-(2-
amino-5-methylphenoxy) ethane-7V,7V,jV',jV'-tetraacetic acid
forced vital capacity
y-glutamyl transferase
gestational day
gamma aminobutyric acid
glycosaminoglycan
cells derived from the V79 cell line
General Cognitive Index
gestational day
guanosine diphosphate
generalized estimating equations
graphite furnace atomic absorption spectroscopy
glomerular filtration rate
y-glutamyl transf erase
growth hormone
gastrointestinal
gel integrated microelectrodes combined with voltammetric
in situ profiling
geographic information system
glutamate
genus mean acute value
genus mean chronic value
guanosine monophosphate
general morphology
gonadotropin releasing hormone
aspartate aminotransferase
gross productivity
glucose-6-phosphate dehydrogenase
glutathione S-transferase P enhancer element
NAD(P)H oxidase
glutamic-pyruvic transaminase
glutathione peroxidase
growth
II-xxxix
-------
GRP78
GSD
GSH
GSIM
GSSG
GST
GSTP
GTP
GV
H+
3H
HA
Hb
HBEF
HBSS
HCG; hCG
Hct
HDL
HEP
HET
HFPLM
Hgb
HGF
HH
H-H
HHANES
H-L
HLA
H-MEM
HMP
HNO3
H2O2
HOME
HOSTE
HPLC
glucose-regulated protein 78
geometric standard deviation
reduced glutathione
gill surface interaction model
glutathione disulfide
glutathione-S-transferase
placental glutathione transferase
guanosine triphosphate
gavage
acidity
hydrogen-3 radionuclide (tritium)
humic acid; hydroxyapatite
hemoglobin
Hubbard Brook Experimenatl Forest
Hank's Balanced Salt Solution
human chorionic gonadotropin
hematocrit
high-density lipoprotein (cholesterol)
habitat evaluation procedure
Binghamton heterogeneous stock
hollow fiber permeation liquid membranes
hemoglobin
hepatocyte growth factor
hydroxylamine hydrochloride
high-high
Hispanic Health and Nutrition Examination Survey
high-low
human leukocyte antigen
minimum essential medium/nutrient mixture-F12-Ham
hexose monophosphate shunt pathway
nitric acid
hydrogen peroxide
Home Observation for Measurement of Environment
human osteosarcoma cells
high-pressure liquid chromatography
II-xl
-------
H3PO4
HPRT
HR
HSI
H2SO4
HSPG
Ht
HTC
hTERT
HTN
IBL
IBL x WRAT-R
ICD
ICP
ICP-AES
ICP-MS, ICPMS
ID-MS
IFN
Ig
IGF-1
IL
ILL
immuno
IMP
iNOS
i.p., IP
IPSC
IQ
IRT
ISEL
ISI
i.v., IV
IVCD
JV
phosphoric acid
hypoxanthine phosphoribosyltransferase (gene)
heart rate
habitat suitability indices
sulfuric acid
heparan sulfate proteoglycan
hematocrit
hepatoma cells
catalytic subunit of human telomerase
hypertension
integrated blood lead index
integrated blood lead index x Wide Range Achievement
Test-Revised (interaction)
International Classification of Diseases
inductively coupled plasma
inductively coupled plasma atomic emission spectroscopy
inductively coupled plasma mass spectrometry
isotope dilution mass spectrometry
interferon (e.g., IFN-y)
immunoglobulin (e.g., IgA, IgE, IgG, IgM)
insulin-like growth factor 1
interleukin (e.g., IL-1, IL-lp, IL-4, IL-6, IL-12)
incipient lethal level
immunohistochemical staining
inosine monophosphate
inducible nitric oxide synthase
intraperitoneal
inhibitory postsynaptic currents
intelligence quotient
interresponse time
in situ end labeling
interstimulus interval
intravenous
intraventricular conduction deficit
juvenile
-------
KABC
KTEA
KXRF, K-XRF
LA
LB
LC
LD50
LDH
LDL
L-dopa
LE
LET
LH
LHRH
LN
L-NAME
LOAEL
LOEC
LOWES S
LPO
LPP
LPS
LT
LT50
LTER
LTP
LVH
liPIXE
liSXRF
MA
MA- 10
MANCOVA
MAO
Kaufman Assessment Battery for Children
Kaufman Test of Educational Achievement
K-shell X-ray fluorescence
lipoic acid
laying bird
lactation
lethal concentration at which 50% of exposed animals die
lethal concentration at which 74% of exposed animals die
lethal dose at which 50% of exposed animals die
lactate dehydrogenase
low-density lipoprotein (cholesterol)
3,4-dihydroxyphenylalanine (precursor of dopamine)
Long Evans (rat)
linear energy transfer (radiation)
luteinizing hormone
luteinizing hormone releasing hormone
lead nitrate
L-7V°-nitroarginine methyl ester
lowest-observed adverse effect level
lowest-observed-effect concentration
locally weighted scatter plot smoother
lipoperoxide
lipid peroxidation potential
lipopolysaccharide
leukotriene
time to kill 50%
Long-Term Ecological Research (sites)
long term potentiation
left ventricular hypertrophy
microfocused particle induced X-ray emission
microfocused synchrotron-based X-ray fluorescence
mature
mouse Ley dig tumor cell line
multivariate analysis of covariance
monoamine oxidase
-------
MATC
MDA
MDA-TBA
MDCK
MDI
MDRD
MEM
MG
Mg-ATPase
MiADMSA
Mi-DMSA
MK-801
MLR
MMSE
MMTV
MN
MND
MNNG
MPH
MRI
mRNA
MROD
MRS
MS
MSCA
mSQGQs
MT
MVV
MW
N, n
N/A
NAAQS
NAC
NAD
NADH
maximum acceptable threshold concentration
malondialdehyde
malondialdehyde-thiobarbituric acid
kidney epithelial cell line
Mental Development Index (score)
Modification of Diet in Renal Disease (study)
Minimal Essential Medium
microglobulin
magnesium-dependent adenosine triphosphatase
monoisamyl dimercaptosuccinic acid
mi monoisoamyl dimercaptosuccinic acid
NMD A receptor antagonist
mixed lymphocyte response
Mini-Mental State Examination
murine mammary tumor virus
micronuclei formation
motor neuron disease
7V-methyl-jV'-nitro-7V-nitrosoguanidine
Morphology
magnetic resonance imaging
messenger ribonucleic acid
methoxyresorufm-O-demethylase
magnetic resonance spectroscopy
mass spectrometry
McCarthy Scales of Children's Abiltities
mean sediment quality guideline quotients
metallothionein
maximum voluntary ventilation
molecular weight (e.g., high-MW, low-MW)
number of observations
not available
National Ambient Air Quality Standards
TV-acetyl cysteine
nicotinamide adenine dinucleotide
reduced nicotinamide adenine dinucleotide
-------
NADP
NAD(P)H, NADPH
NADS
NAF
NAG
Na-K-ATPase
NAWQA
NET
NCBP
NCD
NCS
NCTB
NCV
ND
NDI
NE
NES
NF-KB
NGF
NHANES
NIOSH
NIST
NK
NMDA
NMDAR
NMR
NO
NO2
NO3
NOAEC
NOAEL
NOEC
NOEL
NOM
NORs
nicotinamide adenine dinucleotide phosphate
reduced nicotinamide adenine dinucleotide phosphate
nicotinamide adenine dinucleotide synthase
nafenopin
7V-acetyl-p-D-glucosaminidase
sodium-potassium-dependent adenosine triphosphatase
National Water-Quality Assessment
nitro blue tetrazolium
National Contaminant Biomonitoring Program
nuclear chromatin decondensation (rate)
newborn calf serum
Neurobehavioral Core Test Battery
nerve conduction velocity
non-detectable; not detected
nuclear divison index
norepinephrine
Neurobehavioral Evaluation System
nuclear transcription factor-KB
nerve growth factor
National Health and Nutrition Examination Survey
National Institute for Occupational Safety and Health
National Institute for Standards and Technology
natural killer
7V-methyl-D-aspartate
7V-methyl-D-aspartate receptor
nuclear magnetic resonance
nitric oxide
nitrogen dixide
nitrate
no-observed-adverse-effect concentration
no-observed-adverse-effect level
no-observed-effect concentration
no-observed-effect level
natural organic matter
nucleolar organizing regions
-------
NOS
NOx
NP
NPSH
NR
NRC
NRK
NS
NSAID
NT
NTA
02
ODVP
OH
7-OH-coumarin
1,25-OH-D, 1,25-OHD3
24,25-OH-D
25-OH-D
8-OHdG
O horizon
OR
OSWER
P,P
P300
P450 1A1
P450 1A2
P450CYp3all
PAD
PAH
PAI-1
PAR
Pb
203Pb
204Pb, 206Pb, 207Pb, 208Pb
nitric oxide synthase; not otherwise specified
nitrogen oxides
net productivity
nonprotein sulfhydryl
not reported
National Research Council
normal rat kidney
nonsignificant
non-steroidal anti-inflammatory agent
neurotrophin
nitrilotriacetic acid
oxygen
offspring development
hydroxyl
7-hydroxy-coumarin
1,25-dihydroxyvitamin D
24,25-dihydroxyvitamin D
25-hydroxyvitamin D
8-hydroxy-2'-deoxyguanosine
forest floor
odds ratio; other oral
Office of Solid Waste and Emergency Response
probability value
event-related potential
cytochrome P450 1A1
cytochrome P450 1A2
cytochrome P450 Sal 1
peripheral arterial disease
polycyclic aromatic hydrocarbon
plasminogen activator inhibitor-1
population attributable risk
lead
lead-203 radionuclide
stable isotopes of lead-204, -206, -207, -208, respectively
lead-210 radionuclide
II-xlv
-------
Pb(Ac)2
PbB
PbCl2
Pb(C104)2
PBG-S
PBMC
Pb(NO3)2
PbO
PBP
PbS
PbU
PC12
PCR
PCV
PDE
PDGF
PDI
PEC
PEF
PG
PHA
Pi
PIXE
PKC
plNEpi
PMA
PMN
PMR
PN
P5N
PND
p.o.,PO
POMS
ppb
ppm
lead acetate
blood lead concentration
lead chloride
lead chlorate
porphobilinogen synthase
peripheral blood mononuclear cells
lead nitrate
lead oxides (or litharge)
progressive bulbar paresis
galena
urinary lead
pheochromocytoma cell
polymerase chain reaction
packed cell volume
phosphodiesterase
platelet-derived growth factor
Psychomotor Development Index
probable effect concentration
expiratory peak flow
prostaglandin (e.g., PGE2, PGF2); prostate gland
phytohemagglutinin A
inorganic phosphate
particle induced X-ray emission
protein kinase C
plasma norepinephrine
progressive muscular atrophy
polymorphonuclear leucocyte
proportionate mortality ratio
postnatal (day)
pyrimidine 5'-nucleotidase
postnatal day
per os (oral administration)
Profile of Mood States
parts per billion
parts per million
-------
PPVT-R
PRA
PRL
PROG
PRR
PRWT
PST
PTH
PTHrP
PVC
PWM
PRWT
QA/QC
Q/V
r
R2
r2
226Ra
R/ALAD
RAVLT
86Rb
RBA
RBC
RBF
RBP
RBPH
RCPM
REL
REP
RHIS
222Rn
RNA
ROS
ROS 17.2.8
RPMI 1640
Peabody Picture Vocabulary Test-Revised
plasma renin activity
prolactin
progeny counts or numbers
prevalence rate ratio
progeny weight
percent transferrin saturation
parathyroid hormone
parathyroid hormone-related protein
polyvinyl chloride
pokeweed mitogen
progeny weight
quality assurance/quality control
flux of air (Q) divided by volume of culture (V)
Pearson correlation coefficient
multiple correlation coefficient
correlation coefficient
most stable isotope of radium
ratio of ALAD activity before and after reactivation
Rey Auditory Verbal Learning Test
rubidium-86 radionuclide
relative bioavailablity
red blood cell; erythrocyte
renal blood flow
retinol binding protein
reproductive behavior
Ravens Colored Progressive Matrices
rat epithelial (cells)
reproduction
reproductive organ histology
most stable isotope of radon
ribonucleic acid
reactive oxygen species
rat osteosarcoma cell line
Roswell Park Memorial Institute basic cell culture medium
-------
RR relative risk; rate ratio
RT reaction time
RSEM resorbed embryos
RSUC reproductive success (general)
RT reproductive tissue
ESEM sum of the molar concentrations of simultaneously extracted metal
SA7 simian adenovirus
SAB Science Advisory Board
SAM ^-adenosyl-L-methionine
SBIS-4 Stanford-Binet Intelligence Scale-4th edition
s.c., SC subcutaneous
SCAN Test for Auditory Processing Disorders
SCE selective chemical extraction; sister chromatid exchange
SCP stripping chronopotentiometry
SD Spraque-Dawley (rat); standard deviation
SDH succinic acid dehydrogenase
SDS sodium dodecyl sulfate; Symbol Digit Substitution
SE standard error; standard estimation
SEM standard error of the mean
SES socioeconomic status
sGC soluble guanylate cyclase
SH sulfhydryl
SHBG sex hormone binding globulin
SHE Syrian hamster embryo cell line
SIMS secondary ion mass spectrometry
SIR standardized incidence ratio
SLP synthetic leaching procedure
SM sexually mature
SMAV species mean acute value
SMR standardized mortality ratio
SNAP Schneider Neonatal Assessment for Primates
SNP sodium nitroprusside
SC>2 sulfur dioxide
SOD superoxide dismutase
SOPR sperm-oocyte penetration rate
-------
SPCL
SPCV
SQGs
SRA
SRD
SRIF
SRM
SRT
SSADMF
SSB
SSEP
StAR
STORE!
SVC
SVRT
T
TA
TABL
T&E
TAT
TB
TEARS
TBPS
TCDD
Tcell
TCLP
TE
TEC
TEDG
TEL
TES
TEWT
TF
TG
TGF
sperm cell counts
sperm cell viability
sediment quality guidelines
Self Reported Antisocial Behavior scale
Self Report of Delinquent Behavior
somatostatin
Standard Reference Material
simple reaction time
Social Security Administration Death Master File
single-strand breaks
somatosensory-evoked potential
steroidogenic acute regulatory protein
STOrage and RETrieval
sensory conduction velocity
simple visual reaction time
testosterone
tail
time-averaged blood lead
threatened and endangered (species)
tyrosine aminotransferase
tibia
thiobarbituric acid-reactive species
Total Behavior Problem Score
methionine-choline-deficient diet
T lymphocyte
toxic characteristic leaching procedure
testes
threshold effect concentration
testes degeneration
tetraethyl lead
testosterone
testes weight
transferrin, translocation factor
6-thioguanine
transforming growth factor
-------
TH
232^
TLC
TNF
TOP
tPA
TPRD
TRH
TRY
TSH
TSP
TT3
TT4
TIES
TTR
TU
TWA
TX
U
235-rj 238-rj
UCP
UDP
UNECE
Ur
USFWS
USGS
UV
V79
VA
vc
VDR
VE
VEP
VI
vitC
tyrosine hydroxylase
stable isotope of thorium-232
Treatment of Lead-exposed Children (study)
tumor necrosis factor (e.g., TNF-a)
time-of-flight
plasminogen activator
total production
thyroid releasing hormone
toxicity reference value
thyroid stimulating hormone
triple-super phosphate
total triiodothyronine
serum total thyroxine
total testosterone
transthyretin
toxic unit
time-weighted average
tromboxane (e.g., TXB2)
uriniary
uranium-234 and -238 radionuclides
urinary coproporphyrin
uridine diphosphate
United Nations Economic Commission for Europe
urinary
U.S. Fish and Wildlife Service
United States Geological Survey
ultraviolet
Chinese hamster lung cell line
Veterans Administration
vital capacity; vitamin C
vitamin D receptor
vitamin E
visual-evoked potential
variable-interval
vitamin C
II-l
-------
vitE
VMA
VMI
VSM
VSMC
WAIS
WDS
WHO
wise
WISC-R
WO
WRAT-R
WT
WTHBF-6
ww
XAFS
XANES
XAS
XPS
X-rays
XRD
XRF
ZAP
ZnNa2 DTPA
ZnNa2 EDTA
ZPP
vitamin E
vanilmandelic acid
Visual-Motor Integration
vascular smooth muscle (cells)
vascular smooth muscle cells
Wechsler Adult Intelligence Scale
wavelength dispersive spectrometers
World Health Organization
Wechsler Intelligence Scale for Children
Wechsler Intelligence Scale for Children-Revised
whole organism
Wide Range Achievement Test-Revised
wild type
human liver cell line
wet weight
X-ray absorption fine structure
X-ray absorption near edge spectroscopy
X-ray absorption spectroscopy
X-ray photoelectron spectroscopy
synchrotron radiation
X-ray diffraction
X-ray fluorescence
correction in reference to three components of matrix effects:
atomic number (Z) absorption (A), and fluorescence (F)
zinc disodium diethylenetriaminepentaacetic acid
zinc disodium ethylenediaminetetraacetic acid
zinc protoporphyrin
Il-li
-------
CHAPTER 5 ANNEX
ANNEX TABLES AX5-2
May 2006 AX5-1 DRAFT-DO NOT QUOTE OR CITE
-------
Table AX5-2.1. Effect of Lead on Erythrocyte Morphology, Mobility, and Other Miscellaneous Parameters
to
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Dose & Route of
Exposure
Lead nitrate
0-100 uM,
Free Pb2+
0-20 uM, In vitro
Duration Species
(a) PB recovery Erythrocyte cell lysates from
studies, humans
10 min
(b) Relationship
offreePb2+to
added Pb,
20 min
Blood lead
Effect
— Uptake and transport of Pb in erythrocyte and across erythrocyte cell
a.
b.
c.
d.
membrane under the influence of varying buffers and ions.
Pb can cross the membrane passively in either direction.
Influx and efflux show similar properties
Passive transport of Pb is strongly stimulated by HCO3~
(bicarbonate)
Pb uptake is unaffected by varying the external concentrations of
Na+, K+, and Ca2+
In RBC, Pb binds mainly to hemoglobin. The ratio of bound Pb
to free Pb + in the cytosol is estimated 6000: 1
Authors
Simons (1986a)
Lead nitrate 1.5 mM
In vitro
Ih
Human erythrocytes
Pb uptake and transport are studied in resealed erythrocyte ghosts.
a. Transport of Pb across erythrocyte membranes is passive
b. 90% of Pb uptake by erythrocytes is inhibited by drugs that block
anion transport, indicating the involvement of anion exchanges
c. Pb transport depends upon the presence of a second anion. In the
Simons (1986b)
X
i
to
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p>
H
1
O
o
2|
0
H
O
O
H
W
O
^
O
H
W
10 uM lead, as lead 20 min
acetate,
In vitro
100 ug/dL lead, 1 h
10 mg/dL, lead,
In vitro
24 h
2 uM lead acetate 0- 1 h
2 uM lead
0-2 h
10 or 20 mM lead 5 weeks
acetate, i.p. once a
week
(100 or 200 umoles)/
kg b.wt.
presence ol HCO3 , the rate is stimulated in the order ol C1O4
< N03~ and CH3CO2~ < F" < Cl" < Br~ < L
Erythrocyte ghosts and — In erythrocytes, the anion exchange mechanisms and internal thiol
unsealed erythrocytes groups are critical factors that affect the stimulation of a
Ca2+-dependent process by Pb2+.
Human erythrocytes — Plasma lead uptake was at the rate of 0. 17 u moles/h.
Uptake comparable in erythrocyte ghosts and in intact cells.
No association of lead with membranes at 24 h.
MDCK Kidney epithelial cell — Anion exchange (AE) plays a critical role in regulating intracellular
line, In vitro pH in erythrocytes and epithelial cells and facilitates Pb uptake.
Human erythrocytes, in vitro
Albino rats Control - Exposure to lead significantly decreased the erythrocyte mobility.
1-12 jig/100 mL The decreases in mobility were either simultaneous or prior to the
Exposed - decreases in hemoglobin (Hb) or hematocrit (Ht). In exposed rats, a
100-800 ug/dL significant negative correlation was found between mobility and
blood lead levels. Decreases in ALAD (6-aminolevulinic acid), was
also apparent in exposed animals.
Lai et al. (1996)
Sugawara et al.
(1990)
Bannon et al. (2000)
Terayama et al.
(1986)
-------
to
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X
H
6
o
o
H
O
O
H
W
O
O
HH
H
W
Table AX5-2.1 (cont'd).
Dose & Route of
Exposure Duration
20 mM lead acetate, 5 weeks
i.p. once a week (200
umoles/kg b.wt)
200 uM of lead Once a week for
acetate, i.p. 5 weeks
Lead, i.p. 20 mg/ kg 14 consecutive
b.wt. days
1 uM lead nitrate 1 h
1 uM lead nitrate, 1 h
In vitro
6 and 12 mo
Effect of Lead on
Species
Male Wistar Albino rats
Rat
Male Albino rat
Erythrocyte Morphology, Mobility, and Other Miscellaneous
Blood lead
Control -
l-12ng/100mL
Exposed -
100-800 pg/dL
0-600 ug/dL
Erythrocytes from Controls -
lead-exposed healthy humans 8.3 ug/dL
Exposed -
70.5 ug/dL
Erythrocytes from healthy
human volunteers
Erythrocytes from
lead-exposed rats
—
Effect
Exposure to lead significantly decreased RBC membrane sialic acid
content, erythrocyte survival, hemoglobin, and hematocrit. This was
evident to a minor extent below blood lead levels 100 ug/100 mL
and was generally present from 100 ug/100 mL and higher.
Lead exposure significantly decreases RBC count, Hb values,
hematocrit, mean corpuscular volume, and mean corpuscular
hemoglobin, decreases erythrocyte mobility, membrane sialic acid
content, and deformability.
Acetyl choline esterase (AchE), NADH dehydrogenase, and Na+-K+
ATPase activities in rat erythrocyte membranes were inhibited by
lead exposure. Erythrocyte membrane sialic acid, hexose,
hexosamine were inhibited by lead exposure. Membrane
phospholipids and cholesterol were increased.
Lead exposure in healthy human RBC membranes resulted in
increased levels of arachidonic acid (AA). The increase in AA
correlated in a dose dependent manner with elevation in lead and
with serum iron. On the other hand, a negative correlation was
found between Aa and serum calcium. It is inferred that substitution
of lead to calcium, which is essential for the release of phospholipase
A2 for AA release may be the reason for increased RBC membrane
AA.
Lead inhibits Gordos effect in human erythrocytes; electron spin
labeling studies indicated cell shrinkage and decreased volume.
Cation-osmotic hemolysis (COH) in 12 mo lead-exposed rats was
lower in the areas of lower ionic strength on erythrocyte membranes.
Parameters
Authors
Terayma and
Muratsuga(1988)
Terayama et al.
(1993)
Jehang and Motlag
(1995)
Osterode and Ulberth
(2000)
Eriksson and Beving
(1993)
Mojzis and Nistiar
(2001)
0.1-200 pM, lead l-6h
nitrate in the reaction
buffer
0.1-10 uM lead ions
from 10 mM
Pb(NO3)2 solution,
In vitro
24 h
In vitro, human erythrocytes
Erythrocytes from healthy
human volunteers
Lead crosses the erythrocyte membrane by the anion exchanger and Simons (1993)
can also leave erythrocytes by a vanadate - sensitive pathway,
identified with the calcium pump. The high ratio of erythrocyte to
plasma Pb seen in vivo appeared to be due to the presence of a labile
Pb2+- binding component present in erythrocyte cytoplasm.
Pb activates erythrocyte K+ channels, Ca2+ sensitive erythrocyte Kempe et al. (2005)
Scramblase, triggers Phosphatidyl serine receptors and result in cell
shrinkage and decreased life span.
-------
Table AX5-2.1 (cont'd). Effect of Lead on Erythrocyte Morphology, Mobility, and Other Miscellaneous Parameters
to
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^
X
L/l
rv
O
§
H
6
o
2|
0
H
O
O
H
W
O
O
HH
H
W
Dose & Route of
Exposure Duration Species
20 uM lead ion, 2 min- 2 h Erythrocytes from human
In vitro umbilical cord
20 uM lead ions, 1 h Human umbilical cord
In vitro erythrocytes
Erythrocytes from Duration of Humans
lead-exposed workers exposure not
24-45 yr old white given. RBCs
males were isolated.
Experiments
were performed
in ghosts and
resealed
membranes
0.1 mM lead final Ih Erythrocytes from healthy
concentration, humans
In vitro
1-10 uM lead acetate, 3h Erythrocytes from healthy
In vitro humans
0-1 200 nM lead, Ih Erythrocytes from healthy
In vitro humans
Blood lead Effect
— Pb attenuates prolytic effect on neonatal erythrocytes in iso-or
hypotonic low ionic strength media.
Hemolytic activity of Organo leads increases with their
hydrophobicity: triethyllead chloride < tri-n-propyllead chloride <
tributyl tin chloride.
— Lead ions increase the resistance to lysis in media of diminishing
tonicity. These changes might be mediated by changes in membrane
structure.
1. 17 - 1.54 uM Increased blood lead in exposed workers was associated with a
significant decrease in the average micro viscosity of resealed and
unsealed erythrocyte membranes. Alterations in the microviscosity
of the lipid regions of the hydrophobic core of the erythrocyte
membrane bilayer and in the phospholipid composition of the
membrane may be defects that contribute to the clinical and
biochemical alterations/effects.
— Lead particles adhere to the external and internal surfaces of the
human erythrocyte membrane and disturb the lamellar organization
of lipid bilayers.
— Low concentrations of lead alter the physicochemical properties of
proteins and lipids in erythrocyte membranes.
— Significant increase in the phosphorylation of membrane cytoskeletal
proteins in lead treated human erythrocytes at concentrations above
100 nM mediated by enhanced PKC activity.
Authors
Serranietal. (1997)
Kleszcynska et al.
(1997)
Corchs et al. (2000)
Cooketal. (1987)
Suwalsky et al. (2003)
Slobozhanina et al.
(2005)
Belloni-Olivi et al.
(1996)
-------
Table AX5-2.1 (cont'd). Effect of Lead on Erythrocyte Morphology, Mobility, and Other Miscellaneous Parameters
to
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>
X
1
Dose & Route of
Exposure
Occupational,
Human exposure
Duration Species
— a. 24 adult healthy controls
(humans)
b. 12 patients with lead
poisoning (plumbism)
symptoms (Pb controls)
c. Patients with chronic renal
failure (CRF)
Divided into:
1. Normal urinary blood lead
levels
2. High urinary blood lead
levels
— a. 28 male workers in a lead
refining factory
b. Controls
Blood lead
Controls:
-17.1 ug/dL
Lead controls:
80.5 ug/dL
CRF-1:
18.4 ug/dL
CRF -2
18.0 ug/dL
Urinary lead
CRF 1
-322ug/72h
CRF 2
1785 ug/72 h
Exposed:
-35.97 ug/lOOg
Controls:
5.23 ug/100 g
Effect
Increased erythrocyte Zn protoporphyrin to free protoporphyrin ratio
in lead controls and remained in the normal range in CRF patients.
CRF patients showed minor abnormalities of erythrocyte heme
metabolism, such as low ALAD activity.
SDS polyacrylamide electrophoresis for erythrocyte membrane
proteins showed bands at 3 and 4. 1, that significantly decreased
while bands 2.3, 6, and 7 significantly increased in the lead workers
compared with controls
Authors
Fontanellas et al.
(2002)
Fukumoto et al.
(1983)
H
6
o
RBC—Red blood cells; Hb—Hemoglobin; NADH—Nicotinamide adenine dinucleotide dehydrogenase;
PKC—Protein kinase C; AchE—Acetyl choline esterase
o
H
O
c
o
H
W
O
V
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Table AX5-2.2. Lead, Erythrocyte Heme Enzymes, and Other Parameters
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Dose & Route of
Exposure Duration
Dietary, 35 days
O-lOOug/gdrywt. of
the diet
Lead acetate, oral 3 or 1 1 weeks
gavage, 1.5mg/kg
b.wt, lead acetate
20 |ig/mL as lead 5 weeks
acetate in drinking
water
17|iM Me/kg lead 5 days
acetate, Per OS
1.5 mg lead/ kg body 8 yrs.
wt, oral dose
Occupational 11 -22 yrs
exposure
0-20 mg Pb liter - 1 29 days
Species
Adult male Zebra finches
Red-tailed Hawks
Female Wistar Albino rats
Female
Rabbits
Cynomolgus Monkey,
in vitro
Dogs from urban and rural
areas of Greece.
Human erythrocytes from
exposed populations.
Juvenile Rainbow trout
erythrocytes
Blood lead Effect
0-1.5 ug/mL Significant negative correlation was observed between blood-Pb
concentration and log ALAD activity. RBC ALAD activity ratio is a
sensitive indicator of dietary lead concentration regardless of the mode
of exposure.
0.195-0.752 Erythrocyte phorphobilinogen synthetase was depressed significantly
ug/dL with in the 1st wk of treatment. Rapid but brief increase in free
protoporphyrin. Hematocrit, erythrocyte count, Hb were all decreased
and blood viscosity increased in exposed group.
37.8 ug/dL Lead exposure decreases hematocrit, hemoglobin, and the number of
erythrocytes and enhances blood viscosity
— Lead causes a significant decrease in blood ALAD activity, increases
free erythrocyte protoporphyrins, increases aminolevulinic acid and
coporphyrin excretion in urine.
— Kinetic analyses of erythrocyte 6- aminolevulinic acid revealed
differences in PH optimum and Michaelis constants with lead exposure.
The ALAD enzyme kinetics of lead exposed monkeys and humans are
similar.
326, 97-68 Significant negative correlation existed between blood-lead levels and
ug/L ALAD activity. 807-992 umol/PBG/LRBC/h is established as the
normal erythrocyte ALAD range for dogs
1.39-1.42 Liquid chromatography with inductively coupled plasma spectrometry
umol/1 had revealed ALAD to be the principle lead binding protein. The
percentage of lead bound to ALAD was influenced by a common
polymorphism in the ALAD gene.
— Significant decreases in the erythrocyte ALAD activity after a 29-day
exposure to 121 and 201 mg Pb liter - 1
Authors
Scheuhammer et al.
(1987)
Redigetal. (1991)
Toplan (2004)
Zareba and
Chmelnicka(1992)
Dorward and
Yagminas (1994)
Polizopoulou et al.
(1994)
Bergdahl et al. (1997)
Burden etal. (1998)
-------
Table AX5-2.2 (cont'd). Lead, Erythrocyte Heme Enzymes, and Other Parameters
to
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L/l
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W
Dose & Route of
Exposure
Lead acetate 160
mg/L in water
1.46 jimol/liter
In vitro
Lead 0.34 uM/L-1. 17
jiM/L, subcutaneous
injection
0-60 pM lead ion, in
vitro
200-500 ppm lead in
drinking water
0. 1- 100 uM lead ion,
In vitro
20-5 Jig/kg body wt
1 mg/ kg body wt.
Duration Species
8 weeks Wistar rats
— Fish from regions close to the
smelters and down stream
48 h Human whole blood
erythrocyte hemolysates,
normal and lead intoxicated
individuals
1 h Male albino New Zealand
rabbits
20 min Human erythrocyte lysates
14 or 30 days Male ddY mice
5 min Human erythrocyte ghosts
Pregnancy Erythrocytes from Sprague-
through lactation Dawley rats
Blood lead Effect
>20->40 Lead increases blood and liver lead, erythrocyte porphyrin content,
ug/dL hypoactivity of both hepatocytic and erythrocytic ALAD
— Smelter site fish had elevated lead concentrations, decreased ALAD
activity and species differences in this inhibitory activity were apparent
that could be attributed to Zn levels.
— The effects of various divalent cations on erythrocyte porphobilinogen
are concentration and PH dependent. Zn restores the lead inhibited
activity.
— Lead causes the most inhibition and Zn activation of rabbit Erythrocyte
porphobilinogen activity. Cu2+, Cd2+, and Hg2+ are intermediary. Each
divalent ion has a characteristic effect on the PH- activity relationship of
PBG-S.
— Human erythrocyte lysate porphobilinogen activity is increased by Zn2+
with a Km of 1.6 pM and inhibited by lead with a Ki of 0.07 pM, lead
reduced the affinity for the substrate 5- aminolevulinate, non-
competitively.
24-51 Lead inhibits erythrocyte and bone marrow P5'N activity. Erythrocyte
jig/100 mL ALAD activity was inhibited by 90%. Elevation of Urinary excretion of
ALA with no change in erythrocyte protoporphyrin and urinary co
porphyrin as against in the lead exposed humans indicates that
protoporphyrin metabolism might be more resistant to lead in mice than
humans.
— Under normal incubation conditions lead inhibits, Ca2+ -Mg2+ ATPase
with an IC50 of 6.0jiM. Lead inhibits Ca2+- Mg2+ ATPase related to
sulphahydryl groups above 1.0 jiM lead and by direct action of lead
upon Calmodulin below 1.0 uM.
— Na2+- K+- ATPase and Ca2+- Mg+- ATPase of erythrocyte membranes
from lead-depleted animals did not change in PO generation as compared
to 1 mg/kg b.wt lead animals, where as in Fl generation lead depleted
rats showed reduced activity.
Authors
Santos et al. (1999)
Schmitt et al. (2002)
Farant and Wigfield
(1987)
Farant and Wigfield
(1990)
Simons etal. (1995)
Tomokuni et al.
(1989)
Mas-Oliva(1989)
Eder etal. (1990)
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Table AX5-2.2 (cont'd). Lead, Erythrocyte Heme Enzymes, and Other Parameters
to
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Dose & Route of
Exposure Duration
20 mg Pb acetate/ Kg 14 days
b.wt, i.p, In vivo
Species
Male Albino rats erythrocytes
Blood lead Effect
— Lead significantly decreases erythrocyte membrane acetyl choline
esterase, NADH dehydrogenase, membrane sialic acid, hexose, and
hexosamine.
Lead ions inhibit aerobic glycolysis and diminish ATP level in human
erythrocytes in vitro. Magnesium partly abolishes these effects by
Authors
Jehang and Motlag
(1995)
Grabowska and
Guminska(1996)
stimulating Magnesium dependent enzymes. Effect is seen both by
direct addition of lead acetate to erythrocyte ghosts as well as in the
ghosts obtained after preincubation of erythrocytes with lead acetate.
Ca2+, Mg2+ ATPase is less sensitive and Mg ATPase is practically
insensitive to lead under these conditions.
X
I
oo
10-200 ug/dL lead
ions (lead acetate)
In vitro
Lead acetate through
water or i.p. 1 or 2
mg/Kg b.wt.
20 h
Human umbilical cord
erythrocytes
Every 4th day for Wistar rats
1 mo
— Lead significantly decreased the concentration of ATP, ADP, AMP, Bosiacka and
adenosine, GTP, GDP, GMP, Guanosine, IMP, inosine, hypoxanthine, Hlynczak (2003)
NAD and NADP concentrations.
1.51-35.31 The concentrations of adenosine tri phosphate (ATP), Guanosine Bosiacka and
ug/dL triphosphate (GTP), Nicotinamide adenine dinucleotide NAD+, Hlynczak (2004)
nicotinamide adenine dinucleotide phosphate NADP+ adenylate and
Guanylate (AEC and GEC) were significantly reduced in erythrocytes of
exposed animals. Results indicate lead ions disrupt erythrocyte energy
pathway.
H
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ALAD — Aminolevulinic acid; Cu2+—Copper; Cd2+—Cadmium; Hg2+—Mercury; PBG-S Porphobilinogen synthetase; Zn—Zinc; ATP—Adenosine triphosphate—ADP—Adenosine diphosphate;
AMP-Adenosine monophosphate;
GTP—Guanosine tri phosphate; GDP—Guanosine diphosphate; GMP—Guanosine monophosphate; IMP—Inosine monophosphate;
NAD—Nicotinamide adenine dinucleotide; NADP—Nicotinamide adenine dinucleotide phosphate
O
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S Table AX5-2.3. Lead Binding and Transport in Human Erythrocytes
<<
O Dose & Route of
^ Exposure Duration Species Blood lead Effect Authors
0-60 pM lead ion, In 20 minutes Human erythrocyte lysates — Human erythrocyte lysate porphobilinogen activity is increased by Zn + Simons et al. (1995)
vitro with a Km of 1.6 pM and inhibited by lead with a Ki of 0.07 pM, lead
reduced the affinity for the substrate 5- aminolevulinate, non-
competitively.
Zn—Zinc
X
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Table AX5-2.4. Lead Effects on Hematological Parameters
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Dose & Route of
Exposure Duration
4-6 mg/Kg b.wt, i.p., 15 and 30 days
daily
0.82mglead/kg 3 or 11 weeks
b.wt./day, oral gavage
17 uM Me/Kg b.wt 5 days
lead acetate or 3.5 mg
of Pb/kg body wt, i.p
17 uM Me/Kg b.wt 5 days i.p.
lead acetate or 3.5 mg
of Pb/ kg body wt, i.p.
or per OS 17.5 mg/kg
b.wt single injection
Cu deficient 1 mg 4 weeks
Cu/Kg
Marginal deficient 2
mg/kg
Control 5 mg Cu/Kg
High Zn 60 mg/kg.
0.02 - 40 ppm Pb, 90 days
dietary
20 ug/mL, lead 5 weeks
acetate in drinking
water
Species Blood lead Effect
Intact and splenctamized rats — Lead increases urinary 6-amino levulinic acid (ALA) excretion,
depletion in RBC hemoglobin content, and more number of reticulocytes
in peripheral blood, and results in accumulation of immature
erythrocytes both in intact and splenctomized rats.
Red-tailed hawks erythrocytes 0. 195-0.375 Activity of porphobilinogen synthase/ALAD was depressed significantly
mg/mL in lead exposed rats and did not return to normal values until 5 weeks
after the termination of the treatment. A rapid and relatively brief
increase in erythrocyte free proto porphyrin and a slower, prolonged
increase in Zn complex.
Female Rabbits 17.5 ug/dL Lead causes a significant inhibition of ALAD in the blood , increases
free erythrocyte protoporphyrin, and urinary excretion of
Aminolevulinic acid and coporphyrin
Female Rabbits — Lead induced ALAS activity in liver and kidney, both after i.p and p.o.
administration, i.p. administration of lead also induced kidney heme
oxygen levels.
Rat — Moderately high Zn in the diet reduces plasma copper but not plasma
ceruloplasmin.
Does not affect the recovery of plasma Cu or activity after oral copper
sulphate in Cu deficient diets.
Does not influence RBC Super oxide dismutase activity.
Male and female Swiss mice 0.7-13.0 Increased RBC number and increased hemoglobin and decreased
ug/dL hematocrit up on lead exposure.
Female Wistar Albino rats 37.8 ug/dL Erythrocyte count, hematocrit and hemoglobin were all decreased and
blood viscosity increased in lead exposed workers
Authors
Gautam and Roy
Chowdhury (1987)
Redigetal. (1991)
Zareba and
Chmielnicka(1992)
Chmielnicka et al.
(1994)
Panemangalore and
Bebe (1996)
lavicoli et al. (2002)
Toplan et al. (2004)
-------
Table AX5-2.4 (cont'd). Lead Effects on Hematological Parameters
to
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Dose & Route of
Exposure
Erythrocytes from
humans of
occupational lead
exposure and controls
In vivo and in vitro
Duration Species
— Male lead workers and
13 normal volunteers
Blood lead
Range
-20.6-71.3
ug/dL
Effect
Nicotinamide adenine dinucleotide synthetase activity in the lead
workers ranged from 0.08 to 1.1 umol/h per g of hemoglobin. 50% of
enzyme inhibition was observed at 40 ug/dL. Aminolevulinic acid
dehydratase activity decreased rapidly and reached a plateau at Pb-B
levels 40-60 ug/dL. 50% of enzyme activity inhibition was observed at
20 ug/dL.
Authors
Moritaetal. (1997)
>
X
In vivo; exposure
In vitro assays on
erythrocytes from
exposed populations
1. Workers exposed to
manganese (Mn) and
2. Workers exposed to lead
(Pb) without clinical
manifestations of
intoxication
Erythrocyte concentrations of adenyl nucleotides (ADP and ATP) were Nikolova and
elevated in both groups of workers and that of AMP in lead-exposed Kavaldzhieva et al.
workers. The ratio of ATP/ADP significantly increased in lead-exposed (1991)
workers.
ALA—Aminolevulinic acid; ALAS—Aminolevulinic acid synthetase; ALAD—Aminolevulinic acid dehydratase, RBC—Red blood cells
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Table AX5-2.5. Lead Interactions with Calcium Potassium in Erythrocytes
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Dose & Route of
exposure Duration Species
0-325 uM, lead 0-60 min In vitro
nitrate,
In vitro
0 uM- 5 mM lead, 0-100 min Human erythrocyte
In vitro hemolysates
1-4 uM lead acetate, 0-30 min Rabbit reticulocytes
In vitro
1-50 uM lead ion, 20 min Marine fish erythrocytes
In vitro
Pb depleted rats Gestation Sprague-Dawley rats
Pb concentration through to
<20 ug/kg 15 day of
Diet, oral lactation
Lead controls
200,800 ug/kg Pb2+ in
the form of supra pure
Pb acetate. Diet, oral
Intact or erythrocyte 10 min Healthy human erythrocytes
ghosts
0-100uMleadionor
lead nitrate in the
reaction mix,
In vitro
Blood lead Effect
— Pb modifies the threshold sensitivity of individual K+channels to Ca +
with a biphasic time course. The increase of Pb concentration increased
the extent of the initial inhibition and decreased the duration. The
inhibitory effect was not observed when addition of Calcium preceded
the addition of Pb. Pb decreased the rate of uptake of 86Rb
— Lead and Ca transport was carried out by a passive transport system with
two kinetic components (Michaelis- Menten and Hill) Pb and Ca were
capable of inhibiting the transport of the other metals in a non-
competitive way.
— Pb at low concentrations inhibits the uptake of Fe (II) into all three
(heme, cytosolic and stromal) fractions. The saturable components were
inhibited at lower concentrations of Pb than the non- saturable
components.
— Lead activates Ca2+ activated potassium channels. Treatment of
erythrocytes with 1-2 uM lead led to a minor intra cellular K loss and at
Pb concentrations of 20-50 uM 70% of potassium was lost.
— The concentration of CA2+ ions in erythrocytes of lead-depleted rats was
elevated in FI generation, without changes in P0 generation. The
elevation observed in depleted rats could be because of a reduction in
CA2+-Mg2+ ATPase.
— Modulation of C A2+-activatable K+ permeability was compared with
modulation of a membrane-bound oxidoreductase activity in human
erythrocytes. Lead, anitrion, and menadione had parallel effects on the
channel protein and the enzyme. The results demonstrate that the K+
channel and the enzyme are distinct membrane proteins and the enzyme
activity may influence channel gating.
Authors
Alveraz et al. (1986)
Salinas et al. (1999)
Qian and Morgan
etal. (1990)
Silkin(2001)
Loipfuhrer et al.
(1993)
Fehlau etal. (1989)
Pb—Lead; K+—Potassium; Na2+- K+ ATPase—sodium potassium ATPase; Ca2+- Mg2+ ATPase—Calcium, Magnesium ATPase.
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Table AX5-2.6. Lead, Heme and Cytochrome P-450
Dose & Route of
exposure
0-75mgofPb2+/Kgb.
Duration
0-30 h
Species
C57 BL/6 male mice
Blood lead Effect
— Lead causes an increase in 6-amino levulinic acid levels in plasma and a
Authors
Joveretal. (1996)
wt. i.p., Single
injection
decrease in the heme saturation of hepatic tryptophan -2,3 dioxygenase.
P-450- dependent activities, EROD and O-dealkylation of
alkoxyresorufins decreased progressively. Lead exposure decreased
mRNA levels of the P450 CYp3al 1. The decrease in P450 transcription
was a mechanism dependent on heme by inhibition of heme synthesis
and also by a mechanism independent of heme in which lead decreases
P-450 transcription.
X
EROD—Ethoxy resorufin - O- dealkylase.
CYp3al 1—Cytochrome P-450 Sail.
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Table AX5-2.7. Lead, Erythrocyte Lipid Peroxidation, and Antioxidant Defense
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Dose & Route of
exposure
7.5 mg of lead acetate
or 4.09 mg of lead Kg"
'b.wt, oral
5.46 mg lead as lead
acetate, oral
10 mg/kg b.wt lead
acetate, intra
muscular, daily
Pre treatment with
melatonin
A. ALA 40 mg/kg
b.wt every other day
and /or
B. Melatonin 10
mg/kg
Lead acetate 0.2%, in
drinking water,
followed by
individual or
combined treatment of
lipoic acid (25 mg/Kg
b.wt and DMSA 20
mg/kg b.wt, i.p.)
6-Aminolevulinic
acid, 1-5 mM, In vitro
Duration
28 days,
multiple
analyses at day
7,14,21 and 28
14 days,
multiple
analyses at day
0, 7 and 14
7 days
Every other day
3 times daily for
2 weeks
5 weeks
10 days
Species Blood lead Effect
Erythrocytes from male Calves 0.1-1.6 ppm Lead exposure significantly reduced erythrocyte super oxide dismutase
activity until day 21 followed by a marginal increase by day 28. Total,
protein-bound and non protein- bound -SH content of erythrocytes
declined.
Erythrocytes from female goats 0.09-1. 12 ppm Lead exposure caused a significant increase of erythrocytic GPx, SOD
and CAT activities, total thiol groups and total antioxidant status.
Rat — Lead significantly decreased heme synthesis, decreased Hb, decreased
liver 6- ALAS and erythrocyte ALAD. Markedly elevates hepatic lipid
peroxidation, reduced anti oxidant enzymes such as total sulphahydryl
groups and Glutathione. Pre Treatment with melatonin reduced the
inhibitory effect of lead on both enzymatic and non enzymatic
antioxidants and reduced the iron deficiency caused by lead.
Male Sprague- Dawley rats — Melatonin effectively protects nuclear DNA and lipids in rat lung and
spleen against the oxidative damage caused by the carcinogen ALA.
Male Albino rats 97.5 ug/dL Lead exposure results in decreased blood hemoglobin, hematocrit,
enhanced erythrocyte membrane lipid peroxidation, decline in the
activities of erythrocyte membrane Na+-K+ ATPase, Ca2+ ATPase, and
Mg2+ ATPase. Treatment with lipoic acid and/or DMSA reduced the
lead induced adverse changes in the biochemical parameters
CHO cells — 6- Aminolevulinic acid treatment induces oxidative stress in Chinese
hamster ovary cells by inducing Glutathione, Glutathione disulphide,
Malandialdehyde equivalents, and Catalase. N-acetyl cysteine
administration reverses the decrease in cell survival and colony
formation induced by 6- ALA.
Authors
Patra and Swarup
et al. (2000)
Mousa et al. (2002)
El- Missiry (2000)
Karbownik et al.
(2000)
Siva Prasad et al.
(2003)
Nealetal. (1997)
SOD — Super oxide dismutase; CAT — Catalase; ALAS — Aminolevulinic acid synthatase, ALAD — Aminolevulinic acid dehydratase; ALA — Aminolevulinic acid; GPX — Glutathione peroxidase
-------
ANNEX TABLES AX5-3
May 2006 AX5-15 DRAFT-DO NOT QUOTE OR CITE
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Table AX5-3.1. Summary of Key Studies on Neurochemical Alterations
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Subject
rat
PND16-18
rat
PND50
rat
PND7, 14,21,
28&50
rat
PND21
adult rat
Exposure Protocol
Hippocampal cultures
1500ppmPb(Ac)2
chow 10 d before breeding &
maintained to sacrifice
1500ppmPb(Ac)2
chow 10 d before breeding &
maintained to sacrifice
750 ppm Pb(Ac)2
chow from gestational day 0 to
PND21
Cultured PC 12 cells
water-0.1-1.0%Pb(Ac)2
from gestational day 1 5 to adult
Peak Blood Pb or
[Pb] used
O.l&l.OuM
PbCl2
3 1.9 ug/dL
—
46.5 ug/dL
0.03-10 uM
Pb(NO3)2
61. 8 ug/100 mL
Observed Effects
Pb blockage of IPSCs were partially reversible while EPSCs were not
Decreases the NR1 subunit splice variant mRNA in hippocampus
Alters NMDAR subtypes & reduces CREB phosphorylation
Increased expression of nicotinic receptors
Pb acts as a high affinity substitute for calcium in catecholamine
release
Hippocampal GLU & GABA release exhibits biphasic effects from
chronic Pb
Reference
Braga et al. (2004)
Guilarte and
McGlothan (2003)
Toscano et al. (2002)
Jett et al. (2002)
Westerink and
Vijverberg (2002)
Lasley and Gilbert
(2002)
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O
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O
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HH
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adultrat water-0.1-1.0%Pb(Ac)2
from gestational day 15 to adult
Cultured PC 12 cells
embryonic rat hippocampal neurons
117.6 ug/100 mL NMDA receptor function is upregulated
Lasley etal. (2001)
rat
750orl500ppmPb(Ac)2
chow from 10 d pre-mating to
PND14,21,&28
Cultured PC 12 cells
0.53 uM Pb(Ac)2 PKC is involved in TH upregulation but not downregulation of ChAT Tian et al. (2000)
100 fM-100 nM Decreases [Ca2+]i & increases Ca2+ efflux by a calmodulin-dependent Ferguson et al.
mechanism (2000)
61.1 ug/dL Dose-response effect between level of Pb and expression of NR1 gene Guilarte et al. (2000)
5-20 uM Pb(Ac)2 Induces expression of immediate early genes but requires PKC
Kim et al. (2000)
-------
Table AX5-3.1 (cont'd). Summary of Key Studies on Neurochemical Alterations
to
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X
^
O
H
O
0
H
O
O
H
W
O
O
HH
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W
Subject
rat
PND50
adult rat
adult rat
PND2
rat
rat
PND17
rat
PND7, 14,21,
28
rat
PND7, 14,21,
28
rat
PND22-adult
rat
PND28 56,
112
Exposure Protocol
750orl500ppmPb(Ac)2
chow from 10 d pre-mating to
PND50
calcineurin in mixture
cerebrocortical membranes
0.2% Pb(Ac)2
in water and chow
Cultured hippocampal neurons
Cultured hippocampal neurons
750 ppm Pb(Ac)2
chow from 14 d pre-mating to
experimental use
750 ppm Pb(Ac)2
chow from 14 d pre-mating to
experimental use
water - 0.2% Pb(Ac)2
from gestational day 16 to PND21
water - 1000 ppm Pb(Ac)2
from gestational day 4-use
Peak Blood Pb or
[Pb] used
3 1.9 ug/dL
10- 2000 pM
Pb(NO3)2
0.01-4 uM
free Pb(Ac)2
52.9 ug/100 mL
0.01-10 uMPbC!2
0.1-10uMPbCl2
59.87 ug/dL
59.87 ug/dL
—
39.6 ug/dL
Observed Effects
Reductions in NMDAR receptors result in deficits in LTP and spatial
learning
Has a stimulatory (low) and inhibitory (high) effect on calcineurin
Pb binds to the NMDA receptor channel in a site different from zinc
GLU & GABA release are inhibited independent of Pb exposure period
Inhibits glutamatergic and GABAergic transmission via calcium
channel
Increases tetrodo toxin- insensitive spontaneous release of GLU &
GABA
NMDAR-2A subunit protein expression is reduced in the hippocampus
Alters the levels of NMDA receptor subunits mRNA in hippocampus
Induces loss of septohippocampal cholinergic projection neurons in
neonates lasting into young adulthood
Significant increase in [ HJMK-801 binding after chronic exposure
Reference
Nihei et al. (2000)
Kem and Audesirk
(2000)
Lasley and Gilbert
(1999)
Lasley etal. (1999)
Bragaetal. (1999a)
Bragaetal. (1999b)
Nihei and Guilarte
(1999)
Guilarte and
McGlothan(1998)
Bourjeily and
Suszkiw(1997)
Ma etal. (1997)
-------
Table AX5-3.1 (cont'd). Summary of Key Studies on Neurochemical Alterations
t^J
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Subject Exposure Protocol
rat 50 or 1 50 ppm Pb(Ac)2
PND2 1 -adult water for 2 weeks - 8 mo
adult rat water - 0.2% Pb(Ac)2
from PNDO - adult
rat - 4 mo water - 0.2% Pb(Ac)2
from gestational day 16 to PND28
rat water at 50 ppm Pb(Ac)2
PND 111 for 90 d; start at PND2 1
Cultured bovine chromaffin cells
rat Homogenized cortex
Cultured bovine chromaffin cells
rats neuronal membranes
PND14 or 56
rat cortical synaptosomes
Peak Blood Pb or
[Pb] used
28.0 ug/dL
37.2 ug/100 mL
22.0 ug/dL
18 ug/dL
variable kind &
concentration
ranging Pb(Ac)2
variable kind &
concentration
chow containing
750 ppm Pb(Ac)2
l-50nMfreePb
orl uMPb(NO3)2
Observed Effects
Differential effects in [ HJMK-801 binding with dopamine & D]
agonists
Presynaptic glutamatergic function in dentate gyrus is diminished
Developmental Pb results in long-lasting hippocampal cholinergic
deficit
Decreases in vivo release of dopamine in the nucleus accumbens
Exerts dual stimulatory and inhibitory effects on adrenal PKC
Pb activates PKC in the range of 10'11 to 10'8 M
Pb and calcium activate the exocytotic release of norepinephrine
Review paper discussing Pb-calcium interactions in Pb toxicity
Review paper exploring Pb as a calcium substitute
Inhibitory effect on [3H]MK-801 binding & loss of binding sites in
neonates
Triggers acetylcholine release more effectively than calcium
Reference
Cory-Slechta et al.
(1997)
Lasley and Gilbert
(1996)
Bielarczyk et al.
(1996)
Kala and Jadhav
(1995)
Tomsig and Suszkiw
(1995)
Long etal. (1994)
Tomsig and Suszkiw
(1993)
Simons (1993)
Goldstein (1993)
Guilarte and Miceli
(1992)
Shao and Suszkiw
(1991)
-------
S Table AX5-3.1 (cont'd). Summary of Key Studies on Neurochemical Alterations
<<
O Peak Blood Pb
^ Subject Exposure Protocol or [Pb] used Observed Effects Reference
rat hippocampal neurons 2.5-50 \iM Pb C12 Pb has a blocking effect on the NMDA subtype of glutamate receptors Alkondon et al.
(1990)
rat brain protein kinase C 10"10 MPb salts Stimulates brain protein kinase C and diacylglycerol-activated calcium Markovac and
Goldstein (1988)
>
X
H
6
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o
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O
O
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W
O
O
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W
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Table AX5-3.2. Summary of Key Studies on Neurophysiological Assessments
to
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1
to
o
M
%•/
^
H
6
o
2!
•£—\
O
H
O
O
H
W
O
HH
H
Uv^
Subject
rat
PND22
rat
PND42-64
rat
PND130-210
adult rat
adult rat
adult rat
rat
PND90-130
rat 7- 18 mo
rat
PND13-140
adult rat
rat
PND4-30
rat
Exposure Protocol
250 ppm Pb(Ac)2
3-6 weeks (electro) or 7-13
weeks (immuno)
100,250, or 500 ppm
Pb(Ac)2 in chow for 3-6 w
0.2% Pb(Ac)2
in water
water - 0. 1-1. 0%Pb(Ac)2
from gestational day 16 to adult
0.2% Pb(Ac)2
in water
0.2% Pb(Ac)2
in water PNDO-21
750 ppm Pb(Ac)2
chow from 50 d pre-mating to
experimental use
water - 0.2% Pb(Ac)2
from gestational day 16 to
experimental use
750 ppm Pb(Ac)2
chow from 50 d pre-mating to
experimental use
water - 0.2% Pb(Ac)2
from PNDO - adult
Hippocampal neurons
750 ppm Pb(Ac)2
chow from 50 d pre-mating to
experimental use
Peak Blood Pb
or [Pb] used
30.8 ug/dL
54.0 ug/dL
75.4 ug/dL
11 7.6 ug/dL
30.1 ug/dL
30.1 ug/dL
16.04 ug/1 00 mL
—
28.5 ug/dL
1-100 uMPbC!2
16.2 ug/100 mL
Observed Effects
Reduces midbrain dopamine impulse flow & decreases dopamine D] receptor
sensitivity in nucleus accumbens
Decrease in number of spontaneously active midbrain dopamine neurons
Review paper examining glutamatergic components contributing to
impairments in synaptic plasticity
Deficits in synaptic plasticity in the dentate gyrus from early exposure
Biphasic dose-dependent inhibition of hippocampal LTP
Chronic Pb exposure significantly decreases range of synaptic plasticity
Impairments in LTP and paired-pulse facilitation in the hippocampal DG
NMDA-dependent forms of synaptic plasticity are more vulnerable than
NMDA-independent potentiation or paired pulse-facilitation
Impairs ability to maintain LTP over time in the dentate gyrus
Paired-pulse stimulation of CA3 region shows inhibitory mechanisms
Chronic Pb increases the threshold for LTP in dentate gyrus in vivo
Identified the nicotinic acetylcholine receptor as a target for Pb
LTP and learning are impaired if exposed to Pb in the immature brain
Reference
Tavakoli-Nezhad
and Pitts (2005)
Tavakoli-Nezhad
etal. (2001)
Lasley and Gilbert
(2000)
Gilbert et al.
(1999a)
Gilbert et al.
(1999b)
Zhao etal. (1999)
Ruanetal. (1998)
Gutowski et al.
(1998)
Gilbert and Mack
(1998)
Gutowski et al.
(1997)
Gilbert et al.
(1996)
Ishihara et al.
(1995)
Altmann et al.
(1993)
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Table AX5-3.3. Summary of Key Studies on Changes in Sensory Function
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O
o
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O
O
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W
Subject
mice
PND7-90
rat
PND21or90
monkey
1 3 years
monkey
adult rat
monkey 6 yr
rat
PND90
Exposure Protocol
0.15%Pb(Ac)2
in dams water from PNDO-21
rat retinas
0.02% & 0.2% Pb(Ac)2
in dams water PNDO-21 &
3 weeks as adult
2 mg/kg/day Pb(Ac)2
in capsule for 1 3 y
350 or 600 mg Pb(Ac)2
for 9.75 years
bovine retinas
rat retinas
0.02% & 0.2% Pb(Ac)2
in dams water PNDO-21
glycerine capsule with 25 or
2000 ng/kg/day Pb(Ac)2
0.2% Pb(Ac)2
in dams water PNDO-21
Peak Blood Pb
or [Pb] used
26 |ig/dL
0.01-10 iiM
PbCl2
59.0 |ig/dL
168.0 |ig/dL
55 |ig/dL
50pM-100nM
Pb(Ac)2
10'9tolO-4M
59.4 ng/dL
220 ng/dL
0.59 ppm
Observed Effects
Produces a rod photoreceptor- selective apoptosis inhibited by Bcl-xl
overexpression
Pb & calcium produce rod photoreceptor cell apoptosis via mitochondria
Functional alterations and apoptotic cell death in the retina
Mild increase in detection of pure tones outside of threshold
Consistent prolongations of latencies on the brain stem auditory evoked
potential
Direct inhibition of purified rod cGMP PDE, magnesium can reverse effect
Alters several physiological & biochemical properties of rod photoreceptors
Review paper examining effects upon auditory and visual function
Inhibits adult rat retinal, but not kidney, Na+, K+-ATPase
Morphological damage in the visual cortical area VI and V2
Long-term selective deficits in rod photoreceptor function and biochemistry
Reference
He et al. (2003)
He et al. (2000)
Fox etal. (1997)
Rice (1997)
Lilienthal and
Winneke(1996)
Srivastava et al.
(1995)
Fox etal. (1994)
Otto and Fox
(1993)
Fox etal. (1991)
Reuhl et al.
(1989)
Fox and Farber
(1988)
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Table AX5-3.4. Summary of Key Studies on Neurobehavioral Toxicity
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2
0
H
O
o
H
W
O
O
HH
H
W
Exposure
Subject Protocol
Rat, female, 75 or 300 ppm Pb(Ac)2
22 weeks
Rat 750 ppm Pb(Ac)2
Wistar
Rat, LE, 50 ppm Pb(Ac)2
postweaning
Rat, LE, male 50, or 150 ppm Pb for 3 mos
Postweaning
Rat, LE, male 50, or 150 ppm Pb for 3 mos
Postweaning
Rat, LE, male 50, or 1 50 ppm Pb for 3 mos
Postweaning
Rat, SD, Adult 500 ppm Pb(Ac)2
Rat, SD 0.2% Pb(Ac)2 during gestation
and lactation, postweaning
only, or continuously
Peak Blood Pb
or [Pb] Used
39 (ig/dL
15 ng/dL
15.1 ng/dL
10.8 and
28.5 ng/dL
9. 7 and
26.2 ng/dL after
3 and 7 mos
16.0 and
28.0 ng/dL
20.9 ng/dL
PND56: 3.8,
25.3, and 29. 9
lig/dL
Observed Effects
Significantly impaired on the alteration task with variable intertrial delays.
Pb-induced deficits in AAL in rats exposed to Pb either during pre-weaning
or pre- and postweaning: postweaning-only exposure caused reduced deficits
in AAL.
Quinpirole at 0.05 mg/kg reversed the effects of Pb on FI performance;
eticlopride had no effect on response rates in Pb-treated animals.
FR: 1 50-ppm rats - significantly higher response rates and component resets
than the low dose group and controls. Waiting behavior: wait time was lower
in both treated groups. 1 50-ppm rats - increased number of reinforcers and a
higher response to reinforcement ratio than low dose and controls.
D2 agonist quinpirole reversed the Pb-induced effects on FR-response rate,
FR resets, wait reinforcers, and wait time.
No Pb-induced effects on sustained attention.
Chronic Pb exposure attenuated the reinforcing effect of brain stimulation.
No Pb-associated effects in learning performance with just maternal or
postweaning exposure. Continually exposed rats tended to avoid less
frequently and in two-way active avoidance training, did not respond
more frequently.
Reference
Alber and Strupp
(1996)
Altmann et al.
(1993)
Areola and
Jadhav (2001)
Brockel and
Cory-Slechta
(1998)
Brockel and
Cory-Slechta
(1999a)
Brockel and
Cory-Slechta
(1999b)
Burkey and
Nation (1994)
(Chen et al.
(1997)
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Table AX5-3.4 (cont'd). Summary of Key Studies on Neurobehavioral Toxicity
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Table AX5-3.4 (cont'd). Summary of Key Studies on Neurobehavioral Toxicity
to
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ON
X
to
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6
o
0
H
O
o
H
W
O
O
HH
H
W
Exposure
Subject Protocol
Rat, F344, 2, or 1 0 mg/kg Pb(Ac)2
male
Rat, LE, male 1 00 or 350 ppm Pb(Ac)2
in dam's water PNDO-21
Rat, LE, male 50 or 1 50 ppm Pb(Ac)2
from weaning
Rat, LE, male 50 or 1 50 ppm Pb(Ac)2
Rat, LE, male 50 or 1 50 ppm Pb(Ac)2
in water PND21 -use
Rat, LE, male 50 or 1 50 ppm Pb(Ac)2
from weaning
Rat, LE, male 1 00 or 350 ppm Pb(Ac)2
from weaning
Rat, LE, male 50 or 500 ppm Pb(Ac)2
from weaning
Rat, LE, male 50 or 500 ppm Pb(Ac)2
from weaning
Peak Blood Pb
or [Pb] Used
2mg: 23; 10 mg:
42 (adult),
-48 (old),
- 58 (ig/dL
(young)
34 (ig/dL
-D
35.7 |ig/dL
30.6 ng/dL
15-25
30-50 ng/dL
35.0 |ig/dL
49.1 ng/dL
49.1 ng/dL
Observed Effects
Aging caused impaired accuracy: In both young and old rats: Pb-induced
increase in accuracy, at the longest delay periods (12 s) in young rats, and at the
short delay periods in old rats. Adults: not affected by Pb exposure.
Induced functional D2-D3 supersensitivity to the stimulus properties of agonist.
Altered cholinergic sensitivity due to Pb and several agonists.
Postweaning lead exposure resulted in an MK-801 subsensitivity.
(1) Enhances the stimulus properties of NMD A via a possible
dopaminergic path.
(2) Low level Pb exposure is associated with D] subsensitivity.
Pb exposures attenuated the decrements in rates produced by the two D!
agonists SKF38393 and SKF82958, and at 150 ppm, Pb exposure altered the
rate change associated with the low dose (0.033 mg/kg) of quinpirole.
Post- washout decrease in sensitivity to MK-801 .
Increases FI schedule-controlled behavior in nucleus accumbens.
Both DA and EEDQ, microinjected into the dorsomedial striatum, caused
increases or decreases in FI response rates, which depended on baseline FI
overall rates.
Reference
Cory-Slechta
etal. (1991)
Cory-Slechta
etal. (1992)
Cory-Slechta and
Pokora(1995)
Cory Slechta
(1995)
Cory Slechta et al.
(1996a,b)
Cory Slechta et al.
(1996c)
Cory-Slechta
(1997a)
Cory-Slechta
etal. (1998a)
Cory-Slechta
et al. (2002a)
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Table AX5-3.4 (cont'd). Summary of Key Studies on Neurobehavioral Toxicity
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Table AX5-3.4 (cont'd). Summary of Key Studies on Neurobehavioral Toxicity
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Subject
Monkey,
cynomolgus,
9-10 years of
age
Rat, male,
adult
Rat, LE
Rat, Wistar,
male
Rat, LE,
female
Rat, LE, male
and female
Exposure
Protocol
50 or 100 (ig/kg/day
Pb(Ac)2
500 ppm Pb(Ac)2 in chow for
105 days
1500 ppm Pb(Ac)2 gestation
and lactation
100 mg/kg/body weight by
injection
75 or 300 ppm Pb(Ac)2 in
water GDO experimental use
500 ppm Pb choride during
lactation
Peak Blood Pb
or [Pb] Used
15. 4 and 25. 4
Hg/dL; 10.9 and
13.1 ng/dL,
steady state
28 ng/dL
3.9 (ig/dL at
PND50
not reported
51 ng/dL
42 (ig/dL
Observed Effects
Pb-induced impairment in the presence, but not the absence, of irrelevant cues;
in the lower-dose group monkeys, impairment ended when the irrelevant
stimuli became familiar.
Chronic Pb exposure attenuates cocaine-induced behavioral activation.
Pb + enriched environment: enhanced performance in water maze; increased
gene expression in the hipppocampus of NMDAR subunit 1 and BDNF.
Pb-induced deficits in memory component of the radial arm maze test and in
retention of passive avoidance learning
Impairment of reversal learning as an associative deficit.
PND1 1 : no Pb-induced sex differences, effects on pup activity, and differences
in pup retrieval by dams.
Reference
Gilbert and Rice
(1987)
Grover et al.
(1993)
Guilarte et al.
(2003)
Haider et al.
(2005)
Hilson and Strupp
(1997)
Holloway and
Thor(1987)
PND26: Pb treatment influenced all social behavior tested (i.e., investigation
duration and frequency, crossover frequency, pinning) but did not change
activity levels.
PND36: Pb-treated pups demonstrated increased crossover frequencies but no
change in activity levels compared to controls.
W Rat, LE
H
6
o
250 ppm Pb(Ac)2 chronically Hippocampal
from gestation Pb levels
PND21: 1.73;
PND56: 1.02;
PND91:
0.91 ng/g
Pb-exposure had no effect on working memory at any age tested, but did affect Jett et al. (1 997)
reference memory (significant in females and nearly significant in males) in the
PND21 rats.
0
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O
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Table AX5-3.4 (cont'd). Summary of Key Studies on Neurobehavioral Toxicity
Subject
Rat, LE, male
Monkey,
rhesus
Monkey,
rhesus
Monkey,
rhesus
Monkey,
rhesus,
5-6 years
Monkey,
rhesus,
7-9 years
Monkey,
rhesus
Monkey,
rhesus
Exposure
Protocol
750 ppm Pb(Ac)2
maternally, permanently,
or postweaning only
Pb(Ac)2 testing first 4 wks of
life
1 mg/kg/day Pb(Ac)2
PND5-PND365
Pb(Ac)2 testing first 4 wks of
life
10 mg/kg/day pulses (2) and
chronic 0.7 for first year of life
10 mg/kg/day pulses (2) and
chronic 0.7 for first year of life
10 mg/kg/day pulses (2) and
chronic 0.7 for first year of life
350 or 600 ppm
in utero
Peak Blood Pb
or [Pb] Used
AtPNDlOO
1.8,21.3,22.8,
and26.3|ig/dL
35 (ig/dL
First year
-70 |xg/dL
16-mosPE:
—35 (xg/dL
35 (ig/dL
250-300 ng/dL
peak
80 for rest of year
1-4 wk:
63 |ig/dL
5-6 wk: 174
4yrs: 4
7yrs: 2
wk5: 56 during;
remainder of first
6 mos: 33-43
Hg/dL
50andllOng/dL
Observed Effects
Maternal and permanent exposure: impaired water maze performance, with
maternal exposure producing both the greatest escape latency and longest
escape path length. No effects on performance in the postweaning exposure
groups.
Pb-induced greater agitation, climbing, fear, and exploration of the periphery.
First year: Pb-induced disruption of social play, and increases in both self-
stimulation and fearful behavior were observed. 16 mos: continued
disruption.
Few differences between control and Pb-exposed monkeys were seen; less
stability in SNAP performance.
Pb-induced deficits occurred most commonly with short intertrial delays;
lose-shift errors, possibly due to perseveration.
Chronic L-dopa ameliorated the Pb-induced DSA deficits, which returned
following cessation of L-dopa administration: implicates DA mechanisms in
these impairments.
First 6 wks: Pb-induced lowered muscle tonus and greater agitation, no effects
on sensorimotor measures. PND14: no Pb-related effects on object
permanence task.
2 mos: Pb-induced decreased visual attentiveness in visual exploration task.
At age 12 to 15 mos, the high-dose group exhibited deficits in simple
discrimination learning: both groups showed impairments in the more
complex learning set formation trials; activity at 12-15 mos showed no
Pb-related effects.
Reference
Kuhmann et al.
(1997)
Laskey and
Laughlin (2001)
Laughlin et al.
(1991)
Laughlin et al.
(1999)
Levin and Bowman
(1986)
Levin etal. (1987)
Levin etal. (1988)
Lilienthal et al.
(1986)
-------
Table AX5-3.4 (cont'd). Summary of Key Studies on Neurobehavioral Toxicity
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Subject
Rat, SD, male,
PND60
Rat, Wistar
Rat, LE
RatLE,
PND53
Rat, Wistar
tested at
PND100 and
PND142
Rat, Wistar,
female
Rat, Wistar,
female
Rat, Wistar,
PND80
Exposure
Protocol
8orl6mgPb(Ac)2
500 ppm Pb(Ac)2 through
pregancy and lactation
75 or 300 ppm Pb(Ac)2
continuously
300 or 600 ppm during
gestation or gestation and
lactation
750 ppm though PND16
maternal exposure or
chronically
750 ppm Pb(Ac)2
750 ppm Pb(Ac)2
400 mg/L Pb C12 in dam's
water PND1-30
Peak Blood Pb
or [Pb] Used
6.8 (ig/dL
41.24ng/dL
(dams),
21.24ng/dL
(PND23),
(PND70
36 ng/dL
PND8: 36-43;
PND24: 27-34;
PND53: 131-158
PND110:
maternally
exposed: <3;
chronic:
34 ng/dL
17.3 ng/dL at
PND16
32-39 ng/dL
continuous
exposure
17.3 ng/dL at
PND16
32-39 ng/dL
continuous
exposure
PND8:
10-15 ng/dL;
PND21: -45;
PND80: 2-4
Observed Effects
Long-lasting changes in drug responsiveness to cocaine and related drugs.
PND23: Pb-induced increased ambulation in the open-field tests, decreased
exploratory behavior in the holeboard tests, and no differences from control in
the elevated maze tests.
PND70: Pb-induced increase in head dipping in the holeboard test, decrease
in social interaction time. No differences in the rotarod tests.
Impaired learning of a visual discrimination task.
No Pb-induced differences in learning rate, motivation, or response latency
for correct or incorrect responses. Pb-induced: increases in errors of omission
when a delay was imposed prior to cue presentation, trials that followed an
incorrect response, and response initiation.
Both Pb-treated groups learned the original discrimination comparably to
controls, but showed a deficit in retention; Pb-treated female rats took longer
to reach criterion in the acquisition learning and longer to eat the pellets in the
retention phase.
Pb-induced deficits in acquisition of learning, but not with concurrent
hippocampal lesions. Four weeks later, both lesioned and Pb-treated animals
showed impaired retention.
Pb and lesions of amygdala showed impairments in the acquisition phase of
the maze and impaired passive avoidance; neither treatment affected
locomotor activity. Continuously exposed rats showed greater deficits.
48 h PE: no Pb-induced changes in recall; 5 days PE: decline in recall
latency.
Reference
Miller etal. (2001)
Moreira et al.
(2001)
Morgan et al.
(2000)
Morgan et al.
(2001)
Munozetal. (1986)
Munozetal. (1988)
Munozetal. (1989)
Murphy and Regan
(1999)
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Table AX5-3.4 (cont'd). Summary of Key Studies on Neurobehavioral Toxicity
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0
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o
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O
O
HH
H
W
Subject
Rat, SD, male,
adult
Rat, SD,
PND120
Rat, SD,
PND70
Rabbit, Dutch
Belted, male
Monkey,
squirrel
Monkey,
squirrel
Monkey,
cynomolgus
Monkey,
cynomolgus
Monkey,
cynomolgus,
7-8 yr
Monkey,
cynomolgus,
7 -Syr
Exposure
Protocol
500 ppm Pb(Ac)2
16 mg Pb(Ac)2 via gavage 30 d
pre-pregnancy to PND21
16 mg Pb(Ac)2 via gavage 30 d
pre-pregnancy to PND21
Pb(Ac)2
mother's blood Pb from
gestation week 5-birth
in utero exposure
2 mg/kg/day of Pb(Ac)2
continuously
50 or 100 ^g/kg/day Pb(Ac)2
chronically beginning
atPNDl
1500ng/kg/dayPb(Ac)2
1500ng/kg/dayPb(Ac)2
continuously from birth,
during infancy only, or
beginning after infancy
Peak Blood Pb
or [Pb] Used
28.91 ng/dL
38.0 ng/dL
53.24 ng/dL
20, 40, and
80 (ig/dL
21-79 ng/dL
21-70 ng/dL
maternal
llS^g/dLat
PND100
33 ng/dL by
PND270
PND100: 15.4
and 25.4
PND300: 10.9
and 13.1 ng/dL
36 ng/dL
36 ng/dL
Observed Effects
Decreases sensitization to the locomotor-stimulating effects of cocaine.
Self-administering rats prenatally exposed to Pb demonstrate and increased
sensitivity to the relapse phase of cocaine abuse.
Increased sensitivity to cocaine in rats perinatally exposed to Pb.
Exposed males mated with nonexposed females. Offspring at PND25 showed
Pb-induced effects on exploratory behavior.
Reduced sensitivity to changes in reinforcement contingencies during
behavioral transitions and in steady state.
Pb-induced increase in the number of responses that failed to adequately
displace the bar in the FR schedule and possible subtle motor impairments.
At PND60: Pb-induced increased mean FR pause times, and, decreased FI
pause times. At 3 years of age: Pb-induced increased FI run rate, pause time,
and index of curvature. At both ages, Pb-induced increased variability of
performance.
Delayed alternation at 7-8 years of age: Pb-induced impairment of initial
acquisition of tasks; longer delays between alternations resulted in poorer
performance and perseverative behavior, sometimes lasting for hours.
Pb exposure in infancy only impaired spatial discrimination reversal tasks.
All Pb-treated groups: same impairments of initial acquisition, indiscriminate
responding, greater impairment with longer delays, and preservative
responses.
Reference
Nation etal. (1996)
Nation et al. (2003)
Nation et al. (2004)
Nelson etal. (1997)
Newland et al.
(1994)
Newland et al.
(1996)
Rice (1988)
Rice and Karpinski
(1988)
Rice (1990)
Rice and Gilbert
(1990a)
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Table AX5-3.4 (cont'd). Summary of Key Studies on Neurobehavioral Toxicity
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Subject
Monkey,
cynomolgus,
5-6 yr
Monkey,
cynomolgus,
3 or 7 yr
Monkey,
cynomolgus,
8-9 yr
Monkey,
cynomolgus,
0.5 or 3 yr
Rat, adult
Exposure
Protocol
1500ng/kg/dayPb(Ac)2
continuously from birth, during
infancy only, or beginning after
infancy
1500 (ig/kg/day Pb(Ac)2
1500ng/kg/dayPb(Ac)2
continuously from birth, during
infancy only, or beginning after
infancy
2000 (ig/kg/day Pb(Ac)2
16mgPb(Ac)2
Peak Blood Pb
or [Pb] Used
36 ng/dL
36 ng/dL
36 ng/dL
115ng/dL
83.2 ng/dL
Observed Effects
Post-infancy exposure impairs nonspatial discrimination reversal while
exposure during infancy exacerbates the effect.
Pb exposure during different developmental periods produce different effects
on Fl performance in juveniles versus adults.
Pb-treated monkeys in all three exposure groups learned more slowly, with
less impairment in infancy-only exposures, and showed perseverative
behavior.
Decreased interresponse times and a greater ratio of responses per
reinforcement on the differential reinforcement of low rate schedule.
Developmental Pb exposure results in enhanced acquisition of cocaine self-
Reference
Rice and Gilbert
(1990b)
Rice(1992a)
Rice(1992b)
Rice(1992c)
Rocha et al. (2005)
pregnancy to PND21
o
§
H
6
o
o
H
O
c|
o
H
W
O
HH
7s
O
i — i
H
W
Rat 0.5, 2.0, or 4.0 mM Pb(Ac)2 in
drinking water
Rat, F344
Rat, LE, male 0.2% Pb(Ac)2 from
PND25 until testing at
PND100
Rat, Wistar 750 ppm Pb(Ac)2
gestation and lactation
Rat, Wistar 0.03%, 0.09%,
or 0.27%
Pb(Ac)2
gestationally
ll-SO^g/dL
-42 |xg/dL
-30 in Pb
PND30:
25 |ig/dL
PND90:
0.113ng/dL
-30, -33, and
-42 (ig/dL at
PNDO, tested at
PND49
Pb-induced decreased retention in shuttle avoidance task. Pb-associated
increase in locomotor activity.
Pb-induced better performance using extra-maze spatial cues; Pb-treated rats
spent less time on the periphery of the maze.
Pb-exposure + isolation: spatial learning deficits. Pb-exposure + enrichment:
performed better than the isolated Pb group. Pb-induced decreases in
hippocampal levels of BDNF, NGF-p1, NT-3, and basic FGF.
At PND30 and 90: no Pb-associated changes in elevated maze behavior.
PND30: decreased freezing, increased ambulation, and increased grooming.
PND90: Pb-induced decreased freezing and increased ambulation.
Offspring of Pb-treated females mated with nonexposed males.
F2 generation at PND30
and 90: increased ambulation and decreased grooming.
Male offspring: all three doses impaired memory retrieval. Female offspring:
only the low dose affected memory retrieval. Motor performance and vision
were not affected by Pb
Rodrigues et al.
(1996)
Salinas and Huff
(2002)
Schneider et al.
(2001)
Trombini et al.
(2001)
Yang et al. (2003)
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Table AX5-3.5. Summary of Key Studies on Cell Morphology and Metal Disposition
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Subject Exposure Protocol
Rat, PND 110 Water-0.2% Pb(Ac)2 from
GD16-PND21oruse
Rat C6 glioma cells and human
astrocytoma cells
Rat pup astroglial cell culture
Rat, PND 60 1 500 ppm Pb(Ac)2 for 30-35 days
Rat, Cultured neurospheres
embryos
Young rat PND 0-20 = 600 ug/dL
PND 20-40 = 20-60 ug/dL
Cultured oligodendrite progenitor
cells - PND 2
Cultured oligodendrite progenitor
cells - PND 2
Cultured cerebellar granule neurons
Rat Cultured C6 glioma cells
Human,
l-4yr
Rat and human Cultured rat astroglial,
human neuroblastoma
Cultured GH3, C6, and HEK293
cells
Rat 2 g/1 Pb(Ac)2 in weanlings for 3 mos
Peak Blood Pb
or [Pb] Used
-D
5-10 uMPb(Ac)2
10uMPb(Ac)2
20.0 ug/dL
0.1-100 uM
Pb(Ac)2
131.3 ug/dL
1 uMPb(Ac)2
0. 1-100 uM
Pb(Ac)2
5-50 uM Pb(NO3)2 or
Pb(C104)2
1 uMPb(Ac)2
1 uMPb(Ac)2
l-10uMPb(NO3)2
39 ug/dL
Observed Effects
Reduction in hippocampal neurogenesis with no spatial learning
impairments.
Directly targets GRP78 and induces its compartmentalized
redistribution. GRP78 plays a protective role in Pb neurotoxicity.
Oxidative stress in astroglia results from Pb impairment of the Cu
transporter Atpase (Atp7a).
Significant deleterious effects on progenitor cell proliferation.
Differentially affects proliferation and differentiation of embryonic
neural stem cells originating from different brain regions.
Blood Pb during succimer chelation are not an immediate indicator
of brain. Brain Pb values are slower to respond even though blood
Pb is normal.
Pb inhibition of proliferation and differentiation of oligodendrocyte
cells requires PKC.
Interferes with maturation of oligodendrocyte progenitor cells.
Specific transport systems carry Pb into neurons.
Induces GRP78 protein expression and GRP78 is a strong Pb
chelator.
Half-life of blood Pb was dependent upon exposure
duration,ranging 10-38 mos.
Immature astroglia vs. neuronal cells are most likely to bind Pb in
the brain.
Review paper addressing lead-binding proteins in the brain and
kidney.
Cellular uptake of lead is activated by depletion of intracellular
calcium.
Chronic low Pb levels induces blood brain barrier dysfunction.
Reference
Gilbert et al. (2005)
Qian et al. (2005a)
Qian et al. (2005b)
Schneider et al.
(2005)
Huang and Schneider
(2004)
Stangle et al. (2004)
Deng and Poretz
(2002)
Dengetal. (2001)
Mazzolini et al.
(2001)
Qian et al. (2000)
Manton et al. (2000)
Lindahletal. (1999)
Fowler (1998)
Kerper and Hinkle
(1997)
Struzynska et al.
(1997)
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Table AX5-3.5 (cont'd). Summary of Key Studies on Cell Morphology and Metal Disposition
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Exposure Protocol
Peak Blood Pb
or [Pb] Used
Observed Effects
Reference
X
60 ug/dL
Discovered that albumin rarely enters brain from blood.
Results indicate a positive correlation between p32/6.3 levels and
neuronal maturation.
Examined the relationship between Pb and nuclear protein p32/6.3
and its abundance in intranuclear inclusion bodies.
Attenuation of Pb inhibition of ALAD involves sequestration of Pb
and a donation of zinc to the enzyme.
Injections of Pb-203 showed a linear uptake into three regions of
the brain, suggesting that the blood-brain barrier is rate-limiting.
Blood Pb half-life is affected by duration of exposure, age, and
length of follow-up.
Blood Pb half-life is dependent upon the length of exposure.
Bradbury et al.
(1991)
Klann and Shelton
(1990)
Klann and Shelton
(1989)
Goeringetal. (1986)
Bradbury and Deane
(1986)
Hryhorczuk et al.
(1985)
O'Flaherty et al.
(1982)
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Table AX5-3.6. Key Studies Evaluating Chelation of Pb in Brain
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Exposure Protocol
Chelator
Observed Effects
Reference
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Rat, Male, LE, Group 1: 50 ppm Pb acetate in CaEDTA
PND21 drinking water from PND21 for
3-4 mos, after which they
were given i.p. injections of
75 or 150 mg/kg CaEDTA for
either 1, 2, 3, 4, or 5 days.
Group 2: 25 or 500 ppm Pb
acetate followed by a single
injection of either 75 or
150 mg/kg CaEDTA. Twenty-
four hour urine samples were
collected following CaEDTA
injections.
Rat, Male, LE, 50 ppm Pb acetate from DMSA
PND21 weaning until testing 3-4 mos
later. The rats received either
25 or 50 mg/kg DMSA for 1, 2,
3,4, or 5 days and tissues were
evaluated 24 h following the
last injection.
Rat, Female, given 206Pb-enriched drinking DMSA
Wistar water at 210 ng Pb/mL for
36 h. Following an overnight
fast, the rats were injected with
one 0.25 mL i.p. injection of
0.11 mmol/kg DMSA. Pb
levels in blood, kidney, brain,
and tibia assessed 24 h later.
Group 1: PbB declined after the first CaEDTA injection, but did not drop
further with subsequent CaEDTA and never dropped below control levels
(5 ug/dL). Pb levels in urine increased similarly with both doses of CaEDTA.
Pb was found to be mobilized from both bone and kidney and initially
redistributed to brain and liver. Subsequent CaEDTA injections caused
declines in brain and liver Pb levels, but no net loss of Pb.
Group 2: a single injection of 150 mg/kg CaEDTA caused marked elevation
of brain Pb, which called into question use of injections of CaEDTA in
clinical diagnostic procedures.
Cory-Slechta et al.
(1987a)
PbB was decreased by DMSA dose-dependently, with levels dropping to
<5 ug/dL after 3 injection of the higher dose and 4 injections of the lower
dose. Pb levels dropped in brain and kidney immediately, and in liver
following a delay. Bone Pb did not decline, which contrasts with earlier
studies showing mobilization from bone following DMSA chelation. Another
group in this study received the same five days of DMSA injections, but was
evaluated 4 mos later. Pb concentrations in all tissues were comparable to
those seen in the first group, indicating that chelation therapy must be
continued to lower tissue Pb levels.
PbB declined 40%, Pb in urine increased 1500%, and changes in kidney and
brain tissue Pb levels varied inconsistently. Chelation did not result in
increased excretion of skeletal Pb compared to controls, nor did it show a
redistribution of Pb to brain.
Cory-Slechta (1988)
Smith and Flegal
(1992)
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Table AX5-3.6 (cont'd) Key Studies Evaluating Chelation of Pb in Brain
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Chelator
Observed Effects
Reference
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Rat, Female, 100 ppmPb acetate in drinking CaEDTA
Albino water for 4 weeks. During the
last two day of that exposure,
the rats were administered two
i.p. injections of 1 |ig stable
204Pb tracer. Animals then
received 1 to 5 consecutive
days of 150 mg/kg CaEDTA ;
assayed 24 h following the last
injection.
Rat, Male, SD Chelation with ongoing Pb DMSA
rats at 6 - 7 exposure; PbB were^lS ng/dL;
weeks 550 ppm Pb acetate in drinking
water for 35 days.
Group 1: continued on Pb only
for 21 days.
Group 2: received continued
Pb plus oral DMSA at 16, 32,
120, or240mg/kg/dayfor
21 days.
Group 3: discontinued on Pb
after the first 35 days and
received oral DMSA (16, 32,
or 240 mg/kg/day).
Rat, Male, dosed with 1000 ppm Pb in CaEDTA and
Wistar drinking water for 4 mos, then DMSA
treated for 5 days with: saline;
25 mg/kg DMSA orally, twice
daily; 75 mg/kg CaEDTA i.p.
once daily; or 25 mg/kg DMSA
twice daily plus 75 mg/kg
CaEDTA i.p. once daily. PbB
resulting from these treatments
were 46,22, 28, and 13 ng/dL,
respectively and brain Pb levels
were 49, 38, 26, and 22 ng/g,
respectively.
No redistribution of endogenous Pb into the brain following one CaEDTA
dose, no measurable reduction in brain or bone Pb levels, and reductions in
both kidney and blood Pb levels. Additionally, over the first day of treatment,
CaEDTA reduced the 204Pb tracer more effectively than the Pb from chronic
exposure, indicating greater biologically availability of Pb from recent
exposures.
DMSA treatment increased urinary Pb and decreased levels of Pb in blood,
brain, bone, kidney, and liver, even with continued Pb exposure.
Seatonetal. (1999)
Pappasetal. (1995)
The combined treatments produced an additive response in urinary Pb
elimination and elimination from blood, liver, kidney, brain, and femur.
Flora etal. (1995)
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Table AX5-3.6 (cont'd) Key Studies Evaluating Chelation of Pb in Brain
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Exposure Protocol
Chelator
Observed Effects
Reference
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Rat, Female, Group 1: 325 ug/mL Pb
LE acetate maternally through
weaning, and then to 30 ug/mL
until PND30. Chelation
treatment consisted of 7 days of
30 or 60 mg/kg/day DMSA.
Group 2: 325 ug/dL
maternally and through PND40
and then treated to DMSA for
7 or 21 days.
Rat, LE Exposed gestationally to
600 ug/mL Pb acetate, then
split into high and low dose
groups.
Low dose group: 20 ug/mL
from PND21 - 28, followed by
30 ug/mL from PND29 - 40.
High dose group: 40 ug/mL
from PND21 - 28, followed by
60 ug/mL from PND29 - 40.
DMSA treatment consisted of
50 mg/kg/day for 1 week, then
25 mg/kg/day for 2 weeks.
Rats received either 1 or
2 treatments at PND40 or
40 and 70.
DMSA
Seven days of DMSA effectively removed Pb from both blood and brain.
Treatment beyond 7 days further reduced brain Pb, but not blood Pb.
Reductions in Pb were greater in the second group, which the authors attribute
to the higher exposures used. The authors also hypothesize that DMSA-
mediated reduction in PbB are a poor indicator of reductions in brain Pb.
Smith etal. (1998)
DMSA
One treatment lowered both PbB and brain Pb, but the brain reductions lagged
the blood reductions both temporally and in magnitude. Following the second
DMSA treatment, they observed a rebound in blood, but not brain Pb levels.
Stangle et al. (2004)
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Table AX5-3.6 (cont'd) Key Studies Evaluating Chelation of Pb in Brain
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ANNEX TABLES AX5-4
May 2006 AX5-37 DRAFT-DO NOT QUOTE OR CITE
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Table AX5-4.1. Effect of Lead on Reproduction and Development in Mammals* Effects on Offspring
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Citation
al-Hakkak
et al. (1998)
Appleton
(1991)
Bataineh
et al. (1998)
Berry et al.
(2002)
Bogden et al.
(1995)
Camoratto
et al. (1993)
Corpas
(2002a)
Corpas
(2002b)
Species/
Strain/ Age
Mouse/BALB/c,
weaning
Rat/Long-Evans
hooded, adult
Rat/Sprague-
Dawley, adult
Rat/Sprague-
Dawley, 21 days
old
Rat/Sprague-
Dawley,
12 weeks old
Rat/Sprague-
Dawley, adult
Rat/Wistar, adult
Rat/ Albino
(NOS), adult
Dose/Route/
Form/Duration
0, 25, 50 mg lead monoxide
alloy/kg in chow for 35-70
days
Lead acetate single dose by i.v.
at 30 mg/kg
1000 ppm lead acetate in
drinking water for 12 weeks
Lead nitrate (1000 ppm lead)
in drinking water for 6 weeks
250 mg/L of lead acetate in
drinking water from GD 1 until
after 1 week after weaning
0.02% lead nitrate in drinking
water from gestation day 5 of
dams until PND4 of offspring
Lead acetate 0 or 300 mg/L in
drinking water during gestation
and lactation
Lead acetate 0 or 300 mg/L in
drinking water during gestation
and lactation
Endpoint
Reduced number of spermatogenia and spermatocytes in the 50 mg group after 70 days;
reduced number of implantations after mating (after 35 days exposure).
Increase in serum calcium and phosphorous; SEM analysis revealed 'lead line' in tooth
that was composed of hypomineralized interglobular dentine.
Fertility was reduced; total number of resorptions was increased in female rats
impregnated by males.
Mean plasma growth hormone levels decreased by 44.6%; reduced mean growth
hormone amplitude by 37.5%, mean nadir concentration by 60%, and growth hormone
peak area by 35%; findings are consistent with decreased hypothalamic growth
hormone-releasing factor secretion or reduced somatotrope responsiveness; exogenous
growth hormone in lead-treated and control rats, this response was blunted by the lead
treatment; plasma IGF 1 concentration was not significantly affected by lead treatment.
Dam and pup hemoglobin concentrations, hematocrit, and body weights and lengths
were reduced.
Female pups exposed to lead beginning in utero were smaller, no corresponding effect
in males; pituitary responsiveness to a hypothalamic stimulus.
Alterations in hepatic system of neonates (PND12) and pups (PND21); reductions in
hemoglobin, iron, alkaline and acid phosphatase levels, and hepatic glycogen, and
elevated blood glucose.
Effects energy metabolism; decrease in testis and seminal vesicle weights, and an
increase in DNA and RNA levels on PN day 2 1 ; protein was significantly decreased,
alkaline and acid phosphatase levels of the gonads were reduced; reduction of the
thickness of the epithelium and seminiferous tubule diameter.
Blood Lead Concentration
(PbB)
PbB not reported
PbB not reported
PbB not reported
PbB 37.40±3.60 ug/dL
PbB <15 ug/dL
PbB 17-43 ug/dL
PbB -22 ug/dL
PbB 54-143 ug/dL
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Table AX5-4.1 (cont'd). Effect of Lead on Reproduction and Development in Mammals* Effects on Offspring
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Species/
Strain/Age
Dose/Route/
Form/Duration
Endpoint
Blood Lead Concentration
(PbB)
X
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Table AX5-4.1 (cont'd). Effect of Lead on Reproduction and Development in Mammals* Effects on Offspring
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Species/
Strain/Age
Dose/Route/
Form/Duration
Endpoint
Blood Lead Concentration
(PbB)
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Fox et al.
(1997T)
Gandley et al.
(1999)
Govoni et al.
(1984)
Hamilton et al.
(1994)
Han et al.
(2000)
Hannah et al.
(1997)
Rat/Long-Evans
hooded, adult
Rat/Sprague-
Dawley, adult
Rat/Sprague-
Dawley, adult
Rat/Sprague-
Dawley,
25 days old
Rat/Sprague-
Dawley, 5
weeks old
Mouse/Swiss
ICR
preimplantation
embryos
0.02 or 0.2% lead acetate in
drinking water from PND
0-PND21; 8 female pups per
litter control pups; 8 pups per
litter low level exposure;
8 pups per litter moderate level
exposure (number of litters per
dose unspecified)
Male rats exposed to 25 or
250 ppm acetate lead in
drinking water for at least
35 days prior to breeding
2.5 mg/mL lead acetate in
drinking water from GD 16 to
postnatal week 8
Lead acetate in drinking water
at 250, 500 or 1000 ppm;
8 weeks prior to mating
through GD 21
250 mg/mL lead acetate in
drinking water for 5 weeks
followed by 4 week no
exposure (mated at end of
4-week no exposure period)
In vitro incubation of two- and
four-cell embryos with
0.05-200 uM lead acetate for
72 hours (time required for
blastocyst formation)
Developmental and adult lead exposure for 6 weeks produced age and dose-dependent
retinal degeneration such that rods and bipolar cells were selectively lost; at the
ultrastructural level, all dying cells exhibit the classical morphological features of
apoptotic cell death; decrease in the number of rods was correlated with the loss of
rhodopsin content per eye confirming that rods were directly affected by lead (p<0.05);
single-flash rod ERGs and cone ERGs obtained from lead-exposed rats demonstrated
that there were age- and dose-dependent decreases in the rod a-wave and b-wave
sensitivity and maximum amplitudes without any effect on cones; in adult rats exposed
to lead for three weeks, qualitatively similar ERG changes occurred in the absence of
cell loss or decrease in rhodopsin content (p<0.05); developmental and adult lead
exposure for three and six weeks produced age- and dose-dependent decreases in retinal
cGMP phosphodiesterase (PDE) activity resulting in increased CGMP levels (p<0.05);
retinas of developing and adult rats exposed to lead exhibit qualitatively similar rod
mediated ERG alterations as well as rod and bipolar apoptotic cell death (p<0.05);
similar biochemical mechanism such as the inhibition of rod and bipolar cell cGMP
PDE, varying only in degree and duration, underlies both the lead-induced ERG rod-
mediated deficits and the rod and bipolar apoptotic cell death (p<0.05).
Fertility was reduced in males with PbB in range 27-60 ug/dL, lead was found to affect
initial genomic expression in embryos fathered by male rats with blood lead levels as
low as 15-23 ug/dL; dose-dependant increases were seen in an unidentified set of
proteins with a relative molecular weight of approximately 70 kDa.
Decreased sulpiride binding in the pituitary is consistent with the elevated serum PRL
concentrations previously described in lead-exposed rats; DOPAc concentrations were
reduced by 21% in lead-treated rats.
Altered growth rates; reduced early postnatal growth; decreased fetal body weight.
Pups born to lead-exposed dams had significantly (p<0.0001) lower mean birth weights
and birth lengths.
Exposure of embryos to lead was only toxic at 200 uM, which reduced cell proliferation
and blastocyst formation.
PbB weanlings 19±3 (low
exposure) or 59±8 ug/dL
(moderate exposure), adult
7±2 ug/dL (at PND90)
PbB 27-60 ug/dL (fathers)
15-23 ug/dL (offspring)
PbB 71±8 ug/dL
PbB 40-100 ug/dL
PbB 10-70 ug/dL
PbB not reported
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Table AX5-4.1 (cont'd). Effect of Lead on Reproduction and Development in Mammals* Effects on Offspring
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Citation
lavicoli et al.
(2003)
Species/
Strain/ Age
Mouse/Swiss,
adult
Dose/Route/
Form/Duration
Lead acetate in food (0.02,
0.06,0.11,0.2,2,4,20,
Endpoint
Low-level exposure (PbB 2-13 ug/dL) reduced red cell synthesis (p<0.05); high-level
exposure (PbB 0.6-2 ug/dL) enhanced red cell synthesis (p<0.05).
Blood Lead Concentration
(PbB)
PbB 0.6 to <2.0 ug/dL or
>2.0-13 ug/dL
lavicoli et al. Mouse/Swiss,
(2004) adult
Logdberg Monkey/
et al. (1987) Squirrel, adult
Logdberg Monkey/
et al. (1998) Squirrel, adult
McGivern
etal. (1991T)
Nayak et al.
(1989)
Rat/Sprague-
Dawley, adult
Mouse/Swiss
Webster, adult
Piasek and Rat/Wistar,
Kostial (1991) 10 weeks old
40 ppm) exposure began 1 day
after mating until litter was
90 days old one litter of mice
exposed to each dietary
concentration
Lead acetate in feed; exposure
began 1 day after mating until
litter was 90 days old
Lead acetate p.o. exposure of
gravid squirrel monkeys from
week 9 of gestation through
PNDO
Lead acetate (varying
concentrations < 0.1% in diet);
maternal dosing from 5-8.5
weeks pregnant to PND1;
11 control monkeys, 3 low-lead
exposure group (PbB
24 ug/dL), 7 medium lead
group (PbB 40 ug/dL, 5 high-
lead group (PbB 56 Ug/dL)
0.1% lead acetate in drinking
water from GD 14 to
parturition
Lead nitrate dissolved in NaCl
solution, administered
intravenously, via caudal vein
at dose levels of 100, 150,
200 mg/kg; one time exposure
onGD9
7500 ppm lead acetate in
drinking water for 9 weeks
In females: accelerated time to puberty at PbB <3 ug/dL; delayed time to puberty at 3—
13 ug/dL.
Increase in pre- and perinatal mortality among squirrel monkeys receiving lead acetate
p.o. during the last two-thirds of pregnancy (45% vs. 7-8% among controls); mean
maternal PbB was 54 ug/dL (39-82 ug/dL); statistically significant reductions in mean
birth weight were observed in lead exposed monkeys as compared to controls; effects
occurred without clinical manifestation of toxic effects in the mothers.
Dose-dependent reduction in placental weight (p < 0.0007); various pathological lesions
were seen in the placentas (n = 4), including hemorrhages, hyalinization of the
parenchyma with destruction of the villi and massive vacuolization of chorion
epithelium; effects occurred without clinical manifestation of toxic effects in the
mothers.
Male offspring of dams exhibited reduced sperm counts, altered male reproductive
behavior, and enlarged prostates later in life; females exhibited delayed puberty,
menstrual irregularities, and an absence of observable corpora lutea; males and females
exhibited irregular release patterns of both FSH and LH later in life.
Chemical analysis showed lead was readily transferred across placenta; lead caused
moderate, statistically significant, increase in frequency of SCEs in maternal bone
marrow cells and significant reduction in NORs at the 2 highest dose levels (150 and
200 mg/kg); animals showed several specific chromosomal aberrations, mostly
deletions, in maternal bone, marrow, and fetal cells; aneupoidy was found to be
frequently associated with the lowest dose levels of lead nitrate (100 mg/kg); increased
embryonic resorption and reduced placental weights.
Decrease in litter size, pup survival, and birth weight; food consumption, body weight,
and fertility were not altered in 20 week exposure period.
PbB 0.6 to <2.0 |ig/dL or >2.0-
13 ug/dL
PbB 54 ug/dL (39-82 Ug/dL)
Mean maternal PbB 37 ug/dL
(22-82 ug/dL)
24 (22-26) ug/dL (low dose)
40 (35^16) ug/dL (mid dose)
56 (43-82) ug/dL (high dose)
PbB 73 ug/dL
PbB levels at birth in the
exposure groups for these
studies were >180 ug/dL
Maternal PbB >300 ug/dL
Offspring PbB >220 ug/dL
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Table AX5-4.1 (cont'd). Effect of Lead on Reproduction and Development in Mammals* Effects on Offspring
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Citation
Pinon-
Lataillade
et al. (1995)
Pillai and
Gupta (2005)
Ronis et al.
(1996T)
Ronis et al.
(1998aT)
Species/
Strain/ Age
Mouse/NMRI,
adult
Rat/Charles
Foster,
200-220 g
Rat/Sprague-
Dawley, 22,
55 days or plug-
positive time-
impregnated
Rat/Sprague-
Dawley, various
ages
Dose/Route/
Form/Duration
0-0.5% lead acetate in
drinking water exposed to
lead during gestation until
post-GD 60
Subcutaneous injection of
0.05 mg/kg-d lead acetate for
5-7 days prior to mating
through PND21
0.6% lead acetate in drinking
water for various durations:
PND24-74 (pubertal
exposure), PND60-74 (post
pubertal exposure); 1 1 males
and females in pubertal
exposure group (10 each in
control pubertal group);
6 males and females post-
pubertal exposure and control
groups
0.6% lead acetate in drinking
water ad libitum for various
durations: GD 5 to PND1,
GD 5 to weaning, PND1 to
weaning
3 control litters, 2 gestation
exposure litters, 2 lactation
exposure litters, 2 gestation
and lactation exposure litters,
2 postnatal litters, 2 chronic
litters (4 male and 4 female
pups per litter)
Endpoint
Lead exposure during gestation reduces litter size; reduced birth weight and growth
rates.
Long term exposure of rats (premating, gestational, and lactational) to moderate levels
of lead acetate (s.c.) resulted in reduced activities of hepatic steroid (E2) metabolizing
enzymes (17-6-hydroxy steroid oxidoreductase and UDP glucuronyl transferase) and
decreased hepatic CYP450 content.
Reduction in serum testosterone levels in male, not female; in female suppression of
circulating E2 (p<0.05) and LH (p<0.05); reduction in male secondary sex organ weight
(p<0.0005); delayed vaginal opening and disrupted diestrous in females (p<0.005);
increased incidence of stillbirth (2% control vs. 19% Pb) (p<0.005).
Dose-dependent delay in sexual maturation (delayed vaginal opening) (p<0.0002)
following prenatal lead exposure that continued until adulthood (85 days old); reduced
birth weight (p<0.05), more pronounced among male pups.
Blood Lead Concentration
(PbB)
PbB <4-132 ug/dL
PbB not reported
In utero PbB 250-300 ug/dL
Pre-pubertal PbB 30-60 ug/dL
Post-pubertal PbB 30-60 ug/dL
PbBs in the dams and offspring
in this experiment were
>200 ug/dL.
Group: pup PbB
Naive: ~6 ug/dL
Control: <2 ug/dL
Gest: -10 ug/dL
Lact: —3 ug/dL
Gest+Lact: -13 ug/dL
Postnatal: -260 ug/dL
Chronic: -287 ug/dL
Ronis et al.
(1998bT)
Rat/Sprague-
Dawley, adult
Lead acetate in drinking
water (0.05% to 0.45% w/v);
dams exposed until weaning,
exposure of pups which
continued until PND21, 35,
55, or 85; 5 control litters
(0%), 10 low-dose litters
(0.05%), 8 mid-dose litters
(0.15%), 9 high-dose litters
(0.45%); 4 male and 4 female
pups per litter
Prenatal lead exposure that continues until adulthood (85 days old) delays sexual
maturation in female pups in a dose-related manner (p<0.05); birth weight reduced
(p < 0.05), more pronounced among male pups; decreased growth rates (p<0.05) in both
sexes accompanied by decrease in plasma concentrations of IGF 1 through puberty
(p < 0.05) and a significant increase in pituitary and growth hormone during puberty
(p < 0.05).
PbBs in the pups between the
ages of 21 and 85 days were
>100 ug/dL and reached up to
388 ug/dL.
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Table AX5-4.1 (cont'd). Effect of Lead on Reproduction and Development in Mammals* Effects on Offspring
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Citation
Ronis et al.
(1998c)
Ronis et al.
(200 1T)
Species/ Dose/Route/
Strain/Age Form/Duration
Rat/Sprague- Lead acetate 0.05, 0. 15, or
Dawley, adult 0.45% in drinking water
beginning GD 5 continuing
until PND21, 35, 55, or 85;
5 control litters (0%),
10 low-dose litters (0.05%),
8 mid-dose litters (0. 15%),
9 high-dose litters (0.45%);
4 male and 4 female pups
per litter
Rat/Sprague- Lead acetate in drinking
Dawley, water to 825 or 2475 ppm ad
neonate, male libitum from G'D 4 to GD 55
(100 days) and postpartum; 1 male and
female pup female pup/litter (5 litters per
group) control group, 1 male
and female pup/litter (5 litters
per group) 825 ppm lead
acetate group, 1 male and
female pup/litter (5 litters per
group) 2475 ppm lead acetate
group
Endpoint
Dose-responsive decrease in birth weight (p<0.05), and crown-to-rump length (p<0.05);
dose-responsive delay in sexual maturity in male (p<0.05) and female (p<0.05);
neonatal decrease in sex steroids (p<0.05); pubertal decrease in testosterone (male)
(p<0.05) and E2 (female) (p<0.05); decrease estrous cyclicity at high dose (p<0.05).
Dose-dependent decrease of the load of failure in male (p<0.05); no difference in
plasma levels of vitamin D metabolites; reduced somatic growth (p<0.05), longitudinal
bone growth (p<0.05), and bone strength during the pubertal period (p<0.05); sex
steroid replacement did not restore skeletal parameters in lead exposed rats; L-Dopa
increased plasma IGFi concentrations, rates of bone growth, and bone strength
measures in controls while having no effect in lead exposed groups; DO gap x-ray
density and proximal new endostreal bone formation were decreased in the distration
gaps of the lead-treated animals (p<0.01); distraction initiated at 0.2 mm 30 to 60 days
of age.
Blood Lead Concentration
(PbB)
Dams: 0, 48, 88, or 181
Pups PND1: <1, -40, ~
>120 ug/dL
PupsPND21:50,
-237 ug/dL
PupsPND35:278 ug/dL
Pups PND55: <1, >68,
-380 ug/dL
Pups PND85: <1, >43,
>2 14 ug/dL
PbB at 825 ppm was
67-192 ug/dL
PbB at 2475 ppm was
120-388 ug/dL
ug/dL
70, or
>160, or
>70, or
>137, or
>122, or
Sant'Ana et al. Rat/Wistar,
(2001) 90 days old
Singh et al.
(1993)b
Watson et al.
(1997)
Rat/ITRC,
albino (NOS),
6 weeks old
Rat/Sprague-
Dawley, adult
0.1 and 1% lead in drinking
water
7 days
250, 500, 1000, and 2000
ppm lead nitrate in drinking
water from GD 6 to GD 14
Lead in drinking water at
34 ppm from weaning of
mothers through gestation
and weaning of offspring
until birth; 6 pups control
group, 6 pups experimental
group
1% Pb exposure reduced offspring body weight during treatment, no changes observed
after 0.1% exposure; no altered offspring sexual maturation, higher Pb improved sexual
behavior, while 0.1% reduced it; 0.1% Pb caused decrease in testis weight, an increase
in seminal vesicle weight, and no changes in plasma testosterone levels, hypothalamic
VMA levels were increased compared to control group; reduced birth weight and
growth rates.
Significantly reduced litter size, reduced fetal weight, and a reduced crown-to-rump
length, increased resorption and a higher blood-lead uptake in those groups receiving
1000 and 2000 ppm Pb; these also had a higher placental uptake; however the level was
the same in both groups; fetal lead uptake remained the same whether or not 2000 ppm
lead was given to an iron-deficient or normal iron groups of mothers.
Reduced body weight (p = 0.04); parotid function was decreased by nearly 30%
(p = 0.30); higher mean caries scores than the control pups (p=0.005); pre- and perinatal
lead exposure had significantly increased susceptibility to dental caries (p=0.015).
PbB36.12±9.49ug/dLor
13.08±9.42 ug/dL
PbB not reported
PbB 48±13 ug/dL
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Table AX5-4.1 (cont'd). Effect of Lead on Reproduction and Development in Mammals* Effects on Offspring
Citation
Wiebe et al.
(1998)
Species/
Strain/ Age
Rats/Sprague-
Dawley, adult
Dose/Route/
Form/Duration
20 or 200 ppm lead chloride
in drinking water; prior to
pregnancy, during pregnancy,
lactation
Endpoint
Exposure to lead did not affect tissue weights but did cause a significant decrease in
gonadotropin-receptor binding in the prepubertal, pubertal, and adult females;
conversion of progesterone to androstenedione and dihydrotestosterone was
significantly decreased in 21 -day old rats, in 150-day old females, the exposure to lead
resulted in significantly increased conversion to the 5-alpha-reduced steroids, normally
high during puberty.
Blood Lead Concentration
(PbB)
PbB 4.0±1.4to 6.6±2.3 ug/dL
*Not including effects on the nervous or immune systems.
•(•Candidate key study.
cGMP, cyclic guanosine—3',5'-monophosphate; DO, distraction osteogenesis; DOPAc, 3,4-dihydroxyphenylacetic acid; E2, estradiol; ERG, electroretinographic; FSH, follicle stimulating hormone;
GD, gestational day; IGF!, insulin-like growth factor 1; i.v., intravenous; kDA, kilodalton; LH, luteinizing hormone; NOS, not otherwise specified; PbB, blood lead concentration; PDE,
phosphodiesterase; PND, post-natal day; p.o., per os (oral administration); s.c., subcutaneous; SEM, standard error mean; UDP, uridine diphosphate; VMA, vanilmandelic acid
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Table AX5-4.2. Effect of Lead on Reproduction and Development in Mammals* Effects on Males
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Citation
Acharya et al.
(2003)
Adhikari et al.
(2000)
Adhikari et al.
(2001)
Alexaki et al.,
1990
al-Hakkak
et al. (1998)
Barratt et al.
(1989)
Bataineh et al.
(1998)
Batra et al.
(2001)
Batra et al.
(2004)
Bizarro et al.
(2003)
Boscolo et al.
(1998)
Species/
Strain/Age
Mouse/Swiss,
6-8 weeks old
Rat/Druckrey,
28 days old
Rat/Druckrey,
28 days old
Bulls/Holstein,
3-5 years old
Mouse/
BALB/c,
weaning
Rat/Wistar,
70 days old
Rat/Sprague-
Dawley, adult
Rat/Portan,
8 weeks old
Rat/Portan,
8 weeks old
Mouse/CD-I,
adult
Rat/Sprague-
Dawley,
weanling
Dose/Route/Form/Duration
200 mg/kg lead acetate
through i.p. injection of lead;
one time injection
0.0, 0.4, 4.0, 40.0 uM lead
acetate in vitro for 24 and
48 hours
5, 10, and 20 mg/kg lead in
distilled water by gavage for
2 weeks
In vitro fertilization 2.5 or
0.25 ng/mL
0, 25, 50 mg lead monoxide
alloy/kg in chow for 35-70
days
0, 0.3, 33, 330 mg lead
acetate/kg-d in drinking
water, by gavage for 63 days
1000 ppm lead acetate in
drinking water for 12 weeks
10, 50, 200 mg/kg lead
acetate orally for 3 mo
10, 50, 200 mg/kg lead
acetate orally for 3 mo
0.01 M lead acetate twice a
week for 4 weeks
60 mg lead acetate/mL in
drinking water for 1 8 mo
1 ml point
Testicular weight loss with constant increase in the incidence of abnormal sperm
population; decrease in sperm count; testicular ascorbic acid also declined significantly;
significant rise in LPP of tissue; LPP is indicative of oxidative stress in treated mice
testes.
Germ cells progressively detached from Sertoli cell monolayer into medium in a
concentration and duration dependent manner Viability of the detached cells showed a
decrease with increase in time and concentration of Pb; leakage of LDH recorded at
higher dose of 4.0 and 40.0 uM.
Induced significant numbers of germ cells to undergo apoptosis in the semiferous
tubules of rats treated with highest dose; DNA fragmentation was not detected at any of
the doses; level of lead accumulation in testes increased in a dose-dependent manner.
Sperm motility reduced significantly at 2. 5 ug/mL; lower concentration had no effect
on sperm motility.
Reduced number of spermatogenia and spermatocytes in the 50 mg group after 70 days;
reduced number of implantations after mating (after 35 days exposure).
Increased number of abnormal post-testicular sperm in the highest exposure group;
reduced number of spermatozoa at PbB >4.5 ug/dL.
Fertility was reduced in males.
Lead in testis and epididymis increased with dose; administration of zinc reduced lead
levels; dose related changes in activities of enzyme alkaline phosphatase and Na+-K+-
ATPase, which decreased with increased dose of lead; improvement in activities of
enzymes was seen in groups given lead and zinc; disorganization and disruption of
spermatogenesis with accumulation of immature cells in lumen of tubule; highest dose
of lead resulted in arrest of spermatogenesis, and decrease in germ cell layer
population; highest dose levels, damage of basement membrane, disorganization of
epithelium and vacuolization cells; tubules were found almost empty, indicating arrest
of spermatogenesis.
LH and FSH concentrations were decreased at 200 mg/kg; decrease in fertility status at
200 mg/kg; decline in various cell populations at 200 mg/kg; 50 mg/kg group hormone
levels, cell numbers, and fertility status were found close to normal.
Dose-time relationship was found; ROS role.
Increased vacuolization in Sertoli cells; no other ultrastructural modifications; no
impairment of spermatogenesis.
Blood Lead Concentration
(PbB)
Not reported
PbB not applicable-in vitro study
PbB not reported
PbB not applicable-in vitro study
PbB not reported
PbB 2, 4.5, 7, 80 ug/dL
PbBs >40 ug/dL
PbB not reported
PbB not reported
PbB not reported
PbB not reported
PbB 4-17 ug/dL
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Table AX5-4.2 (cont'd). Effect of Lead on Reproduction and Development in Mammals* Effects on Males
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Qtation
Chowdhury
etal. (2001)
Chowdhury
et al. (1984)
Chowdhury
et al. (1986)
Chowdhury
et al. (1987)
Coffigny et al.
(1994|)
Corpas et al.
(1995)
Corpas et al.
(2002a)
Corpas et al.
(2002b)
Species/
Strain/Age
Mouse/
BALB/c,
3 months old
Rat/ Albino,
(NOS), adult
Rat/NOS, adult
Rat/Charles
Foster, 150±5 g
Rat/Sprague-
Dawley, adult
Rat/Wistar,
adult
Rat/Wistar,
adult
Rat/Wistar,
adult
Dose/Route/Form/Duration
0.0,0.2,0.5, 1.0, 2.0ug/mL
lead acetate in culture
medium for 2 hours
(superovulated ova and
sperm)
Dietary concentrations of
0.25, 0.50, or 1.0 g/L lead
acetate for 60 days
0, 1, 2, 4, 6 mg lead
acetate/kg-d i.p. for 30 days
0, 1, 2, 4, 6 mg lead
acetate/kg-d/i.p. for 30 days
Inhalation exposure to 5
mg/m3 lead oxide daily for
13 days during gestation
(GD 2, 3, 6-10, 13-17, 20)
300 mg/L lead acetate via
drinking water beginning GD
1 through 5 day postnatal or
throughout gestation and
early lactation
300 mg/L acetate lead in
drinking water beginning at
mating until PND12 and 21
300 mg/L acetate lead in
drinking water beginning at
mating until PND12 and 21
Blood Lead Concentration
Endpoint (PbB)
Significant dose dependent decrease in the number of sperm attaching to the ova in both PbB not applicable-in vitro study
exposed groups; decrease in the incorporation of radio-labeled thymdine, uridine, and
methionine.
Testicular atrophy along with cellular degeneration was conspicuous at 1 g/L; high PbB 54-143 ug/dL
cholesterol concentration and significantly low ascorbic acid concentration were found
in the testes at 1 g/L; lowest dose (0.25 g/L) had no significant morphological and
biochemical alterations, whereas as 0. 5 g/L resulted in partial inhibition of
spermatogenesis.
Dose-related decrease oftestis weight; at 187 ug/dL: degenerative changes in testicular PbB 20, 62, 87, 187, or
tissues; at 325 ug/dL: degenerative changes and inquiry of spermatogenetic cells; 325 ug/dL
edematous dissociation in interstitial tissue.
Dose related decrease oftestis weight at 56 |lg of spermatoids; at 91 ug/dL: inhibition PbB 56-3332 ug/dL
of post-meiotic spermatogenic cell; at 196 ug/dL: decreased spermatogenic cell count
(6), detachment of germinal call layers; at 332 ug/dL: Decreased spermatogenic cell
count, degenerative changes, Interstitial edema, and atrophy of Ley dig cells.
Adult male offspring exhibit no change in sperm parameters or sex hormones T, FSH, PbB 71.1 ug/dL (dam)
and LH (because of duration or timing). PbB 83.2 ug/dL (fetal)
Testicular weight and gross testicular structure were not altered; seminiferous tubule PbB 14 ug/dL
diameter and the number of prospermatogonia were reduced; total DNA, RNA, and
protein content of the testes in treated rats was significantly reduced, DNA: RNA ratio
remained unaltered.
Neither abnormalities in the liver structure nor depositions of lead, toxicant produced PbB 22 ug/dL
biochemical alterations; pups exhibited decrease in hemoglobin, iron and alkaline, and
acid phosphatase levels and an increase in Pb content; protein, DNA, and lipid total
amounts were reduced, and hepatic glycogen content was diminished at 12 and 21 PN,
with a higher dose of glucose in blood; decrease in alkaline phosphatase in liver of pups
at day 2 1 PN, but acid phosphatase was unaltered.
Neither abnormalities in the liver structure nor depositions of lead, toxicant produced PbB 22 ug/dL
biochemical alterations; pups exhibited decrease in hemoglobin, iron and alkaline, and
acid phosphatase levels and an increase in Pb content; protein, DNA, and lipid total
amounts were reduced, and hepatic glycogen content was diminished at 12 and 21 PN,
with a higher dose of glucose in blood; decrease in alkaline phosphatase in liver of pups
at day 2 1 PN, but acid phosphatase was unaltered.
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Table AX5-4.2 (cont'd). Effect of Lead on Reproduction and Development in Mammals* Effects on Males
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Citation
Cory-Slechta
et al. (2004|)
Foote (1999)
Foster et al.
(1993)
Foster et al.
(1996a)
Foster et al.
(1998)
Gandley et al.
(1999)
Species/
Strain/Age
Rat/Long-Evans,
adult
Rabbit/Dutch-
belted, adult
Monkey/
Cynomolgus,
adult
Monkey/
Cynomolgus,
adult
Monkey/
Cynomolgus,
adult
Rat/Sprague-
Dawley, adult
Dose/Route/Form/Duration
Lead acetate in drinking
water beginning 2 months
before breeding until the end
of lactation
0, 0.005, 0.01, and 0.025 mM
PbC12 in vitro; one time dose
0-1500 |lg lead acetate/kg-d
in gelatin capsules p.o. for
various durations: 9 control
monkeys, 4 monkeys in
lifetime group (birth to 9
years), 4 in infancy group
(first 400 days of life), 4 in
post-infancy exposure (from
300 days to 9 years)
0-1500 |lg lead acetate/kg-d
in gelatin capsules p.o. from
birth until 9 years of age: 8
control monkeys, 4 monkeys
in low group (6-20 ug/dL), 7
monkeys in high group (22-
148 Ug/dL)
0-1500 |lg lead acetate/kg-d
in gelatin capsules p.o. for
various durations: birth to
10 years (lifetime); PND300
to 10 years (post-infancy);
birth to 300 days (infancy);
3 control monkeys, 4 lifetime,
4 infancy, 5 post-infancy
Male rats received lead
acetate 25 or 250 ppm in
1 ml point
Observed potential effects of lead and stress in female; Pb alone (in male) and Pb plus
stress (in females) permanently elevated corticosterene levels in offspring.
Six out of 22 males tested showed appreciable spontaneous hyperactivation, lead did
not affect hyperactivation, or associated capacitation.
Suppressed LH response to GnRH stimulation in the lifetime group (p=0.0370); Sertoli
cell function (reduction in the inhibin to FSH ratio) (p=0.0286) in lifetime and post-
infancy groups.
Mean PbB of 56 ug/dL showed no significant alterations in parameters of semen
quality (count, viability, motility, or morphology).
Circulating concentrations of FSH, LH, and testosterone were not altered by treatment;
semen characteristics (count, motility, morphology) were not affected by treatment
possibly because not all Sertoli cells were injured; degeneration of seminiferous
epithelium in infancy and lifetime groups (no difference in severity between these
groups); ultrastructural alterations in seminal vesicles, most prominent in infancy and
post-infancy groups.
High dose reduced fertility; low dose altered genomic expression in offspring.
Blood Lead Concentration
(PbB)
PbB 30-40 ug/dL
PbB not applicable-in vitro study
Lifetime group 3-26 ug/dL at 4-5
years; infancy group 5-36 ug/dL
at 100-300 days, 3-3 ug/dL at
4-5 years; post-infancy group
20-35 ug/dL
PbB 10±3 or 56±49 ug/dL
PbB -35 ug/dL
PbB 15-23 ug/dL or
27-60 ug/dL
Gandley et al.
(1999)
Gorbel et al.
(2002)
Rat/Sprague-
Dawley, adult
Rat/(NOS),
90 days old
Male rats received lead
acetate 25 or 250 ppm in
drinking water for 35 days
prior to mating
3 mg (PI) or 6 mg (P2) lead
acetate in drinking water for
15, 30, 45, 60, or 90 days
High dose reduced fertility; low dose altered genomic expression in offspring.
Male rats, absolute and relative weights of testis, epididymis, prostate and seminal
vesicles were found to significantly decrease at day 15 in P2 group and at day 45 in PI
group, at day 60 these absolute values and relative weights returned to control values; at
day 15 arrest of cell germ maturation, changes in the Sertoli cells, and presence of
apoptotic cells were observed; serum testosterone level was found to be lowered at day
15 in both PI and P2, and peaked at day 60, then returned to normal values.
PbB 15-23 ng
27-60 ug/dL
;/dL or
PbB not reported
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Table AX5-4.2 (cont'd). Effect of Lead on Reproduction and Development in Mammals* Effects on Males
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Qtation
Graca et al.
(2004)
Hsu et al.
(1997)
Hsu et al.
(1998a)
Hsu et al.
(1998b)
Huang et al.
(2002)
Johansson
(1989)
Johansson and
Pellicciari
(1998)
Johansson and
Wide (1986)
Species/
Strain/Age
Mouse/CD- 1,2
months old
Rat/Sprague-
Dawley,
7 weeks old
Rat/Sprague-
Dawley,
100-120 g
Rat/Sprague-
Dawley,
7 weeks old
Mouse MA- 10
cells
Mouse, 9 weeks
old
Mouse/NMRI, 9
weeks old
Mouse/NMRI, 9
weeks old
Dose/Route/Form/Duration
Subcutaneous injection of
74 mg/kg-d of lead chloride
for 1 to 3 days
10 mg/kg lead acetate
through i.p. injection to males
for 6 or 9 weeks
20 or 50 mg lead acetate via
i.p. route weekly to males for
6 weeks
10 mg/kg lead acetate weekly
via i.p. injection to males for
6 weeks
10"8 to 10"5 M lead incubated
for 3 hours
0-1 g lead chloride/L in
drinking water for 112 days
1 g/L lead chloride in
drinking water for 16 weeks
0-1 g/L lead chloride in
drinking water for 84 days
1 ml point
Reversible changes in sperm (count) and ultrastructural changes in testes (reduced
diameter of seminiferous tubules).
Six- week group had unchanged epididymal sperm counts, percent of motile sperms, and
motile epididymal sperm counts compared with control group; 9-week group showed
statistically lower epididymal sperm counts, and lower motile epididymal sperm counts;
good correlation between blood lead and sperm lead; significantly higher counts of
chemiluminescence, they were positively associated with sperm lead level; epididymal
sperm counts, motility, and motile epididymal sperm counts were negatively associated
with sperm chemiluminescence; SOPR were positively associated with epididymal
sperm counts, motility and motile epididymal sperm counts, sperm chemiluminescence
was negatively associated with SOPR.
Serum testosterone levels were reduced; percentage of capacitation and the
chemiluminescence were significantly increased in fresh cauda epididymal
spermatozoa; serum testosterone levels were negatively associated with the percentage
of acrosome-reacted spermatozoa; sperm chemiluminescence was positively correlated
with the percentage of both capacitated and acrosome-reacted spermatozoa; SOPR was
negatively associated with the percentage of both capacitated and acrosome-reacted
spermatozoa.
Intake of VE and/or VC in lead exposed rats prevented the lead associated sperm ROS
generation, increased the epididymal sperm motility, enhanced the capacity of sperm to
penetrate eggs harvested from unexposed female rats in vitro; protective effect of VE
and VC not associated with reduced blood or sperm lead levels.
Higher decreases in human chorionic gonadotropin (hCG)-stimulated progesterone
production, expressions of StAR protein, and the activity of 36-HSD compared to
2 hours; no affect on P450scc enzyme activity.
No effects on frequency of motile spermatozoa, nor on swimming speed; decreased
fertilizing capacity of the spermatozoa by in vitro fertilization; premature acrosome
reaction .
Decreased uptake of PI was found in spermatozoa from the vas deferens of the lead-
exposed mice; after thermal denaturation of the DNA, the spermatozoa showed a higher
uptake of PI in comparison to those of the controls; after reductive cleavage of S-S
bonds with DTT and staining with a thiol-specific reagent significantly fewer reactive
disulfide bonds were also observed in the spermatozoa; significant delay in the capacity
for NCD was noted.
No effects on sperm count; no effects on serum testosterone; reduced number of
implantations after mating.
Blood Lead Concentration
(PbB)
PbB not reported
PbB after 6 weeks 32 ug/dL,
after 9 weeks 48±4.3 ug/dL
PbBs >40 ug/dL
PbB30.1±3.4to36.1±4.6
PbBs >40 ug/dL
PbB not applicable-in vitro study
PbB 0.5-40 ug/dL
PbB 42±1.6 ug/dL
PbB <0.5-32 ug/dL
Mean tissue lead content
difference between lead treated
and controls: testicular 11 ug/g
(epididymal 67 ug/g)
PbB<0.5 ug/lOOmL
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Table AX5-4.2 (cont'd). Effect of Lead on Reproduction and Development in Mammals* Effects on Males
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Citation
Johansson
etal. (1987)
Kempinas
etal. (1998)
Kempinas
etal. (1990)
Kempinas
etal. (1994)
Klein et al.
(1994)
Liu et al.
(2001)
Liu et al.
(2003)
Species/
Strain/Age
Mouse/NMRI,
9-10 weeks old
Rat/Wistar,
adult
Rat/NOS,
pubertal
Rat/Wistar,
50 days old
Rat/Sprague-
Dawley,
100 days old
Mouse,
MA- 10 cells
Mouse,
MA- 10 cells
Dose/Route/Form/Duration
1 g/L lead chloride in
drinking water for 16 weeks
0.5 g/L and 1.0 g/L lead
acetate in drinking water for
90 days
(1.0 g/L) lead acetate in
drinking water in addition to
i.v. injections of lead acetate
(O.lmg/100gbw)every
10 days, 20 days
(1.0 g/L) lead acetate in
drinking water in addition to
i.v. injections of lead acetate
(O.lmg/lOOgbw) every
1 5 days, 9 months
0-1 g/lead acetate/L in
drinking water + 0.1 mg/kg
i.v. every 10 days for 20 days
0-1 g lead acetate/L in
drinking water + 0.1 ug/kg
i.v. every 15 days for
270 days
0.1, 0.3, or 0.6% lead acetate
in distilled water for 21 days
10'8to 10'5 lead acetate in
vitro for 2 hours
10"8 to 10"5 lead acetate in
vitro for 6 hours incubated
1 ml point
Spermatozoa had significantly lower ability to fertilize mouse eggs; morphologically
abnormal embryos were found.
PbB exhibited a significant increase in both groups; decrease in hematocrit and
hemoglobin, together with a rise in glucose levels; no signs of lesion were detected
upon histological examination of testes, caput, and cauda epididymidis; an increase in
ductal diameter, and a decrease in epithelial height were observed in the cauda
epididymidis; concentration of spermatozoa stored in the caudal region of the
epididymis exhibited a significant increase in lead-treated animals.
Basal levels of testosterone were higher both in the plasma and in the testes of acutely
intoxicated animals; levels of LH were not affected in either group, nor was the LHRH
content of the median eminence; density of LH/hCG binding sites in testicular
homogenates was reduced by saturnism in both groups, apparent affinity constant of the
hormone-receptor, complex significantly increased.
Increased plasma and testicular testosterone concentrations; no effects on testicular
weight; reduced weight of prostate; increased weight of seminal vesicle and seminal
secretions.
2-3 fold enhancement of mRNA levels of GnRH and the tropic hormone LH; 3-fold
enhancement of intracellular stores of LH; mRNA levels of LH and GnRH and pituitary
levels of stored LH are proportional to blood levels of lead.
Significantly inhibited hCG- and dbcAMP-stimulated progesterone production in MA-
10 cells; steroid production stimulated by hCG or dbcAMP were reduced by lead;
expression of StAR protein and the activities of P450 side-chain cleavage (P450) and
36-HSD enzymes detected; expression of StAR protein stimulated by bdcAMP was
suppressed by lead at about 50%; progesterone productions treated with 22R-
hydroxycholesterol or pregnenolone were reduced 30^10% in lead treated MA-10 cells.
Lead significantly inhibited hCG- and dbcAMP-stimulated progesterone production
from 20 to 35% in MA-10 cells at 6 hours; lead suppressed the expression of
steriodogenesis acute regulatory (StAR) protein from 30 to 55%; activities P450 side-
chain cleavage (P450scc) enzyme and 36-HSD were reduced by lead from 15 to 25%.
Blood Lead Concentration
(PbB)
PbB not reported
PbB 65-103 ug/dL
PbB -40 ug/dL
PbB 10-41 ug/dL
PbB 8.5^10 ug/dL
PbB 42-102 ug/dL
PbB not applicable-in vitro study
PbB not applicable-in vitro study
-------
Table AX5-4.2 (cont'd). Effect of Lead on Reproduction and Development in Mammals* Effects on Males
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Species/
Strain/Age
Dose/Route/Form/Duration
1 ml point
Blood Lead Concentration
(PbB)
X
I
o
H
6
o
o
H
O
o
H
W
O
O
HH
H
W
Marchlewicz Rat/Wistar,
et al. (1993) 90 days old
McGivern Rat/Sprague-
et al. (1991|) Dawley, adult
McMurry Rat/Cotton,
et al. (1995) adult
Mishra and Mouse/Swiss,
Acharya(2004) 9-10 weeks old
Moorman
et al. (1998)
Murthy et al.
(1991)
Murthy et al.
(1995)
Nathan et al.
(1992)
Pace et al.
(2005)
Rabbit/NOS,
adult
Rat/ITRC,
(NOS),
weanling
Rat/Druckrey,
adult
Rat/Sprague-
Dawley, adult
Mouse/BALB/c,
adult
Piasecka et al. Rat/Wistar,
(1995) adult
0-1% lead acetate in drinking
water for 270 days
0.1% lead acetate in drinking
water from GD 14 to
parturition: 8 control litters;
6 lead acetate litters (5 males
per litter)
0, 100, or 1000 ppm lead in
drinking water for 7 or
13 weeks
10 mg/kg lead acetate in
drinking water for 5 to
8 weeks
3.85 mg/kg lead acetate
subcutaneous injection for
15 weeks
0-250 ppm lead acetate in
drinking water for 70 days
Pb 5 mg/kg i.p. lead acetate
in drinking water for 16 days
0,0.05, 0.1, 0.5, or 1% lead
acetate in drinking water for
70 days
0.1 ppm lead acetate in
drinking water (lactational
exposure as neonates and
drinking water from PND21
to PND42)
1% aqueous solution of lead
acetate for 9 months
No histological or weight changes in testicle or epididymis; fever spermatozoa in all
zones of the epididymis.
Decreased sperm count (21% at 70 days and 24% at 165 days; p<0.05); reduced male
behavior (p<0.05); enlarged prostate (25% increase in weight; p<0.07); irregular release
patterns of both FSH and LH (p<0.05).
Immune function was sensitive to lead exposure; spleen mass was reduced in cotton rats
receiving 100 ppm lead; total leukocytes, lymphocytes, neutrophils, eosinophils, total
splenocyte yield, packed cell volume, hemoglobin, and mean corpuscular hemoglobin
were sensitive to lead exposure; reduced mass of liver, seminal vesicles, and epididymis
in males after 7 week exposure.
Stimulates lipid peroxidation in the testicular tissue, associated with increased
generation of noxious ROS; reduced sperm count, increased sperm abnormality
Increased blood levels associated with adverse changes in the sperm count, ejaculate
volume, percent motile sperm, swimming velocities, and morphology; hormonal
responses were minimal; dose-dependent inhibition of sperm formation; semen quality,
threshold estimates ranged from 16 to 24 ug/dL.
At 20 ug/dL no impairment of spermatogenesis; vacuolization of Sertoli cell cytoplasm
and increase in number and size of lysosomes.
Swelling of nuclei and acrosomes round spermatids; in Sertoli cells, nuclei appeared
fragmented, whereas the cytoplasm exhibited a vacuolated appearance and a few
structures delimitated by a double membrane that contains microtubules arranged in
parallel and cross-striated fin fibrils, cell tight junction remain intact; no significant
change in epididymal sperm motility and counts, testicular blood levels were found to
be elevated after lead exposure.
No effects on spermatogenesis in all groups; at 124 ug/dL: decreased seminal vesicle
weight; decreased serum testosterone in the 0.5% group at 10 weeks; no effects in the
other exposure categories; no effects on serum FSH, LH, nor pituitary LH content.
Reduction in fertility when mated with unexposed females; no change in sperm count;
increase in number of apoptotic cells intestes.
Lead-loaded (electron dense) inclusions were found in the cytoplasm of the epididymal
principal cells, especially in the caput of epididymis, also present, but smaller, in
smooth muscle cells; inclusions were located in the vacuoles, rarely without any
surrounding membrane; similar lead-containing structures were found in the epididymal
lumen.
PbB not reported
Control PbB <5 ug/dL at birth
Maternal PbB 73 ug/dL at birth
Pup PbB 64 ug/dL at birth
PbB not reported
PbB not reported
PbB 0, 20, 40, 50, 70, 80, 90, and
110 ug/dL
PbB 20.34± 1.79 ug/dL
PbB 7.39 ug/dL
PbB 2.3, 40, 44, 80, or 124 ug/dL
Neonatal PbB 59.5 ug/dL
Post PND21 PbB 20.3 ug/dL
PbB not reported
-------
Table AX5-4.2 (cont'd). Effect of Lead on Reproduction and Development in Mammals* Effects on Males
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Qtation
Piasek and
Kostial(1987)
Pinon-
Lataillade
et al. (1993)
Pinon-
Lataillade
et al. (1995)
Rodamilans
et al. (1998)
Ronis et al.
(1996T)
Ronis et al.
(1998a)
Species/
Strain/Age
Rat/ Albino,
(NOS), adult
Rat/Sprague-
Dawley,
90 days old
Mouse/NMRI,
adult
Mouse/BALB/c,
63 days old
Rat/Sprague-
Dawley, adult
Rat/Sprague-
Dawley, adult
Dose/Route/Form/Duration
1500, 3500, and 5500 ppm of
lead acetate in drinking water
for 18 weeks
0-0.3% lead acetate in
drinking water for 70 days
5 mg/m2 lead oxide in
aerosol for 6 hours/day,
5 days/week, 90 days
0-0.5% lead acetate in
drinking water, day 1 of
gestation until 60 days of age
0-366 mg lead acetate/L in
drinking water for 30, 60, 90,
120, 150, 180 days
0.6% lead acetate in drinking
water for various durations:
PND24-74 (pubertal
exposure); PND60-74 (post
pubertal exposure); 1 1 males
and females in pubertal
exposure group (10 each in
control pubertal group);
6 males and females post-
pubertal exposure and control
groups
0.6% lead acetate in drinking
water ad libitum for various
durations as follows: GD 5 to
PND1;GD 5 to weaning;
PND1 to weaning; 3 control
litters, 2 gestation exposure
litters, 2 lactation exposure
litters, 2 gestation and
lactation exposure litters,
2 postnatal exposure litters,
2 chronic exposure litters;
4 male and 4 female pups
per litter.
1 ml point
No overt signs of general toxicity in adult female rats, only at the end of the exposure
period the mean body weight of males exposed to two higher levels was slightly lower;
no affect of lead exposure on male fertility either after first or after second mating;
values in the pups did not differ from control group.
Decreased weight of seminal vesicles in inhalation study; no effects on spermatogenesis
(epididymal sperm count, spermatozoal motility or morphology) or plasma testosterone,
LH, and FSH; no effects on fertility; decrease in epididymal sperm count of progeny of
sires of the inhalation group, however without effect on their fertility.
No effects on testicular histology, nor on number and morphology of epididymal
spermatozoa; no effects on plasma FSH, LH, and testosterone, nor on testicular
testosterone; decreased weight of testes, epididymis, seminal vesicles, and ventral
prostate; no effects on fertility.
Reduction of intratesticular testosterone concentrations after 30 days; reduction of and
renostenedione concentrations after 150 days; no changes in intratesticular progesterone
and hydro xy-progesterone.
PbB>250 |ig/dL reduced circulating testosterone levels in male rats 40-50% (p<0.05);
reduction in male secondary sex organ weight (p<0.005); delayed vaginal opening
(p<0.0001); disrupted estrous cycle in females (50% of rats); increased incidence of
stillbirth (2% control vs. 19% Pb) (p<0.005).
Suppression of adult mean serum testosterone levels was only observed in male pups
exposed to lead continuously from GD 5 throughout life (p<0.05).
Blood Lead Concentration
(PbB)
PbB not reported
PbB 58±1.7 ug/dL (oral)
PbB 51.1±1.8 ug/dL (inhalation)
PbB <4-132 ug/dL
PbB 48-67 ug/dL
Pubertal PbB 30-60 ug/dL
Post pubertal PbB 30-60 ug/dL
Mean PbBs in male rats
30-60 ug/dL, respectively
Group: male PbB
Naive: 5.5±2.0 ug/dL
Control: 1.9±0.2 ug/dL
Gest:9.1±0.7ug/dL
Lact: 3.3±0.4 ug/dL
Gest+Lact: 16.1±2.3 ug/dL
Postnatal: 226.0±29 ug/dL
Chronic: 316.0±53 ug/dL
-------
Table AX5-4.2 (cont'd). Effect of Lead on Reproduction and Development in Mammals* Effects on Males
to
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Citation
Species/ Strain/Age Dose/Route/Form/Duration
1 ml point
Blood Lead Concentration
(PbB)
>
X
to
H
6
o
o
H
O
O
H
W
O
O
HH
H
W
Ronis et al.
(1998b)
Ronis et al.
(1998c|)
Sant'Ana
etal. (2001)
Saxena et al.
(1984)
Saxena et al.
(1986)
Saxena et al.
(1987)
Rat/Sprague-
Dawley, adult
Rat/Sprague-
Dawley, adult
Rat/Wistar, 90 days
old
Rat/ITRC, albino
(NOS), 12 weeks
old
Rat/ITRC, albino
(NOS), 40-50 g
Rat/Wistar, 40-50 g
Lead acetate in drinking
water (0.05% to 0.45% w/v);
dams exposed until weaning,
exposure of pups which
continued until PND21, 35,
55, or 85; 5 control litters
(0%), 10 low-dose litters
(0.05%), 8 mid-dose litters
(0.15%), 9 high-dose litters
(0.45%); 4 male and 4 female
pups per litter
Lead acetate 0.05, 0.15, or
0.45% in drinking water
beginning GD 5 continuing
until PND21, 35, 55, or 85;
5 control litters (0%), 10 low-
dose litters (0.05%), 8 mid-
dose litters (0.15%), 9 high-
dose litters (0.45%); 4 male
and 4 female pups per litter
0.1 and 1% lead acetate in
drinking water for 7 days
8 mg/kg lead acetate i.p. for
15 days
5, 8, or 12 mg Pb+2/kg lead
acetate i.p. for 15 days
8 mg Pb2/kg-d lead acetate
i.p. for 100 days (from
PND21 to PND120)
Dose-response reduction in birth weight (p<0.05), more pronounced in male pups;
decreased growth rates in both sexes (p<0.05) were accompanied by a statistically
significant decrease in plasma concentrations of IGF1 through puberty PND35 and 55
(p<0.05); increase in pituitary growth hormone during puberty (p<0.05).
Dose-responsive decrease in birth weight (p<0.05); dose-responsive decrease in
crown-to-rump length (p<0.05); dose-dependent delay in sexual maturity (p<0.05);
decrease in prostate weight (p<0.05); decrease in plasma concentration of testosterone
during puberty (p<0.05); decrease in plasma LH (p<0.05); elevated pituitary LH
content (p<0.05); decrease in plasma testosterone/LH ratio at high dose (p<0.05).
0.1% Pb caused decrease in testis weight, an increase in seminal vesicle weight and
no changes in plasma testosterone levels, hypothalamic VMA levels were increased
compared to control group.
Histoenzymic and histological alterations in the testes; degeneration of seminiferous
tubules; patchy areas showing marked loss in the activity of succinic dehydrogenase
and adenosine triphosphatase, whereas alkaline phosphatase activity showed only
slight inhibition.
Increasing dose of lead resulted in significant loss of body weight, as well as
testicular weight in groups 3 and 4; cholesterol in the testis of rats markedly decreased
at all given doses of lead and was statistically significant in groups 3 and 4; in
phospholipid contents, the significant decrease was observed only at two highest
doses, while at the lowest dose the decrease was not significant; activity of ATPase
remained unaffected at all three doses of lead; no significant increase in lead content
in the testis was noticed at lower dose levels as compared to control; however,
significant increase was found in groups 3 and 4 which was dose dependent.
Disturbed spermatogenesis; Leydig cell degeneration; altered enzyme activity
(G6PDH).
Mean PbB in offspring at 0.05%
(w/v) 49±6 ug/dL
Mean PbB in offspring at 0.15%
(w/v) 126±16 ug/dL
Mean PbB in offspring at 0.45%
(w/v) 263±28 ug/dL
Dams: 0, 48, 88, or 181 ug/dL
PupsPNDl: <1, 40, 83, or
120 ug/dL
Pups PND21: <1, 46, 196, or
236 ug/dL
PupsPND35:
-------
Table AX5-4.2 (cont'd). Effect of Lead on Reproduction and Development in Mammals* Effects on Males
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Citation Species/Strain/Age
Saxena et al. ITRC albino,
(1990) (NOS), adult
Singh et al. Monkey/
(1993a) Cynomolgus, birth
Birth:
300 days:
Sokol (1987) Rat/Wistar, 52 days
old
Sokol (1989) Rat/Wistar, 27 days
old
52 days old
Sokol (1990 ) Rat/Wistar, 52 days
old
Sokol and Rat/Wistar, NOS
Berman
(1991)
Dose/Route/ Form/Duration
8 mg/kg-day lead acetate for
45 days
0-1500 |lg lead acetate/kg-d
in gelatin capsules for various
durations: 3 control monkeys,
4 monkeys in infancy group
(exposure first 400 days), 5 in
post-infancy group (exposure
300 days to 9 years of age),
4 in lifetime group (exposure
from birth until 9 years)
0-0.3% lead acetate in
drinking water for 30 days
0-0.6% lead acetate in
drinking water for 30 days +
30 days recovery
0-0.6% lead acetate in
drinking water for 30 days +
30 days recovery
0-0.6% lead acetate in
drinking water for 7, 14, 30,
60 days
0, 0.1, or 0.3% lead acetate in
drinking water for 30 days
beginning at 42, 52, or 70
days old; 8-1 1 control rats
for each age, 8-11 rats for
each age in 0.1% group,
8-11 rats for each age in
0.3% group
1 ml point
Alterations in SDH, G6PDH activity, cholesterol, and ascorbic acid contents and
reduced sperm counts associated with marked pathological changes in the testis,
after combined treatment with lead and immobilization stress in comparison to
either alone.
Degeneration of seminiferous epithelium in all exposed groups (frequency not
specified); ultrastructural alterations in seminal vesicles, most prominent in infancy
and post-infancy groups (frequency not specified).
Hyper-responsiveness to stimulation with both GnRH and LH (10); blunted
response to naloxone stimulation (10).
Suppressed intratesticular sperm counts, sperm production rate, and serum
testosterone in both lead treated groups (10-10); sperm parameters and serum
testosterone normalized at the end of the recovery period in the pre-pubertal animals
(27 days at start) (10) but not in the pubertal animals (52 days at start) (5).
Decreased sperm concentration, sperm production rate and suppressed serum
testosterone concentrations after 14 days of exposure; not dose related (NS).
Dose-related suppression of spermatogenesis (decreased sperm count and sperm
production rate) in the exposed rats of the two highest age groups (p<0.05); dose-
related suppression of serum testosterone in 52-day old rats (p=0.04) and in 70-day
old rats (p<0.003).
Blood Lead Concentration
(PbB)
PbB >200 ug/dL
Chronic PbB <40-50 ug/dL
PbB 30±5 ug/dL
<3-43 ug/dL (<4-18 ug/dL after
recovery period)
Bl <3^13 ug/dL (<4-18 ug/dL
after recovery period)
Controls: <8 ug/dL at any time
exposed: 42, 60, 58, 75 ug/dL
after 7, 14, 30, and 60 days,
respectively
0% All <7 ug/dL
42 d 25 ug/dL
0.1% 52 d 35 ug/dL
70 d 37 ug/dL
49 H ^6 iio/HT
^tZ U JO f^g/Ulj
0.3% 52 d 60 ug/dL
70 d 42 ug/dL
-------
Table AX5-4.2 (cont'd). Effect of Lead on Reproduction and Development in Mammals* Effects on Males
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1
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M
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o
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Qtation
Sokol et al.
(1985)T
Sokol et al.
(1994)
Sokol et al.
(2002)
Thoreux-
Manlay et al.
(1995a)
Thoreux-
Manlay et al.
(1995b)
Wadi and
Ahmad (1999)
Wenda-
Rozewicka
et al. (1996)
Yu et al.
(1996)
*Not including
^Candidate key
Species/
Strain/Age
Rat/Wistar,
52 days old
Rat/Sprague-
Dawley, 100
days old
Rat/Sprague-
Dawley, adult
Rat/Sprague-
Dawley, 97 days
old
Rat/Sprague-
Dawley, adult
Mouse/CF-1,
adult
Rat/Wistar,
adult
Rat/Sprague-
Dawley,
neonates
Dose/Route/Form/Duration
0. 1 or 0.3% lead acetate in
drinking water for 30 days
0.3% lead acetate in drinking
water for 14, 30, or 60 days
lead acetate in water for
1 week
0-8 mg lead acetate/kg i.p.
for 5 days/week, 35 days
8 mg/kg-d lead for
5 days/week, 35 days
0.25 and 0.5% lead acetate in
drinking water for 6 weeks
1% aqueous solution of lead
acetate for 9 months
Neonatal and lactational
exposure to 0.3% lead acetate
in drinking water beginning
PNDltoPND21
1 ml point
Negative correlations between PbB levels and serum and intratesticular testosterone
values; dose-dependent reduction in intratesticular sperm count; FSH values were
suppressed; no change in LH; decrease in ventral prostatic weight; no difference in
testicular or seminal vesicle weights.
Lead exposed fertilized fewer eggs; increased duration of exposure did not result in
more significant percentage of eggs not fertilized; no ultrastructural changes were noted
in the spermatozoa of animals; no difference in histogram patterns of testicular cells.
Dose-related increase in gonadotropin-releasing hormone (GnRH) mRNA; no effect
on the serum concentrations of hypothalamic gonadotropin-releasing hormone (GnRH)
orLH.
No effects on spermatogenesis; decreased plasma and testicular testosterone by 80%;
decreased plasma LH by 32%, indications for impaired Leydig cell function, no effects
on fertility.
Germ cells and Sertoli cells were not major target of lead, accessory sex glands were
target; epididymal function was unchanged; plasma and testicular testosterone dropped
about 80%, plasma LH only dropped 32%.
Low dose significantly reduced number of sperm within epididymis; high dose reduced
both the sperm count and percentage of motile sperm and increased the percentage of
abnormal sperm within the epididymis; no significant effect on testis weight, high dose
significantly decreased the epididymis and seminal vesicles weights as well as overall
body weight gain; LH, FSH, and testosterone were not affected.
Electron microscopic studies did not reveal any ultrastructural changes in the
semiferous epithelium or in Sertoli cells; macrophages of testicular interstitial tissue
contained (electron dense) lead-loaded inclusions, usually located inside
phagolisosome-like vacuoles; x-ray micro-analysis revealed that the inclusions
contained lead.
Neonatal exposure to lead decreased cold-water swimming endurance (a standard test
for stress endurance) and delayed onset of puberty in males and female offspring, which
was exacerbated by swimming stress.
Blood Lead Concentration
(PbB)
PbB 34±3 ug/dL or PbB
60±4 ug/dL
PbB -40 ug/dL
PbB 12-28 ug/dL
PbB not reported
PbB 1700 ug/dL
PbB not reported
PbB not reported
PbB 70 ug/dL
effects on the nervous or immune systems.
study.
36-HSD, 36-hydroxysteriod dehydro;
dehydrogenase; GD, gestational day;
genase; dbcAMP, dibutyryl cyclic
adenosine-3',5'-monophosphate; DTT, dithiothreitol; FSH, follicle stimulating hormone; G6PDH, glucose-6-phosphate
GnRH, gonadotropin releasing hormone; hCG, human chorionic gonadotropin; IGFi, insulin-like growth factor 1; i.p., intraperitoneal; LDH, lactate
dehydrogenase; LH, luteinizing hormone; LHRH, luteinizing hormone releasing hormone; LPP, lipid peroxidation potential; NCD, nuclear chromatin decondensation rate; NOS, not otherwise
specified; PbB,
blood lead concentration; PND, post-natal day; p.o., per os (oral administration); ROS, reactive oxygen species; SDH, succinic acid dehydrogenase;
SOPR, sperm-oocyte penetration
rate; StAR, steroidogenic acute regulatory protein; VC, vitamin C; VE, vitamin E; VMA, vanilmandelic acid
-------
Table AX5-4.3. Effect of Lead on Reproduction and Development in Mammals* Effects on Females
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W
Citation
Burright et al.
(1989)
Coffigny et al.
(1994|)
Corpas et al.
(2002a)
Cory-Slechta
et al. (2004|)
Dearth et al.
(2002|)
Dearth et al.
(2004)
Species/
Strain/Age
Mouse/HET,
neonates
Rat/Sprague-
Dawley, adult
Rat/Wistar,
adult
Rat/Long-Evans,
adult
Rat/Fisher 344,
150-175 g
Rat/Sprague-
Dawley and
Fisher-344,
adult
Dose/Route/
Form/Duration
0.5% lead acetate solution via
milk, or drinking water
chronic beginning PND1
Inhalation exposure to 5
mg/m3 lead oxide daily for
13 days during gestation
(GD 2, 3, 6-10, 13-17, 20)
300 mg/L acetate lead in
drinking water from mating
until PND12 or PND21
Lead acetate in drinking
water beginning 2 months
before breeding through
weaning
12 mg/mL lead acetate
gavage from 30 days prior
breeding until pups were
weaned 21 day after birth;
10-32 litters per group,
control group, gestation and
lactation exposure, gestation
only exposure, lactation only
exposure
12 mg/mL lead acetate by
gavage 30 days prior to
breeding through PND21
(gestation and lactation
exposure)
Endpoint
Plasma prolactin levels implied that lead exposure alone decreased circulating prolactin
in primiparous; low prolactin levels in non-behaviorally tested females suggests that
dietary lead alone may alter plasma-hormone in these lactating HEX dams; pattern of
plasma prolactin appear to be inconsistent with the observation that lead exposure
decreases dopamine; prolactin levels of lead exposed dams were very low.
No effects on the incidence of pregnancy, prenatal death, or malformations when male
and female rats from mothers who had been exposed.
Neither abnormalities in the liver structure nor depositions of lead, toxicant produced
biochemical alterations; pups exhibited decrease in hemoglobin, iron and alkaline, and
acid phosphatase levels and an increase in Pb content; protein, DNA, and lipid total
amounts were reduced, and hepatic glycogen content was diminished at 12 and 21 PN,
with a higher dose of glucose in blood; decrease in alkaline phosphatase in liver of pups
at day 2 1 PN, but acid phosphatase was unaltered.
Observed potential effects of lead and stress in female; Pb alone (in male) and Pb plus
stress (in females) permanently elevated corticosterene levels in offspring.
Delay in onset of puberty (p<0.05); reduced serum levels of IGF1 (p<0.001), LH
(p<0.001), and E2 (p<0.001).
Lead delayed the timing of puberty in PbB 37.3 ug/dL lead group and suppressed
serum levels of LH and E2, these effects did not occur in PbB 29.9 ug/dL lead group,
when doubling dose to 29.9 ug/dL group the PbB levels rose to 62.6 ug/dL, yet no
effect was noted; results indicate that offspring are more sensitive to maternal lead
exposure with regard to puberty related insults than are 29.9 ug/dL rats.
Blood Lead Concentration
(PbB)
PbB -100 ug/dL
PbB 7 1.1 ug/dL (dam)
PbB 83.2 ug/dL (fetal)
PbB 22 ug/dL
PbB 30-40 ug/dL
Maternal PbB: -40 ug/dL
Pups PbB as follows:
Gest+lact: -38 ug/dL PND10
Gest+lact: -15 ug/dL PND21
Gest+lact: -3 ug/dL PND30
Gest: -14 ug/dL PND10
Gest: -3 ug/dLPND21
Gest: -1 ug/dL PND30
Lact: -28 ug/dL PND 10
Lact: -15 ug/dLPND21
Lact: -3 ug/dL PND30
PbB 29.9 ug/dL (Sprague-
Dawley)
PbB 37.3 ug/dL (Fisher)
-------
Table AX5-4.3 (cont'd). Effect of Lead on Reproduction and Development in Mammals* Effects on Females
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Species/
Citation Strain/Age
Foster (1992) Monkey/
Cynomolgus,
0-10 years old
Foster et al. Monkey/
(1992) Cynomolgus,
10 years old
Foster et al. Monkey/
(1996b) Cynomolgus,
15-20 years old
Franks et al. Monkey/Rhesus,
(1989) adult
Fuentes et al. Mouse/Swiss,
(1996) adult
Gorbel et al. Rat/NOS,
(2002) 3 months old
Dose/Route/
Form/Duration
Daily dosing for up to 10
years with gelatin capsules
containing lead acetate
(1.5 mg/kg); 8 control group
monkeys, 8 lifetime exposure
(birth-10 years), 8 childhood
exposure (birth-400 days),
and 8 adolescent exposure
(postnatal day 300-10 years
of age)
Daily dosing for up to
10 years with gelatin capsules
containing lead acetate
(1.5 mg/kg); 8 control group
monkeys, 8 childhood
(birth-400 days), 7 adolescent
(postnatal day 300-10 years),
7 lifetime (birth-10 years)
Chronic exposure to lead
acetate 50 to 2000 ug/kg-d
p.o. beginning at birth for
15-20 years; 20 control
monkeys, 4 monkeys in
50 ug/kg-d group, 3 monkeys
in 100 ug/kg-d, 2 monkeys in
500 ug/kg-d group, and
3 monkeys in 2000 ug/kg-d
group
Lead acetate in drinking
water (2-8 mg/kg-d) for
33 months; 7 control and
10 lead monkeys
14, 28, 56, and 112 mg/kg
lead acetate via i.p; one time
exposure on GD 9
3 mg (PI) or 6 mg (P2) lead
acetate in drinking water for
15, 30, 45, 60, or 90 days
Endpoint
Statistically significant reductions in circulating levels of LH (p<0.042), FSH
(p<0.041), and E2 (p<0.0001) during menstrual cycle; progesterone concentrations
were unchanged and menstrual cycle was not significantly affected.
No effect on endometrial response to gonadal steroids as determined by ultrasound.
Reduced corpora luteal production of progesterone (p=0.04), without alterations in E2,
20alpha-hydroxyprogesterone, or menstrual cyclicity.
Reduced circulating concentration of progesterone (p<0.05); treatment with lead did not
prevent ovulation, but produced longer and more variable menstrual cycles and shorter
menstrual flow.
Absolute placental weight at 1 12 mg/kg and relative placental weight at 14, 56, and 112
mg/kg were diminished significantly; most sections of placenta showed vascular
congestion, and increase of intracellular spaces, and deposits of hyaline material of
perivascular predominance; trophoblast hyperplasia was also observed, whereas there
was a reinforcement of the fibrovascular network in the labyrinth
Female rats absolute and relative weights of ovary and uterus were unchanged, vaginal
smears practiced in females revealed the estrus phase; fertility was found to be reduced;
lead level in blood was poorly correlated with the level of poisoning.
Blood Lead Concentration
(PbB)
PbB <40 ug/dL
PbB <40 ug/dL
PbB 10-15 ug/dL in low group
(50 or 100 ug/kg-d)
PbB 25-30 ug/dL in moderate
group (500 or 2000 ug/kg-d)
PbB 68.9±6.54 ug/dL
PbB not reported
PbB not reported
-------
Table AX5-4.3 (cont'd). Effect of Lead on Reproduction and Development in Mammals* Effects on Females
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Citation
lavicoli et al.
(2004)
Junaid et al.
(1997)
Laughlin et al.
(1987)
Logdberg
et al. (1987)
Logdberg
et al. (1998)
McGivern
etal. (1991|)
Nilsson et al.
(1991)
Piasek and
Kostial(1991)
Pinon-
Lataillade
et al. (1995)
Species/
Strain/Age
Mouse/Swiss,
33-37 days old
Mouse/Swiss,
adult
Monkey/Rhesus,
adult
Monkey/
Squirrel, adult
Monkey/
Squirrel, adult
Rat/Sprague-
Dawley, adult
Mouse/NMRI,
adult
Rat/Wistar,
10 weeks old
Mouse/NMRI,
adult
Dose/Route/
Form/Duration
0.02,0.06,0.11,0.20,2.00,
4.00, 20.00, 40.00 ppm in
food lead acetate
concentration beginning GD
1 to 3 months after birth
0, 2, 4, or 8 mg/kg-d lead
acetate, subchronic exposure,
5 days/week, 60 days
Lead acetate in drinking
water at 3. 6, 5.9, or 8.1
mg/kg-d for 1-2 years; 7
control and 10 experimental
monkeys per group
Lead acetate in drinking
water from 9th week of
gestation to PND1; per oral
exposure similar to Laughlin
et al. (1987)
Lead acetate maternal dosing
from 5-8.5 weeks pregnant to
PND1
1 1 control monkeys, 3 low-
lead exposure group (PbB
24 ug/dL), 7 medium lead
group (PbB 40 ug/dL, 5 high-
lead group (PbB 56 ug/dL)
0. 1% lead acetate in drinking
water from GD 14 to
parturition
75 ug/g bw lead chloride via
i.v.; one time injection on
gestation day 4
7500 ppm lead acetate in
drinking water for 9 weeks
0-0.5% lead acetate in
drinking water exposed to
lead during gestation until
post-GD 60
Endpoint
Increase in food consumption; however, did low-dose group increase food consumption
because of sweet nature of lead? body weight may contribute to delay in onset of
puberty and confound results.
Altered follicular development.
Reductions in cycle frequency (p<0.01); fewer days of flow (p<0.01); longer and more
variable cycle intervals (p<0.025).
Increase in pre- and perinatal mortality during the last two-thirds of pregnancy;
statistically significant reduction in mean birth weight was observed in lead exposed
monkeys as compared to controls.
Dose-dependent reduction in placental weight (p<0.0007); various pathological lesions
were seen in the placentas, including hemorrhages, hyalinization of the parenchyma
with destruction of the villi, and massive vacuolization of chorion epithelium.
Female rats showed delay in vaginal opening; 50% exhibited prolonged and irregular
periods of diestrous and lack observable corpora lutea; both sexes showed irregular
release patterns of both FSH and LH.
Electron microscopy showed that the uterine lumen, which was closed in control mice,
was opened in lead-injected mice; suggested that lead caused increase in uterine
secretion; study suggested lead could have a direct effect on the function of the uterine
epithelium and that lead was secreted into the uterine lumen and affect the blastocysts.
Decrease in litter size, pup survival, and birth weight; food consumption, body weight,
and fertility were not altered in 20 week exposure period.
Exhibited reduced fertility as evidenced by smaller litters and fewer implantation sites.
Blood Lead Concentration
(PbB)
PbB 0.69, 1.32, 1.58, 1.94,3.46,
3.80, 8.35, 13.20 ug/dL
PbB 22.3-56.5 ug/dL
PbB 44-89 ug/dL
51.2 ug/dL (low dose)
80.7 ug/dL (mid dose)
88.4 ug/dL (high dose)
Mean maternal PbB 54 ug/dL
(39-82 ug/dL)
PbB 37 ug/dL (22-82 ug/dL)
24 (22-26) ug/dL (low dose)
40 (35^16) ug/dL (mid dose)
56 (43-82) ug/dL (high dose)
PbB 73 ug/dL
PbB not reported
Maternal PbB >300 ug/dL
Offspring PbB >220 ug/dL
PbB 70 ug/dL
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Table AX5-4.3 (cont'd). Effect of Lead on Reproduction and Development in Mammals* Effects on Females
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Citation
Priya et al.
(2004)
Ronis et al.
(1996)
Ronis et al.
(1998a|)
Species/
Strain/Age
Rat/Charles
Foster,
6-9 months old
Rat/Sprague-
Dawley, various
ages
Rat/Sprague-
Dawley, adult
Dose/Route/
Form/Duration
0.03 uM lead in vitro for
1 hour
lead acetate in the drinking
water or male and female rats
for the following durations:
PND24-74 (pubertal
exposure); PND60-74
(post pubertal exposure)
0.6% lead acetate in drinking
water; ad libitum for various
durations as follows: GD 5 to
PND1, GD 5 to weaning,
Endpoint
LH binding was dropped to 84% in Pb treated cells; lead exposed cells showed 31%
reduction in the enzymes 176-HSDH and 176-HS; lead can cause a reduction in LH and
FSH binding, which significantly alters steroid production in vitro and exerts a direct
influence on granulose cell function.
Data suggest that both the temporary and the long-lasting effects of lead on
reproductive endpoints in male and female experimental animals are mediated by the
effects of lead on multiple points along the hypothalamic-pituitary-gonad axis;
exposure of male and female Sprague-Dawley rats pre-pubertally (age 24—74 days) to
lead acetate in the drinking water resulted in significant reduction in testis weight and in
the weight of secondary sex organs in males; these effects were not observed in rats
exposed post-pubertally (day 60-74); there is convincing evidence that pre-pubertal
female rats exposed in utero and during lactation have reduced levels of circulating E2
andLH.
Female pups exposed to lead from birth through adulthood or from GD 5 through
adulthood were observed to have significantly delayed vaginal opening and disrupted
estrus cycling; these effects on female reproductive physiology were not observed in
animals where lead exposure was confined only to pregnancy or lactation.
Blood Lead Concentration
(PbB)
PbB not applicable-in vitro study
Materal PbB 30-60 ug/dL
Offspring PbB >200 ug/dL.
Pups continuously exposed to
lead 225 to 325 ug/dL
Ronis et al.
(1998b)
Ronis et al.
(1998c)
Sierra and
Tiffany-
Castiglioni
(1992)
Srivastava
et al. (2004)
Rat/Sprague-
Dawley, adult
PND1 to weaning
Ad libitum intake of lead
acetate (0.05 to 0.45% w/v);
lead exposure of dams until
weaning, exposure of pups
until day 21, 35, 55, 85
Rat/Sprague-
Dawley, adult
Guinea
pig/NOS, adult
Rat/Fisher 344,
adult
0.05, 0.15, or 0.45% lead
acetate in drinking water
beginning GD 5 for 21, 35,
55, 85 days
0, 5.5, or 11 mg/kg lead
acetate, oral dose from GD 22
until GD 52 or 62
12 mg/mL lead acetate by
gavage for 30 days prior to
breeding until weaning
Prenatal lead exposure that continues until adulthood (85 days old) delays sexual
maturation in female pups in a dose-related manner; dose-dependent delay in sexual
maturation (delayed vaginal opening) among female rats following prenatal lead
exposure that continued until adulthood (85 days old); a growth hormone-mediated
effect on growth that differs depends upon the developmental state of the animal, birth
weight was significantly reduced and more pronounced among male pups; decreased
growth rates in both sexes were accompanied by a statistically significant decrease in
plasma concentrations of IGF 1 through puberty and a significant increase in pituitary
growth hormone during puberty; growth suppression of male and female rats involves
disruption of growth hormone secretion during puberty.
Dose-responsive decrease in birth weight and crown-to-rump length was observed in
litters; dose-dependent delay in sexual maturity (delay in vaginal opening); decrease in
neonatal sex steroid levels and suppression of E2 during puberty; elevation in pituitary
LH content was observed during early puberty; E2 cycle was significantly disrupted at
the highest lead dose; data suggests that the reproductive axis is particularly sensitive to
lead during specific developmental periods, resulting in delayed sexual maturation
produced by sex steroid biosynthesis.
Hypothalamic levels of SRIF; lower serum concentrations of progesterone at higher
dose only; hypothalamic levels of GnRH and SRIF were reduced in a dose-dependent
manner by lead treatment in both dams and fetuses; reduction of SRIF levels in 52-day
old fetus was particularly severe (92%) in the 11 mg group.
Lead decreased StAR protein expression and lowered E2 levels; suggested that the
primary action of Pb to suppress E2 is through its known action to suppress the serum
levels of LH and not due to decreased responsiveness of StAR synthesizing machinery.
Mean PbB in offspring at 0.05%
(w/v) 49±6 ug/dL
Mean PbB in offspring at 0.15%
(w/v) 126±16 ug/dL
Mean PbB in offspring at 0.45%
(w/v) 263±28 ug/dL
PbB in dams 181±14 ug/dL
PbB in pups ranged from 197±82
to 263±38 ug/dL, increasing with
age of pups
PbB not reported
PbB of dams 39±3.5 SEM ug/dL
and offspring PbB 2.9±0.28 SEM
Ug/dL
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Table AX5-4.3 (cont'd). Effect of Lead on Reproduction and Development in Mammals* Effects on Females
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Citation
Taupeau et al.
(2001)
Tchernitchin
et al. (1998a)
Tchernitchin
et al. (1998b)
Wide, 1985
Wide and
D'Argy (1986)
Wiebe and
Barr(1998)
Wiebe et al.
(1998)
Yu et al.
(1996)
Species/
Strain/Age
Mouse/C57blxC
BA, 8 weeks old
Rat/Sprague-
Dawley, 14 days
old
Rat/Sprague-
Dawley, 20 or
21 days old
Mouse/NMRI,
10 weeks old
Mouse/NMRI,
adult
Rat/Sprague-
Dawley, adult
Rat/Sprague-
Dawley, adult
Rat/Sprague-
Dawley, adult
Dose/Route/
Form/Duration
10 mg/kg-d lead nitrate via
i. v for 1 5 days
172 ug/g bw lead from day
14 every 2nd day until day 20
(75 mg/g bw) lead via i.v. one
time exposure at 1 or
24 before hormone
stimulation
20 ug/dL/g bw lead chloride
via i.v. single exposure on
days 8, 12, or 16 after mating
20 ug/g bw by i.v. single
injection on GD 8
20 or 200 ppm lead chloride
in drinking water; 3 exposure
durations; prior to mating
through weaning, GD 7 to
weaning, PND21 to PND35
20 or 200 ppm lead chloride
in drinking water; 4 exposure
durations; prior to mating
through weaning, GD 7 to
weaning, PND21 to PND35,
prior to mating only
Neonatal and lactational
exposure to 0.3% lead acetate
in drinking water (PND30)
Endpoint
Low lead concentration in the ovary caused dysfunction of folliculogenesis, with fewer
primordial follicles and an increase in atretic antral follicles.
Lead inhibits estrogen-induced uterine eosinophilia at 6 and 24 hours after treatment; lead
also inhibits estrogen-induced edema in deep and superficial endometrial stoma at 24
hours but not 6 hours after treatment; myometrial hypertrophy is inhibited under the
effect of exposure at 24 hours of treatment.
Enhanced some parameters of estrogen stimulation and inhibited other estrogenic
responses; interaction with responses to estrogen was different depending on whether lead
pretreatment was 1 or 24 hours before hormone stimulation; estrogenic responses mostly
affected were uterine eosinophilia, endometrial edema, uterine liminal epithelial,
hypertrophy, and mitosis in various, but not all, uterine cell types, in some cell types,
estrogen-induced mitotic response developed earlier under the effect of lead exposure.
Litter size and fetal survival varied significantly; small litters and increased numbers of
fetal deaths were observed in mice exposed to lead on day 8 of intrauterine life; live
fetuses were normal with respect to weight and morphological appearance; ovarian
follicle counts revealed a significantly smaller number of primordial follicles in the latter
group, it suggested that the exposure to lead at a time of early organogenesis caused the
observed fertility decrease by interfering with the development of the female germ cells.
Primordial germ cells showed a normal body distribution but were significantly fewer at
all four stages compared with those of control embryos of corresponding age; lead had
interfered with the production or activity of alkaline phosphatase.
Treatment with lead prior to mating resulted in significant increase in E2-receptor affinity
in 21 -day old offspring without a change in E2 receptor number; treatment from day 7 of
pregnancy until weaning of the pups resulted in approximately 35% decrease in E2
receptors per mg uterine protein when these offspring reached 150 days of age; lead
treatment from 21-35 days old or until 150 days resulted in a significant decrease in
uterine E2 receptor number at 35 and 150 day, respectively.
Exposure to lead did not affect tissue weights but did cause a significant decrease in
gonadotropin-receptor binding in the pre-pubertal, pubertal, and adult females;
conversion of progesterone to androstenedione and dihydrotestosterone was significantly
decreased in 21-day old rats and in 150-day old females; significantly increased
conversion to the 5-alpha-reduced steroids, normally high during puberty.
Neonatal exposure to lead decreased cold-water swimming endurance (a standard test for
stress endurance); delayed onset of puberty in males and female offspring, which was
exacerbated by swimming stress.
Blood Lead Concentration
(PbB)
PbB not reported
PbB 47 ug/dL
PbB not reported
PbB not reported
PbB not reported
PbB likely 4.0±1.4to 6.6±2.3
ug/dL (similar design as Wiebe
etal. (1988))
PbB range 4.0±1.4 to
6.6±2.3 ug/dL
PbB 70 ug/dL
*Not including effects on the nervous or immune systems.
^Candidate key study.
E2, estradiol; FSH, follicle stimulating hormone; GD, gestational day; GnRH, gonadotropin releasing hormone; HET, Binghamton Heterogeneous Stock; IGFi, insulin-like growth factor 1;
i.p., intraperitoneal; LH, luteinizing hormone; NOS, not otherwise specified; PbB, blood lead concentration; PND, post-natal day; p.o., per os (oral administration); SRIF, somatostatin; StAR,
steroidogenic acute regulatory protein.
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ANNEX TABLES AX5-5
May 2006 AX5-60 DRAFT-DO NOT QUOTE OR CITE
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Table AX5-5.1. In Vivo and In Vitro Studies of the Effects of Lead Exposure on Production and Metabolism of Reactive
Oxygen Species (ROS), Nitric Oxide (NO), and Soluble Guanylate Cyclease (sCG).
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Species/
Reference Tissue Age/Weight
Khalil- Male SD 200 g
Manesh et al. rats
(1994)
Gonick et al. Male SD 2 months
(1997) rats 200 g
Ding et al. Male SD 2 months
(1998) rats
Vaziri (1997) Male SD 190 g
rats
Dursun Male SD
(2005) rats
Pb Exposure
n Dosage Duration
N/A Pb-acetate, 6 months
100 ppm in
water
6 Pb-acetate, 3 months
100 ppm in
water
N/A Pb-acetate, 3 months
100 ppm in
water
12 Pb-acetate, 3 months
100 ppm in
water
24 Pb acetate 2 weeks
8 mg/kg IP
Measured Parameters
Pb Level CVS Other
7 ± 3.6 ug/d BP, tail art. ET-3, cGMP
ring response
toNE
12.4 ± 1.8 ug/dL BP cGMP, NO2 +
NO3, ET-1,
ET-3, MDA,
eNOS, iNOS
3.2 ± 0.2 ug/dL BP urine NO2 +
NO3> plasma
MDA
17±9|ig/dL BP plasma MDA
urine NO2 +
NO3
BP, RBF UrNa, Ur
N02 + N03,
24 hr UrNa
(Na+ intake
Not given )
Interventions Results
DMSA Rx Pb caused HTN, |ET3,
|U cGMP (NS) (no
effect on NE reactivity).
DMSA Rx lowered BP
and Vase response to NE
& raised cGMP
— HTN, |MDA, feNOS,
fiNOS (protein and
activity in kidney)
DMSA (0. 5% H2O) Pb caused HTN, |urine
x 2 wks, IV NO2+NO3, tplasma
infusions of L. MDA. DMSA lowered
Arg., SOD & SNP BP, blood lead & MDA
+ raised urine NO2 +
NO3 L-Arg lowered BP
and MDA, raised
N02+N03, SNP lowered
BP
Antioxidant Rx Pb caused HTN, fMDA,
(Lazaroid) jurine NO2+NO3 in
untreated animals.
Antioxidant Rx improved
HTN, urine NO2+NO3
and lowered MDA
without changes in blood
Pb level
|BP, |RBF, |UrN02 +
NO3, unchanged UrNa+
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Table AX5-5.1 (cont'd). In Vivo and In Vitro Studies of the Effects of Lead Exposure on Production and Metabolism of
Reactive Oxygen Species (ROS), Nitric Oxide (NO), and Soluble Guanylate Cyclease (sCG).
X
4 per 0 and 1 ppm 24 hrs w/Pb 1 ppm medium
Ding (2001) coronary experiment lead acetate or Na acetate
endothelial followed
cells by 24 hrs
w/tempol or
vehicle
Measured Parameters
CVS Other Interventions
BP Aorta & kidney Subgroups
eNOS protein treated with high-
abundance, Ur dose vitamin E
N02 + N0
BP Aorta, heart, Subgroups
kidney & brain studied after 2
NOS isoforms, wksofRx
urine NO2 + w/tempol and
NO3 those studied 2
wks after
cessation of
tempol Rx
N/A eNOS Co-treatment
expression w/O2' scavenger,
tempol
Results
Pb exposure resulted in a
time-dependent rise in
BP, aorta & kidney
eNOS&iNOS. This
was associated w/a
paradoxical fall in NO
availability (Ur NO2 ±
NO3). Antioxidant Rx
attenuated upregulation
ofiNOS&eNOS&
raised NO availability.
Pb exposure resulted in
rises in BP, eNOS, iNOS
& nNOS in the tested
tissues + jurine NOX.
Tempol administration
attenuated HTN, reduced
NOS expressions &
increased urine NOX.
The effects of tempol
disappeared within 2
weeks of its
discontinuation.
Pb exposure for 48 hours
upregulated eNOS
expression. Co-treatment
w/ tempol resulted in
dose-dependent reversal
of Pb-induced
upregulation of eNOS
but had no effect on
control cells.
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Table AX5-5.1 (cont'd). In Vivo and In Vitro Studies of the Effects of Lead Exposure on Production and Metabolism of
Reactive Oxygen Species (ROS), Nitric Oxide (NO), and Soluble Guanylate Cyclease (sCG).
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Pb Exposure
Species/
Reference Tissue Age/W eight n Dosage Duration Pb Level
Vazirietal. Male SD rats 200 g 6 per lOOppmin 3 months 8.3 - 10.8 ug/g
(1999) group water kidney tissue
Vaziri et al. Male SD rats 200 g 6 per 100 ppm in 3 months N/A
(2003) group water
Nietal. Cultured N/A >4per 0,1 & 10 ppm short 0,1, 10 ppm
(2004) human experime Pb acetate exposure
coronary nt (5-30 min) &
endothelial & long exposure
VSM cells. (60 hours)
Measured Parameters
CVS Other Interventions
BP Urine NO2 + Antioxidant Rx
NO3, tissue and (Vit E)
plasma
nitrotyrosine
(marker of
NO-ROS
interaction).
BP Urine NO2 + Tempol (O2'
NO3, kidney, scavenger
heart, brain infusion)
SOD, catalase,
GPX, NAD(P)H
oxidase
abundance
N/A O2' and H2O2 None
productions
SOD, catalase,
GPX&
NAD(P)H
oxidase
(gp91phox)
Results
Pb exposure raised BP,
reduced Ur NO2 + NO3
& increased nitrotyrosine
abundance in plasma,
heart, kidney, brain &
liver. Anti ox Rx
ameliorated HTN,
lowered nitrotyrosine &
raised Ur NO2 + NO3.
Pb caused HTN,
|NAD(P)H oxidase
(gp91phox), fSOD, un-
changed catalase and
GPX, |UrNO2 + NO3.
Tempol resulted in |BP
+ furine NO2 + NO3 in
lead-exposed but not
control rats.
Short-term incubation
with Pb at 1 & 10 ppm
raised O2'& H2O2
productions by both
endothelial & VSM cells,
long-term incubation
resulted in further rise in
H2O2 generation &
normalization of
detectable O2y . This was
associated with increases
in NAD(P)H oxidase &
SOD & reduced or
unchanged catalase &
GPX.
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Table AX5-5.1 (cont'd). In Vivo and In Vitro Studies of the Effects of Lead Exposure on Production and Metabolism of
Reactive Oxygen Species (ROS), Nitric Oxide (NO), and Soluble Guanylate Cyclease (sCG).
X
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Species/
Reference Tissue
Ding et al. Male SD rats
(2001)
Ding et al. Cultured rat
(2000) aorta
endothelial
cells
Attri (2003) Male Wistar
Kyoto rats
Malvezzi Male Wistar
(2001) rats
Pb Exposure
Age/Weight n Dosage Duration Pb Level
2 months N/A 100 ppm 3 months Blood lead 12.4
200 g ±1.8ng/dLvs.
1 mg/dL in
controls
N/A > 4 0-1 ppm 1, 2, 24, 84 0-1 ppm culture
experiment hours media
150-200 g 10 per Pb acetate, 1-3 months Blood Pb
group 100 ppm in 1.5 mg/dL at
water ± 1 mo
Vit C 20
mg/day/rat 2.4 mg/dL at
2 mo
4. 1 mg/dL at
3 mo
5-6 wks 4-10 per Pb acetate 100 days Blood, bone,
(170 g) group 750 ppm in kidney, aorta,
water liver
Measured Parameters
CVS Other Interventions
BP Response to IV infusion of
DMTU DMTU
administration,
tissue
nitrotyrosine,
hydro xyl radical
N/A Hydroxyl radical None
production using
the following
reaction (Na
Salicylate + OH
— > 2,3dihydroxy
benzoic acid),
MDA
BP Total antioxidant Response to
capacity, ferric- vitamin C.
reducing
antioxidant
power, NO
metabolites,
MDA,
8-hydroxy guano
sine
BP — Response to L.
arg, DMSA, L.
arg .+ DMSA
(given together
w/Pb in last 30
days
Results
Pb caused HTN, fplasma
nitrotyrosine,
tplasma.OH
concentration all reversed
with .OH-scavenger,
DMTU infusion
Pb exposure resulted in
cone-dependent rise in
MDA and . OH
production by cultured
endothelial cells.
Pb caused |BP, |MDA,
|DNA damage/
oxidation, |NOX,
jantioxidant. and ferric-
reducing antioxidant.
Concomitant Rx with Vit-
C ameliorated all
abnormalities.
|BP w/lead, partial |BP
w/L. Arg or DMSA,
greater reduction w/both,
blood and aorta PB
remained | in all but
DMSA + L. Arg group.
Significant Pb
mobilization shown in
other organs.
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Table AX5-5.1 (cont'd). In Vivo and In Vitro Studies of the Effects of Lead Exposure on Production and Metabolism of
Reactive Oxygen Species (ROS), Nitric Oxide (NO), and Soluble Guanylate Cyclease (sCG).
X
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Species/ Age/
Reference Tissue Weight
Khalil- Male SD 8 wks
Manesh et al. rats
(1993)
Marques et al. Male 3 months
(2001) Wistarrats
Farmland Male SD 200 g
et al. (2005) rats
Courtois et al. Rat N/A
(2003) thoracic
aorta
Pb Exposure Measured Parameters
n Dosage Duration Pb Level CVS Other Interventions
N/A Pb acetate 100 or 1-12 months 29±4ug/dL BP, vascular cGMP, ET-3, ANP —
5000 ppm in contractility to
water NE in vitro
20 Pb acetate 5 ppm 30 day N/A BP, arch-, SNP- sGC protein mRNA CoTx with Vit C
± Vit C (3 vasorelaxation & activity.
mmol/L) in response in aorta cGMP production,
water rings eNOS protein
8 Pb acetate 100 3 months N/A BP Aorta sGC, SOD, —
ppm in water catalase,
glutathione
peroxidase
6/experiment 0-1 ppm 24 hr 0-1 ppm cGMP sGC expression, Vit C, COX-2
production superoxide inhibitor
production, COX-2
Results
Pb caused HTN,
jserum and urine
cGMP, tserum ET-3
without changing ANP
or response to NE
Pb caused HTN,
^relaxation to Ach &
SNP, feNOS, |sGC
protein mRNA and
activity. These
abnormalities were
prevented by
antioxidant Rx.
|sGC, fCuZn SOD
activity, unchanged
catalase & GPX
activities.
Pb caused |sGC,
|cGMP, tO2, TCOX-2.
All abnormalities
improved by Vit C.
COX-2 inhibitor
improved sGC
expression but not O2
production.
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Table AX5-5.2. Studies of the Effects of Lead Exposure on PKC Activity, NFkB Activation, and Apoptosis
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X
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Table AX5-5.3. Studies of the Effects of Lead Exposure on Blood Pressure and Adrenergic System
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Pb Exposure Measured Parameters
Species/ Age/
Reference Tissue Weight n
Chang etal. Wistarrats 190-200 g 20
(1997)
Tsaoetal. Wistarrats 190-200 g 70
(2000)
Carmignani Male SD rats 3 mo 24
et al. (2000)
Dosage
Pb acetate
0.5% in
drinking
water
Pb acetate
0-2% in
drinking
water
60 ppm
Duration Pb Level CVS
2 months blood 29. 1± BP
1.9 ug/dL
aorta; 1.9±
0.2 Mg/g
2 months blood, heart, BP, p agonist-
aorta, kidney stimulated
cAMP
production (10
uM
isoproterenol
in vitro)
10 months Blood 22. 8 ± BP, HR,
1.2 ug/dL cardiac
contractility
(dP/dt), blood
flow
Other
Plasma
catecholamines +
aorta; p receptor
binding assay &
cAMP generation
pi NEpi, cAMP p
receptro densities
Plasma NE, Epi,
dopamine, monoamine
oxidase (MAO)
activity, histology
Reference Species/Tissue
— Pb exposure caused
HTN, elevated plasma
NE (unchanged
plasma Epi).
I isoproterenol-
stimulated plasma
cAMP, I p receptor
density in aorta.
— Pb exposure raised BP
and pi NE + lowered
aorta and heart p
receptor density, basal
and stimulated cAMP
productions +
increased kidney p
receptor density and
basal and stimulated
cAMP productions.
— Pb exposure raised BP
and dp/dt, lowered
carotid blood flow, (no
change in HR) raised
plasma NE and Epi
and MAO (all tissues)
lowered plasma NOx
+ 4 aorta media
thickness,
| lymphocyte
infiltration in
periaortic fat,
nonspecific change in
kidney (congestion,
edema, rare prox.
tubular cell necrosis).
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Table AX5-5.3 (cont'd). Studies of the Effects of Lead Exposure on Blood Pressure and Adrenergic System
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Species/ Age/
Reference Tissue Weight n
Lai et al. Male SD rats 300 g Acute
(2002) response
Chang et al. Male Wistar 10 wks 70
(2005) rats
Pb Exposure
Dosage Duration Pb Level
In vivo: — —
Intrathecal
injection of
PbC12, 10-100
uM.
In vitro:
Thoracic cord
slices exposed
to 5-50 uM
PbC12
2% Pb acetate 2 mo, blood:
(drinking observed for 7 85 ug/dL
water) mo after
cessation aorta:
8 n/g/g
heart:
1 ug/g
kidney:
60 ug/g
Measured Parameters
Intervention
CVS Other s Results
BP, HR, (In vivo) Electophysiologic — In vivo: IT injection
w/without measures (In vitro) of PbC12 raised BP
ganglionic blockade before/after saline and HR. This was
(Hexomethonium) washout reversed by ganglionic
blockade. In vitro: Pb
raised excitatory &
lowered inhibitory
postsynaptic potentials
which were reversed
by removal of lead
(saline washout)
BP Plasma NE, p Cessation of Pb exposure raised
recaptor density Pb exposure BP, plasma NE, &
(aorta, heart, renal tissue p receptor
kidney) & lowered aorta/heart
p receptor density.
Plasma and tissue lead
fell to near-control
values within 7 mo.
after Pb cessation.
This was associated
with significant
reductions (not
normalization) of BP,
plasma NE and partial
correction of tissue p
receptor densities
(Bone lead was not
measured).
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Table AX5-5.4. Studies of the Effects of Lead Exposure on Renin-angiotensin System, Kallikrein-Kinin System,
Prostaglandins, Endothelin, and Atrial Natriuretic Peptide (ANP)
X
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Table AX5-5.5. Studies of Effect of Lead on Vascular Contractility
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Pb Exposure
Species/ Age/
Reference Tissue Weight n Dosage Duration Pb Level
Shelkovnikov Rat aorta rings Pb acetate short —
andGonick 10"8to W4 incubations
(2001)
Purdyetal. Male SD rats 8 weeks Pb acetate 100 3 months —
(1997) ppm in water
Oishi (1996) Male Wistar Pb acetate 1-3 months
rats
Valencia et al. Wistar rat 7 weeks 6 sets/ Pb acetate rapid response —
(2001) thoracic aorta experiment 0.1-3.1mM in vitro
rings
Measured Parameters
CVS Other
Vasoconstriction/
vasodilation
BP Aorta ring
response to NE,
phenylephrine,
acetylcholine, and
nitroprusside
Mesenteric art &
aorta response to
acetylcholine in
presence or
absence of NOS
inhibitor (L-
NAME)
In vitro
contractile
response
Interventions Results
Lead acetated did not
cause Vasoconstriction &
did not modify the
response to NE,
isoproterenol, phorbol
ester or acetylcholine but
raised contractile
response to submaximal
Ca2+ concentration
Pb exposure raised BP.
Aorta ring
vasoconstrictive response
to NE & phenylephrine
& vasodilatory response
to acetylcholine &
nitroprusside were
unchanged.
Vasorelaxation response
to acetylcholine in
presence of L-NAME
was significantly reduced
in mesenteric art, but not
aorta of lead-exposed
animals (Inhibition of
hyperpolarizing factor)
Lead induced a
concentration-dependent
Vasoconstriction in intact
& endothelial-denuded
rings in presence or
absence of a- 1 blacker,
PKC inhibitor, L. type
Ca + channel blocker or
intra- & extracellular
Ca2+ depletion.
However, the response
was abrogated by
lanthanum (a general Ca
channel blocker)
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Table AX5-5.6. Effects of Lead on Cultured Endothelial Cell Proliferation, Angiogenesis, and
Production of Heparan Sulfate Proteoglycans and tPA
X
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Reference
Kaji et al.
(1995)
Kaji et al.
(1995)
Fujiwara et al.
(1998)
Kishimoto
etal. (1995)
UedaD.,
etal. (1997)
Pb Exposure
Species/ Age/
Tissue Weight n Dosage Duration Pb Level
Bovine aorta — 4 sets per Pb nitrate 24 hrs
endothelial experiment 5-50 uM
cells
— 4-5 sets per Pb nitrate 24 hrs
experiment 0.5-5 uM
Bovine aorta — 6 set per Pb nitrate 48 hrs
endothelial experiment 5 and 10 uM
cells
Human- — 3 sets per Pb acetate 24 hrs
umbilical vein experiment 1-100 //M
endothelia
cells
Human — 3 sets per Pb acetate 24 hrs
umbilical vein experiment 1-100 uM
endothelial
cells
Measured Parameters
CVS Other
— endothelial
damage
— 3H-thymidine
incorporation, cell
count,
morphology,
LDH release
— appearance of
cells in denuded
areas of
monolayer, DNA
synth
— formation of tube-
like structures
(angio-genesis
assay, on Matrigel
(BM)
— tube formation on
Matrigel matrix
Interventions Results
Co-incubation Addition of Pb alone
with cadmium resulted in mild
deendothelialization of
the monolayers &
markedly increased
cadmium-associated
endothelial damage.
stimulation Incubation w/Pb resulted
w/pFGF & in a concentration-
aFGF dependent reduction of
DNA synthesis & cell
count, caused some shape
change (polygonal
— >spindle) & reduced
pFGF- and aFGF-
mediated proliferation.
stimulation Pb inhibited appearance
w/Zn of endothelial cells in the
denuded section of
monolayer & attenuated
the healing response to
Zinc
— Lead inhibited tube
formation concentration-
dependently & tube
lengthening time
dependently.
PKC activator Lead inhibited tube
and inhibitor formation concentration-
dependently & tube
lengthening time
dependently. These
effects were independent
of PKC.
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Table AX5-5.6 (cont'd). Effects of Lead on Cultured Endothelial Cell Proliferation, Angiogenesis, and
Production of Heparan Sulfate Proteoglycans and tPA
X
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Fujiwara &
Kaji (1999)
Kaji et al.
(1992)
Table AX5-5.6 (cont'd). Effects of Lead on Cultured Endothelial Cell Proliferation, Angiogenesis, and
Production of Heparan Sulfate Proteoglycans and tPA
Pb Exposure
Measured Parameters
Reference
Species/
Tissue
Age/
Weight
n
Dosage
Duration
Pb Level
CVS
Other
Interventions
Results
Bovine aorta
endothelial
cells (growing
10% BCS)
Human
umbilical vein
endothelial
cells
(confluent)
4 sets per
experiment
Pb nitrate 0.1-
uM
48hrs
5 sets per
experiment
0.01-1 uM
Sulfate & glucosamine
incorporation in GAGs,
quantification of high
& low MW-HSPG,
identification of
perlecan core protein
t-PA release, DNA
synth, protein synth
(leucine incorporation)
Thrombin and
ET-1
stimulations
In growing cells, Pb
depressed high-MW
HSPGs production but
had little effect on low-
MW HSPGx (-50 KD).
The core protein of
perlecan (400 KD) was
significantly reduced by
Pb exposure.
Lead exposure reduced
basal & thrombin-
stimulation t-PA release
& worsened ET-1
induced inhibition of t-
PA release.
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Table AX5-5.7. Studies of the Effect of Lead on Cultured Vascular Smooth Muscle Cells
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Reference
Fugiwara
etal. (1995)
Corsia RV
(1995)
Yamamoto C
(1997)
Pb Exposure
Species/ Age/
Tissue Weight n Dosage Duration Pb Level
Bovine aorta 4 sets per lead nitrate 24 hrs
vascular experiment 0.5-10 uM
smooth
muscle cell
rat aorta > 3 sets per lead citrate time to
VSMC cells experiment 100 & 500 confluence
(80-90% ug/L (-90% for
confluent) control
experiments)
Human aorta 5 sets per lead chloride 24 hrs
VSMC& experiment 0.5-10 uM
fetal lung
fibroblasts
(confluent)
Measured Parameters
CVS Other
— DNA synthesis
cell density (cell
#/Cm2), cell
morphology,
membrance lipid
analysis, receptor
densities (Ang-II,
a, P, ANP
t-PA&PAI-l
release
Reference Species/Tissue
Coincubation Pb caused a
w/pFGF, concentration-dependent
aFGF. pDGF increase in DNA
synthesis. Co-incubation
w/pFGF & Pb resulted in
an additive stimulation of
VSMC DNA synth.
However, Pb inhibited
PDGF & aFGF-induced
DNA synthesis.
At low concentration, Pb
caused VSMC
hyperplasia, phenotypic
transformation from
spindle-to-cobblestone
(neointima-like) shape,
reduced Ang II receptor
density without changing
a, P, ANP receptors,
increased arachidonic
acid content of cell
membrane.
At 2 uM or higher
concentrations, lead
resulted in a
concentration-dependent
decline in t-PA release in
both cell types. Lead
increased P AI- 1 release
in fibroblasts but lowered
PAI-linVSMC.
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ANNEX TABLES AX5-6
May 2006 AX5-75 DRAFT-DO NOT QUOTE OR CITE
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Table AX5-6.1. Genotoxic/Carcinogenic Effects of Lead - Laboratory Animal Studies
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Compound
Dose and Duration
Cell Type
Co-exposure
Effects
Reference
X
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Table AX5-6.1 (cont'd). Genotoxic/Carcinogenic Effects of Lead - Laboratory Animal Studies
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Compound
Dose and Duration
Cell Type
Co-exposure
Effects
Reference
Lead Acetate 50 or 1000 ppm 50 or 1000 ppm
given in drinking water for 280
days.
Number of mice pre group in initial
exposures not given.
Number of mice at end were 50 per
dose.
Female albino Swiss
Mice - 8 weeks old
None Mice have high rate of spontaneous leukemia from endemic viral Blakley
infection. (1987)
No signs of lead poisoning. No lead effects on growth or weight
gain.
Lead did increase leukemia-related mortality possibly due to
immunosuppression.
Lead levels did increase in tissues.
Data indicate that lead may be immunosuppressive, though the
exact mechanism is not understood.
X
Lead Acetate 60 mg/kg injected s.c. weekly for 5
weeks followed by observation for
80 weeks. 13 treated and 14 control
rats.
Fisher F344/NSle rats -
3 weeks old
None Lead induced tumors at the site of injection in 42% of rats Teraki and
though data was not shown. Uchiumi (1990)
Control data not indicated or shown.
Lead accumulated in tumor tissue, tooth, and bone. This data
was shown.
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Lead Acetate 1 or 100 ug/L given in the drinking Male Wistar Rats -
water for 31 weeks weanlings
8 animals per group
0.2-4 % calcium
carbonate given in the
diet for 31 weeks.
No differences in drinking water or food consumption.
High lead and high calcium reduced growth.
No deaths in low calcium groups.
10/24 rats from high calcium diet died (4 from controls and 3
each from lead groups). All 10 had kidney or bladder stones.
0/8 rats in low calcium no lead had kidney pathology
2/8 rats in low calcium low lead had nephrocalcinosis.
7/8 rats in low calcium high lead had nephrocalcinosis.
3/4 rats in high calcium no lead had nephrocalcinosis.
1/5 rats in high calcium low lead had a renal pelvic
carcinoma. 3/5 rats had nephrocalcinosis.
3/5 rats in high calcium high lead had transitional cell
hyperplasia. 2/5 rats had invasive renal pelvic carcinoma.
Lead tissue levels were same regardless of dietary calcium
levels.
Bogden et al.
(1990)
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Table AX5-6.2. Genotoxic/Carcinogenic Effects of Lead - Human Cell Cultures
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Compound
Lead
Chromate
Lead
Chromate
Lead
Chromate
Dose and Duration
Anchorage Independence
(0.1-1 uMfor48h)
Anchorage Independence
(0.1-1 uMfor48h)
Morphological Transformation
(2 ug/mL for 24 h, performed 3
Cell Type
Human Foreskin
Fibroblasts
InH-MEM+15%FCS
Human Foreskin
Fibroblasts
InH-MEM+15%FCS
HOS TE 85 in DMEM
+ 10% FBS
Co-exposure Effects
None Lead chromate-induced concentration-dependent increase in
anchorage independence.
None Lead chromate-induced concentration-dependent increase in
anchorage independence.
None Lead chromate induced foci of morphological
transformation after repeated exposure and passaging.
Reference
Biedermann and
Landolph (1987)
Biedermann and
Landolph (1990)
Sidhu et al. (1991)
X
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Lead Acetate
times immediately after passage)
Anchorage Independence (0.2-2
ug/mL or cells isolated during
morphological transformation)
Neoplastic Transformation (cells
isolated during morphological
transformation)
Anchorage Independence
(500-2000 uM for 24 h)
Human Foreskin
Fibroblasts (Chinese)
In DMEM +10% FCS
3-aminotriazole (3-AT)
(80 mM to inhibit
catalase)
Lead chromate did not induce anchorage independence, but
cells from the foci obtained during morphological
transformation.
Lead chromate did not induce neoplastic transformation in
the cells from the foci obtained during morphological
transformation.
Studied as a chromate compound. Role of lead not
mentioned or considered.
Lead acetate-induced concentration-dependent increase in
anchorage independence. Anchorage independence not
affected by 3-AT.
Hwua and Yang
(1998)
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Table AX5-6.3. Genotoxic/Carcinogenic Effects of Lead - Carcinogenesis Animal Cell Cultures
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Compound
Lead Acetate
Lead Chloride
Lead
Chromate
Lead
Chromate
Lead
Chromate (and
pigments
containing
lead chromate)
Lead
Chromate
Dose and Duration
Morphological Transformation
(10-50 uM for 48 h)
Morphological Transformation
(doses not given)
Enhancement of Simian
Adenovirus (SA7) induced
morphological transformation.
(80- 1,240 uM for 20 h)
Morphological Transformation
(10-50 uM for 24 h)
Anchorage Independence for cells
isolated during morphological
transformation
Neoplastic Transformation for cells
isolated during morphological
transformation.
Morphological Transformation
(0.04 - 8 ug/mL as Cr for 7 days)
Anchorage Independence for cells
isolated during morphological
transformation
Neoplastic Transformation for cells
isolated during morphological
transformation
Morphological Transformation
(0.02 - 0.88 ug/mL as Cr for 7
days)
Cell Type
Primary SHE cells in
AMEM + 10% FBS
C3H10T1/2 cells in
EMEM +10% FBS
Primary SHE cells in
DMEM + 10% FBS
C3H10T1/2 cells in
EMEM + 10% FBS
Primary SHE cells in
DMEM + 10% FCS
Primary SHE cells in
DMEM + 10% FCS
Co-exposure Effects
None Lead acetate was weakly positive inducing a 0. 19-1.6%
increase in transformation. There was a weak dose
response. There were no statistical analyses of these data.
None Lead chloride did not induce morphological transformation.
None Lead chromate enhanced SA7-induced morphological
transformation.
Studied as a chromate compound. Role of lead not
mentioned or considered.
None Lead chromate induced morphological and neoplastic
transformation.
Cells exhibiting morphological transformation grew in soft
agar and grew in nude mice.
Studied as a chromate compound.
None Lead chromate induced morphological and neoplastic
transformation.
Cells exhibiting morphological transformation grew in soft
agar and grew in nude mice.
Studied as a chromate compound.
None Lead chromate induced morphological transformation more
potently (9-fold) than other chromate compounds.
Reference
Zelikoffetal. (1988)
Patierno et al. (1988),
and Patierno and
Landolph (1989)
(both papers present
the same data)
Schechtman et al.
(1986)
Patierno et al. (1988)
and Patierno and
Landolph (1989)
(both papers present
the same data)
Elias et al. (1989)
Elias et al. (1991)
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Table AX5-6.3 (cont'd). Genotoxic/Carcinogenic Effects of Lead - Carcinogenesis Animal Cell Cultures
Compound
Lead Nitrate
Dose and Duration
Morphological Transformation
(0.04 - 8 ng/mL as Cr for 7 days)
Cell Type
Primary SHE cells in
DMEM + 10% PCS
Co-exposure
Calcium chromate
Effects
Lead nitrate alone did not induce significant levels of
transformation.
Reference
Elias et al. (1991)
Lead nitrate plus calcium chromate increased the potency
of calcium chromate to that of lead chromate. Data suggest
lead ions are synergistic with chromate ions in inducing
neoplastic transformation.
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Abbreviations:
Cells
SHE = Syrian hamster embryo;
C3H10T/12 cells are a mouse embryo cell line
Medium and Components
AMEM = Alpha Minimal Essential Medium;
DMEM = Dulbecco's Minimal Essential Medium;
EMEM = Eagle's Minimal Essential Medium;
FBS = Fetal Bovine Serum
ECS = Fetal Calf Serum
H-MEM = Minimum essential medium/nutrient mixture-F12-Ham
HOS TE = Human osteosarcoma cell line TE
Differences between the serum are unclear as insufficient details are provided by authors to distinguish.
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Table AX5-6.4. Genotoxic/Carcinogenic Effects of Lead - Genotoxicity Laboratory Animal Studies.
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Compound
Dose and Duration
Species
Co-exposure
Effects
Reference
Lead Acetate
2.5 mg/100 g given i.p. as
daily injection for 5-15
days
10-20 mg/100 g given i.p. a
single injection and animals
studied after 15 days
5 animals per group.
Chromosome damage in
bone marrow
Female Norway Rat
Selenium (0.012-0.047
mg/lOOg or 0.094-0.188
mg/100 g given i.p.
with lead)
Lead induced chromosome damage after chronic treatment.
It was not dose dependent as only 1 dose was studied. The
effects of selenium on lead effects are unclear as selenium
alone induced substantial chromosome damage.
The single dose exposure also induced chromosome damage,
but untreated controls were not done in this regimen. There
is some mention that this dose regimen is toxic to the animals
as selenium modulated the lethal effects, but no explanation
of how many animals died.
Chakraborty
etal. (1987)
Lead Acetate
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Table AX5-6.4 (cont'd). Genotoxic/Carcinogenic Effects of Lead - Genotoxicity Laboratory Animal Studies.
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Compound
Lead Nitrate
Lead Nitrate
Lead Nitrate
Dose and Duration Species
100-200 mg/kg given iv on ICR Swiss Webster Mice
9th day of gestation onwards - 6-8 week old
for 9 days.
Mothers and fetuses
analyzed on G18.
Group size not given.
Resorptions, fetal viability
and chromosome damage,
SCE and NOR in the
mother and fetus were
examined. Mother -bone
marrow; fetus liver or lung
3 mothers and fetuses per
dose were analyzed.
10, 20, or 40 mg/kg given Swiss Albino Mice -
i.p. 24 h 8 weeks old
6 animals per group.
Chromosome damage and
Mitotic Index in bone
marrow
5, 10, or 20 mg/kg given Swiss Albino Mice -
i.p. 24 h 8 weeks old
6 animals per group.
Chromosome damage in
bone marrow.
50 metaphases per animal
for a total of 300 (X6).
Co-exposure
None
Phyllanthus fruit extract
(685 mg/kg) or ascorbic
acid (16.66 mg/kg)
given by gavage for
7 days
Ferric chloride
(18 mg/kg) given i.p.
for 24 h administered
1 h
before-, 1 h after- or
together with- lead
nitrate
Effects
Lead levels were found in both mother and fetus indicating
no problems crossing the placenta.
All doses indicated increased resorption and decreased
placental weights. No effects on fetal weight.
Significant increase in SCE in mothers at 150 and 200 mg/kg.
No increase in SCE in fetuses.
Significant decrease in NOR in both mother and fetuses.
No gaps or breaks in mothers or fetuses.
Some weak aneuploidy at lowest dose.
Some karyotypic chromosome damage was seen.
No explanation of how many cells analyzed per animal
(3 animals per dose were analyzed) as only 20- 40 cells were
analyzed.
There was no dose response and no statistical analyses for
chromosome damage. No details on how many animals
analyzed for metaphase damage or how many cells per
animal.
Data interpretation is also complicated as too few metaphases
were analyzed 10-25 for SCE. Not given for CA.
No detail on potential maternal toxicity.
Lead nitrate increased the amount of chromosome damage at
each dose. But there was no dose response and a similar
level of damage was seen for each dose.
Phyllanthus fruit extract reduced the amount of damage at
each dose. Ascorbic acid reduced the damage at the lowest
dose but increased it at the higher doses.
Higher concentrations of lead nitrate reduced the mitotic
index. This effect was reversed by ascorbate and Phyllanthus
only at the moderate dose.
Lead nitrate increased the amount of chromosome damage in
a dose-dependent manner.
Iron exhibited some modifications of lead induced damage:
If administered 1 h before lead plus simultaneously it reduced
the damage. If administered with lead only at same time it
reduced damage in the lower doses. If lead was started 1 h
before iron there was no effect.
Thus iron may antagonize lead perhaps by blocking uptake.
Reference
Nayaketal. (1989b)
Dhiretal. (1990)
Dhiretal. (1992)a
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Table AX5-6.4 (cont'd). Genotoxic/Carcinogenic Effects of Lead - Genotoxicity Laboratory Animal Studies.
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Compound
Lead Nitrate
Lead Nitrate
Lead Nitrate
Lead Nitrate
Lead Nitrate
Dose and Duration
5 or 10 mg/kg given by
gavage for 24 h
6 animals per group.
Chromosome aberrations in
bone marrow
10, or 20 mg/kg given i.p.
48 h
12 animals per group.
Micronucleus formation in
bone marrow
10, 20, or 40 mg/kg given
i.p. 24 h
5 animals per group.
SCE in bone marrow
0.625-80 mg/kg given i.p.
for 12, 24 or 36 h.
12 animals per group
Micronucleus formation in
bone marrow. 4000
erythorocytes scored per
animal
0.7-89.6 mg/kg given by
gavage for 24, 48, or 72 h,
or 1 or 2 weeks.
5 animals per group.
Cell viability by trypan blue
Single strand breaks in
white blood cells
Species
Swiss Albino Mice -
7-8 weeks old
Swiss Albino Mice -
6 weeks old
Swiss Albino Mice -
6-8 weeks old
Swiss Albino Mice -
6-8 weeks old
Swiss Albino Mice -
4 weeks old
Co-exposure
Zirconium oxychloride
(110 or 220 mg/kg)
given by gavage for
24 h administered 2 h
before-, 2 h after- or
together with- lead
nitrate
Phyllanthus fruit extract
(685 mg/kg) or ascorbic
acid (16.66 mg/kg)
given by gavage for 7
days
Phyllanthus fruit extract
(685 mg/kg) or ascorbic
acid (16.66 mg/kg)
given by gavage for
/ days
None
None
Effects
Lead nitrate increased the amount of chromosome damage in
a dose-associated manner.
Zirconium induced a dose-associated increase in
chromosome damage.
Zirconium exhibited minimal modification of lead nitrate-
induced damage when administered 2 h before or after lead
nitrate.
Administering the two together increased the damage.
Lead nitrate increased the amount of micronuclei at both
doses in a dose-associated manner. The 48 h recovery time
was lower than 24 h but still elevated.
Phyllanthus fruit extract reduced the amount of damage at
both doses. Ascorbic acid reduced the damage at the lowest
dose but increased it at the higher dose.
Lead nitrate increased the amount of SCE in a dose-
dependent manner. Lead nitrate had no effect of the
proliferative rate index (consideration of metaphases in
different division numbers)
Phyllanthus fruit extract and ascorbic acid reduced the
amount of damage at each dose.
Lead nitrate induced micronuclei but they did not increase
with dose.
Lead induced more micronuclei in males than in females.
The ratio of polychromatic to normochromatic erythrocytes
was elevated in lead nitrate treated cells, but again did not
increase with dose.
Viability was high (92-96%) at all doses.
Lead nitrate induced single strand breaks but they did not
increase with dose. In fact the 3 highest doses were all
similar in magnitude and less than the 5 lowest doses. The 5
lowest doses were also similar in magnitude.
Reference
Dhiretal. (1992)b
Kumar et al. (1990)
Dhiretal. (1993)
Jagetia and Aruna
(1998)
Devi et al. (2000)
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to
o
o
Table AX5-6.4 (cont'd). Genotoxic/Carcinogenic Effects of Lead - Genotoxicity Laboratory Animal Studies.
Compound
Dose and Duration
Species
Co-exposure
Effects
Reference
Lead Acetate
10 mg/kg given by gavage
5 times a week for 4 weeks.
10 animals per group
Chromosome Aberrations
with 20 metaphases scored
per animal
Male Wistar rats -
30 days old
Cypermethrin No effects on weight gain.
Lead Acetate induced an increase in aneuploidy, and the
percent of cells with damage, but did not increase structural
damage or alterations in organ weight.
Cypermethrin and lead together increased structural
aberrations that were predominately acentric fragments.
However, this was compared to untreated controls and not the
individual treatments. Considering the individual treatments,
the two together are less than additive.
Nehez et al. (2000)
X
-------
Table AX5-6.5. Genotoxic/Carcinogenic Effects of Lead - Genotoxicity Human Cell Cultures Mutagenesis
to
o
o
Compound
Dose and Duration
Cell Type
Co-exposure
Effects
Reference
X
-------
Table AX5-6.6. Genotoxic/Carcinogenic Effects of Lead - Genotoxicity Human Cell Cultures Clastogenicity
to
o
o
Compound
Assay (Concentration
and Exposure Time)
Cell Type and Culture
Medium
Co-exposure
Effects
Reference
Lead Chromate Chromosome Aberrations Human Foreskin Fibroblasts
(0.08-2 jig/cm2 for 24 h) (Caucasian) in EMEM + 15%FBS
X
oo
Oi
H
6
o
o
H
O
O
H
W
O
Lead Chromate Chromosome Aberrations
(0.1-5 ug/cm2 for 24 h)
Lead Chromate Chromosome Aberrations
(0.1-5 ug/cm2 for 24 h)
Lead Chromate Chromosome Aberrations
(0.1-5 ug/cm2 for 24 h)
Lead Chromate Chromosome Aberrations
(0.05-5 ug/cm2 for 24 h)
Lead Chromate Chromosome Aberrations
(0.05-5 jig/cm2 for 24 h)
Primary Human Lung Cells in
DMEM/F12 + 15%FBS
Primary Human Lung Cells and
WTHBF-6 -human lung cells with
hTERT in DMEM/F12 + 15%FBS
WTHBF-6 -human lung cells with
hTERT in DMEM/F12 + 15%CCS
WTHBF-6 -human lung cells with
hTERT in DMEM/F12 + 15%CCS
WTHBF-6 -human lung cells with
hTERT in DMEM/F12 + 15%CCS
None
None
None
Vitamin C
(2mM
co-exposure
for 24 h)
Vitamin C
(2mM
co-exposure
for 24 h)
None
Lead Chromate induced chromosome damage in a Wise et al.
concentration dependent manner. (1992)
This study was focused on chromate.
Lead Chromate induced chromosome damage in a Wise et al.
concentration dependent manner. (2002)
This study was focused on chromate.
Lead Chromate induced chromosome damage in a Wise et al.,
concentration dependent manner. Effects were similar in (2004a)
both cell types establishing the WTHBF-6 cells as a useful
model.
This study was focused on chromate.
Lead Chromate induced chromosome damage in a Xie et al. (2004)
concentration dependent manner.
Vitamin C blocked Cr ion uptake and the chromosome
damage after lead chromate exposure.
This study was focused on chromate.
Lead Chromate induced chromosome damage in a Wise et al.
concentration dependent manner. (2004b)
This study was focused on showing chromate and not lead
ions were the clastogenic species.
Lead Chromate induced chromosome damage in a Wise et al.
concentration dependent manner. (2004c)
This study was focused on comparing particulate chromate
compounds.
O
HH
H
W
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Table AX5-6.6 (cont'd). Genotoxic/Carcinogenic Effects of Lead - Genotoxicity Human Cell Cultures Clastogenicity
to
o
X
1
-------
Table AX5-6.7. Genotoxic/Carcinogenic Effects of Lead - Genotoxicity Human Cell Cultures DNA Damage
to
o
o
X
oo
oo
H
6
o
o
H
O
O
H
W
O
O
HH
H
W
Compound
Lead Acetate
Lead Acetate
Lead Chromate
Lead Chromate
Lead Acetate
Assay (Concentration and
Exposure Time)
DNA strand breaks as nucleiod
sedimentation (500 uM for
20-25 h)
DNA strand breaks as nucleoid
sedimentation assay (100 uM
for 30 min - 4 h)
DNA adducts (0.4-0.8 jig/cm2
for 24 h)
DNA double strand breaks
(0.1-5 ug/cm2 for 24 h) by
Comet assay and H2A.X foci
formation
DNA strand breaks and DNA
protein crosslinks and oxidative
lesions by comet assay (1-100
uM for 1 h)
Cell Type and
Culture Medium
HeLa Cells in AMEM
+ 5%FBS
HeLa Cells in
HEPES/
glucose buffer
Primary Human Small
Airway Cells in
Clonetics growth
medium
WTHBF-6 -human
lung cells with
hTERT in
DMEM/F12 +
15%CCS
Primary lymphocytes
in RPMI 1640 without
serum
Co-exposure
None
See also Table
AX5-6-16
Buthionine
sulfoximine (BSO) to
deplete cells of thiols
None
None
Vitamins A (10 uM),
C (10 uM), E
(25 uM), calcium
chloride (100 uM)
magnesium chloride
(lOOnM)orzinc
chloride (100 uM)
Effects
Lead acetate alone did not induce single strand breaks.
Lead acetate did not induce DNA strand breaks.
Lead chromate induced lead inclusion bodies and Cr-DNA adducts and
Pb-DNA adducts in a concentration-dependent manner.
Lead Chromate induced DNA double strand breaks in a concentration
dependent manner.
This study showed the damage was due to chromate and not lead.
Lead acetate induced an increase in DNA single strand breaks at 1 uM
that went down with increasing dose. The highest concentration was
significantly less than the damage in untreated controls. For double
strand breaks, all concentrations had more damage than the controls,
but there was less damage in the highest concentrations than the two
lower ones. Lead only induced a slight increase in the amount of
DNA-protein crosslinks at the highest concentration.
Co exposure to magnesium had no effect. Co-exposure to Vitamins A,
C, and E or zinc exacerbated the DNA single strand break effects at the
highest concentration. Co-exposure to calcium exacerbated the single
strand break effect at all concentrations.
Reference
Hartwig et al
(1990)
Snyder and
Lachmann
(1989)
Singh et al.
(1999)
Xie et al.
(2005)
Wozniak and
Blasiak (2003)
-------
Table AX5-6.7 (cont'd). Genotoxic/Carcinogenic Effects of Lead - Genotoxicity Human Cell Cultures DNA Damage
to
o
o
H
6
o
o
H
O
O
H
W
O
O
HH
H
W
Compound
Lead Nitrate
Assay (Concentration and
Exposure Time)
DNA-protein crosslinks by
SDS precipitation
(l-10mMfor6h)
Cell Type and
Culture Medium
Human Burkitt's
lymphoma cells -
EBV transformed in
RPMI 1640 +
10%FCS
Co-exposure
Effects
None Lead nitrate did not induced DNA protein crosslinks. Independent
samples were analyzed by 5 different laboratories.
Reference
Costa et al.
(1996)
Abbreviations:
hTERT = hTERT is the catalytic subunit of human telomerase.
Medium and Components
AMEM = Alpha Minimal Essential Medium;
EMEM = Eagle's Minimal Essential Medium;
DMEM/F12 = Dulbecco's Minimal Essential Medium/Ham's F12;
FBS = Fetal Bovine Serum
FCS = Fetal Calf Serum
^ Differences between the serum types are unclear as insufficient details are provided by authors to distinguish
X
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VO
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to
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o
ON
Table AX5-6.8. Genotoxic/Carcinogenic Effects of Lead - Genotoxicity Animal Cell Cultures Mutagenicity
Compound
Lead Acetate
Lead Acetate
-insoluble
precipitate at high
dose.
Assay and Duration
Cytotoxicity (1-25 uM for 24 h)
Mutagenesis- HPRT (0.5-5 uM for 44 h)
Cytotoxicity (0.5-2000 uM for 5 days)
Mutagenesis - gpt (0.5-1700 uM for 5 days)
Cell Type
V79 in AMEM +
10%FBS
G12-CHV79 cells
with 1 copy gpt gene
in Ham's F 12 +
5%FBS
Co-exposure
None
See also
Table AX5-6-16
See also Table
AX5-6-17
Effects
LC50 = 3 uM
Lead acetate alone was not mutagenic.
LC50 = 1700 uM
Lead acetate was mutagenic, but only at toxic
concentration (1700 uM) where precipitate formed
not at lower concentrations (500 or 1000 uM).
Reference
Hartwig et al (1990)
Roy and Rossman
(1992)
There were no statistical analyses of these data.
X
H
6
o
o
H
O
O
H
W
O
O
HH
H
W
Lead Chloride
Lead chloride
Lead chloride
Lead Chromate
Lead Chromate
Cytotoxicity (0.1-1 uM for 1 h) AS52-CHO-gpt, lack None
hprt in HBSS
Mutagenicity - gpt assay (0.1-1 uM for 1 h) followed by Ham's
F12 + 5%FBS
Cytotoxicity (0.1-1 uM for 1 h) AS52-CHO-gpt, lack Allopurinol (50 uM)
hprt in HBSS to inhibit xanthine
Mutagenicity - gpt assay (0.1-1 uM for 1 h) followed by Ham's oxidase
F12 + 5%FBS
Mutagenicity - gpt assay (0.1-1 uMfor 1 h) AS52-CHO-gpt, lack
hprt in HBSS
followed by Ham's
sequence F12 + 5%FBS
PCR/Southern to analyze mutants for
Cytotoxicity (10 -100 uM for 24 h)
HGPRT assay (10 -100 uM for 24 h)
V79 CHL - HPRT
low clone in MEM +
10% FCS
Mutagenicity as Sodium/potassium ATPase
(ouabain resistance) or 6-thioguanine
resistance (25-100 uM for 5 h)
C3H10T1/2 cells in
EMEM + 10% FBS
None
NTA
None
LC74 = 1 uM (maximum concentration tested)
Lead chloride induced a dose-dependent increase in
the number of 6 thioguanine resistant mutants.
Did not adjust and compare as previous studies.
LC74 = 1 uM. Allopurinol had no effect on
Cytotoxicity.
Lead chloride was mutagenic (0.8 and 1 uM).
Allopurinol reduced mutagenesis.
Lead chloride (0.1-0.4 uM) caused mostly point
mutations. Higher concentrations (0.5-1 uM)
caused more deletions.
There were no statistical analyses of these data.
Usually examined fewer mutations than control.
Mutagenesis was assessed with HGPRT assay. Lead
chromate was not mutagenic.
Co-exposure to NTA caused Lead chromate to
become mutagenic through increased solubilization.
This mutagenic effect was completely attributed to
the Cr(VI) ions.
Lead chromate was not mutagenic.
Arizaetal. (1996)
Arizaetal. (1998)a
Arizaetal. (1998)b
Celotti et al. (1987)
Patierno et al.
(1988) and Patierno
and Landolph
(1989) (both papers
present the same
data)
-------
Table AX5-6.8 (cont'd). Genotoxic/Carcinogenic Effects of Lead - Genotoxicity Animal Cell Cultures Mutagenicity
to
o
o
Compound
Lead Nitrate
Precipitate at 1000
uM and higher.
Lead nitrate
-no insoluble
precipitate
Lead Sulfide
Assay and Duration
Cytotoxicity (50-5,000 uM for 5 days)
Mutagenesis at HPRT locus (50-2,000 uM
for 5 days)
Cytotoxicity (0.5-2000 uM for 5 days)
Mutagenesis - gpt (0.5-1700 uM for 5 days)
Cytotoxicity (100-1,000 uM for 24 h)
Mutagenicity at HPRT locus (100-1,000 uM
for 24 h)
Cell Type
V79CHL-HPRT
low clone in Ham's
F12+10%FBS
G12-CHV79 cells
with 1 copy gpt gene
in Ham's F 12 +
5%FBS
V79CHL-HPRT
low clone in Ham's
F12+10%FBS
Co-exposure
None
See also Table
AX5-6-17
None
Effects
LC50 = 2950 uM
Lead nitrate was mutagenic at 500 uM, but there
was no dose response as higher doses less
mutagenic though still 2-4 fold higher. There were
no statistical analyses of these data.
LC 50 = 1500 uM
Lead nitrate was not mutagenic. There were no
statistical analyses of these data.
LC50 = 580 uM; did not increase with longer
exposures.
Mutagenic at 376 and 563 uM. Not mutagenic
Reference
Zelikoff etal.
(1988)
Roy and Rossman
(1992)
Zelikoff etal.
(1988)
lower or higher. Suggested Cytotoxicity prevented
mutagenesis at higher concentrations. There were
no statistical analyses of these data.
X
H
6
o
o
H
O
O
H
W
O
O
HH
H
W
Abbreviations:
V79 are a Chinese Hamster Lung Cell Line;
G12 - CHV79 are derived from V79;
CHO are a Chinese Hamster Ovary Cell Line ;
AS52 are derived from CHO;
C3H10T/12 cells are a mouse embryo cell line
Medium and Components
AMEM = Alpha Minimal Essential Medium;
EMEM = Eagle's Minimal Essential Medium;
HBSS = Hank's Balanced Salt Solution
FBS = Fetal Bovine Serum
FCS = Fetal Calf Serum
Differences between the serum types are unclear as insufficient details are provided by authors to distinguish.
-------
Table AX5-6.9. Genotoxic/Carcinogenic Effects of Lead - Genotoxicity Animal Cell Cultures Clastogenicity
to
o
o
Oi
X
i
to
o
H
O
0
H
O
O
H
W
O
O
HH
H
W
Assay (Concentration and
Compound Exposure Time)
Lead Chromate Chromosome Aberrations
(0.4-8 ug/cm2 for 24 h)
Lead Chromate Chromosome Aberrations
(0.8-8 ug/cm2 for 24 h)
Lead Chromate Chromosome Aberrations
(0.8 or 8 ug/cm2 for 24 h)
Lead Chromate Chromosome Aberrations
(0.8-8 ug/cm2 for 24 h)
Lead Glutamate Chromosome Aberrations
(500-2,000 uM for 24 h)
Lead Nitrate Chromosome Aberrations
(500-2,000 uM for 24 h)
Insoluble precipitate at all
concentrations
Lead Nitrate Chromosome Aberrations
(3-30 uM for 2h +16 h recovery)
Lead Nitrate Chromosome aberrations
(0.05- 1 uM for 3-12 h)
Cell Type and Culture
Medium
Chinese Hamster Ovary AA8
cells in AMEM + 10%FBS
Chinese Hamster Ovary AA8
cells in AMEM + 10%FBS
Chinese Hamster Ovary AA8
cells in AMEM + 10%FBS
Chinese Hamster Ovary AA8
cells in AMEM + 10%FBS
Chinese Hamster Ovary AA8
cells in AMEM + 10%FBS
Chinese Hamster Ovary AA8
cells in AMEM + 10%FBS
Chinese Hamster Ovary cells
in EMEM + 10%FBS
Chinese Hamster Ovary AA8
cells in DMEM +10% NCS
Co-exposure
None
Vitamin C (1 mM for
24 h as co-exposure to
block Cr uptake)
Vitamin E (25 uM as
pretreatment for 24 h)
Vitamin C (1 mM as
pretreatment for 24 h)
Vitamin E (25 uM as
pretreatment for 24 h)
Vitamin E (25 uM as
pretreatment for 24 h)
Vitamin E (25 uM as
pretreatment for 24 h)
None
Crown ethers to
modify effect through
chelation and uptake
Effects
Lead chromate induced chromosome damage in a
concentration dependent manner.
This study was focused on chromate.
Lead chromate induced chromosome damage in a
concentration dependent manner. This effect and uptake
of Cr ions were blocked by vitamin C.
This study was focused on chromate.
Lead chromate induced chromosome damage in a
concentration dependent manner. Vitamin E blocked
clastogenic activity of lead chromate, but had no effect on
other lead compounds.
This study found that the chromosome damage was
mediated by chromate ions and not lead ions
Lead chromate induced chromosome damage in a
concentration dependent manner. Vitamins C and E
blocked clastogenic activity of lead chromate.
This study was focused on chromate.
Lead glutamate induced chromosome damage at 1 mM
but not at higher or lower concentrations.
Vitamin E did not modify this effect.
Lead nitrate did not induce chromosome damage.
Lead nitrate did not induce chromosome damage.
Lead nitrate did not induce chromosome damage.
Reference
Wise et al.
(1992)
Wise et al.
(1993)
Wise et al.
(1994)
Blankenship
etal. (1997)
Wise et al.
(1994)
Wise et al.
(1994)
Lin et al.
(1994)
Cai and
Arenaz
(1998)
-------
H
6
o
o
H
O
O
H
W
O
O
HH
H
W
Table AX5-6.9 (cont'd). Genotoxic/Carcinogenic Effects of Lead - Genotoxicity Animal Cell Cultures Clastogenicity
to
o
o
Assay (Concentration and
Compound Exposure Time)
Cell Type and Culture
Medium
Co-exposure
Effects
Reference
Lead Acetate
SCE (1-10 uM for 26 h+)
V79 in AMEM + 10%FBS
None
See also
Table AX5-6-17
Lead acetate alone did not induce SCE. Only 25 cells per
treatment analyzed.
Hartwig et al
(1990)
Lead Acetate Micronucleus assay (0.01-10 uM for
18 h)
Chinese Hamster V79 cells in
DMEM + 10% ECS
None Lead acetate induced an increase in micronuclei that Bonacker
increased with concentration and reached a plateau. Two et al. (2005)
experiments were done and presented separately as a
Figure and a Table. The magnitude of the effects was
small to modest and statistics were not done.
Lead Nitrate SCE Formation (500-3,000 uM for 24 h) V79 CHL - HPRT low clone in
Precipitate at 1000 uM and higher. Ham's F12 +10% FBS
None
No SCE. Only 30 cells analyzed per treatment.
Zelikoff
etal. (1988)
X
vo
OJ
Lead Nitrate
Lead Nitrate
Micronucleus Formation (3-30 uM for CHO cells in EMEM +
2h+16 h recovery) 10%FBS
SCE (3-30 uM for 2h +16 h recovery)
SCE (0.05- 1 uM for 3-12 h)
CHO AA8 in DMEM +10%
NCS
None Lead nitrate did not induce micronuclei formation Lin et al.
Lead nitrate induces a concentration-dependent increase (1994)
in SCE (3, 10, 30 uM).
Crown ethers to Lead nitrate caused a weak concentration-dependent Cai and
modify effect increase in SCE. These data were not statistically Arenaz
through chelation analyzed. The effect was reduced by a crown ether (1998)
and uptake probably because a similar reduction was seen in
spontaneous SCE.
-------
Table AX5-6.9 (cont'd). Genotoxic/Carcinogenic Effects of Lead - Genotoxicity Animal Cell Cultures Clastogenicity
to
o
o
Compound
Lead Sulfide
Assay (Concentration and
Exposure Time)
SCE Formation (100-1,000 uM for
24 h)
Cell Type and Culture
Medium
V79 CHL - HPRT low clone
in Ham's F12 +10% FBS
Co-exposure
None
Effects
No SCE. Only 30 cells analyzed per treatment.
Reference
Zelikoffetal.
(1988)
H
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H
O
O
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W
O
O
HH
H
W
Abbreviations:
V79 are a Chinese Hamster Lung Cell Line;
CHO are a Chinese Hamster Ovary Cell Line ;
Medium and Components
AMEM = Alpha Minimal Essential Medium;
DMEM = Dulbecco's Minimal Essential Medium;
EMEM = Eagle's Minimal Essential Medium;
. HBSS = Hank's Balanced Salt Solution
r*j FBS = Fetal Bovine Serum
£; FCS = Fetal Calf Serum
^ NCS = Newborn Calf Serum
4^
Differences between the serum types are unclear as insufficient details are provided by authors to distinguish.
-------
Table AX5-6.10. Genotoxic/Carcinogenic Effects of Lead - Genotoxicity Animal Cell Cultures DNA Damage
to
o
o
X
H
6
o
o
H
O
O
H
W
O
O
HH
H
W
Compound
Lead Acetate
Lead Acetate
Assay (Concentration
and Exposure Time)
DNA damage as alkaline
elution (exposure time and
dose not given)
Precipitate at 1000 uM and
higher.
DNA strand breaks as nick
translation (1700 uM for
Cell Type and Culture Medium
V79 CHL - HPRT low clone in Ham's
F12+10%FBS
G12 - CHV79 cells with 1 copy gpt
gene in Ham's F12 + 5%FBS
Co-exposure
None
See also Table
AX5-6-17
Effects
No DNA damage (Single strand breaks, DNA-
protein crosslinks or DNA-DNA crosslinks).
However, the data was not shown
Lead acetate did not induce SSB. Lead acetate
(1700 uM) did increase nick translation when an
Reference
Zelikoff etal. (1988)
Roy and Rossman
(1992)
Lead Chromate
Lead Chromate
Lead Nitrate
5 days)
Or Supercoiled relaxation
(1000 uM for 5 days)
Insoluble precipitate at high
dose.
DNA damage as alkaline
elution (0.4 -8 ug/cm for
24 h plus 24 recovery)
DNA adducts
(0.8or8ng/cm2for24h)
DNA Protein Crosslinks as
SDS precipitation
50-5,000 uM for 4 h)
Chinese Hamster Ovary AA8 cells in
AMEM + 10%FBS
Chinese Hamster Ovary AA8 cells in
AMEM + 10%FBS
Novikoff ascites hepatoma cells
None
Vitamin C (1 mM as
pretreatment for 24 h)
Vitamin E (25 uM as
pretreatment for 24 h)
None
exogenous polymerase was added. There were no
statistical analyses of these data.
Lead chromate induced DNA single strand breaks Xu et al. (1992)
in a concentration dependent manner, which were
all repaired by 24 h post-treatment.
Lead chromate induced DNA protein crosslinks in
a concentration dependent manner, which
persisted at 24 h post-treatment.
Lead chromate did not induce DNA-DNA
crosslinks.
This study was focused on chromate.
Lead chromate induced DNA adducts in a Blankenship et al.
concentration dependent manner. Vitamins C and (1997)
E blocked DNA adducts induced by lead
chromate.
This study was focused on chromate.
Lead Nitrate induced DNA protein crosslinks in a Wedrychowski et al.
concentration dependent manner. (1986)
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to
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Oi
Table AX5-6.10 (cont'd). Genotoxic/Carcinogenic Effects of Lead - Genotoxicity Animal Cell Cultures DNA Damage
Compound
Lead Nitrate
Assay (Concentration and
Exposure Time)
DNA strand breaks as nick
translation (1700 uM for 5 days)
Cell Type and Culture
Medium
G12 - CHV79 cells with 1 copy
gpt gene in Ham's F12 + 5%
FBS
Co-exposure
See also Table
AX5-6-17
Effects
Lead nitrate (1700 uM) did increase nick translation when an
exogenous polymerase was added. There were no statistical
analyses of these data.
Reference
Roy and Rossman
(1992)
Abbreviations:
G12 - CHV79 are derived from V79;
V79 are a Chinese Hamster Lung Cell Line;
CHO are a Chinese Hamster Ovary Cell Line ;
Medium and Components
AMEM = Alpha Minimal Essential Medium;
FBS = Fetal Bovine Serum
FCS = Fetal Calf Serum
Differences between the serum types are unclear as insufficient details are provided by authors to distinguish.
H
6
o
o
H
O
O
H
W
O
O
HH
H
W
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S Table AX5-6.11. Genotoxic/Carcinogenic Effects of Lead - Genotoxicity Non-Mammalian Cultures
&>
<<
K> Compound Assay and Concentration Cell Type Co-exposure Effects Reference
o
*-^ Lead Chromate Mutation Frequency (50-500 ug/plate) Salmonella Nitrilotriacetic acid Lead chromate and its related pigments did not induce Connor and Pier
(and 13 Anchorage Independence for cells isolated typhimurium+/-S9 (NTA to dissolve mutagenicity. (1990)
pigments during morphological transformation fraction insoluble compounds) A few did when dissolved in NTA.
containing lead and
chromate) Neoplastic Transformation for cells Silica Encapsuiation Encapsulation prevented mutagenesis in those that
isolated during morphological were positive when dissolved in NTA.
transformation S9 had no effect.
Studied as a chromate compound.
X
H
6
o
o
H
O
O
H
W
O
O
HH
H
W
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Table AX5-6.12. Genotoxic/Carcinogenic Effects of Lead - Genotoxicity as it Pertains to Potential Developmental Effects
to
o
o
X
oo
H
6
o
o
H
O
O
H
W
O
O
HH
H
W
Compound
Lead Acetate
Assay and Concentration
25-400 mg/kg given i.p as single injection
and animals studied after 24 h
Sperm morphology
Species
Male Swiss Mice -
9-12 weeks old
Co-exposure
None
Effects
Lead induced sperm head abnormalities at 50-100
mg/kg. A lower dose was negative and higher doses
were not done.
Reference
Fahmy (1999)
Lead Acetate 200 or 400 mg/kg given by gavage daily Male Swiss Mice - Calcium chloride (40 or Lead induced sperm abnormalities at 200 and 400 Aboul-Ela (2002)
for 5 days 9-12 weeks old 80 mg/kg by gavage mg/kg. A lower dose was negative and higher doses
5 animals r>er arouc daily for 3 days given 2 were not done. Calcium appeared to block this effect.
weeks after lead
Sperm Morphology exposure)
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to
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o
X
H
6
o
o
H
O
O
H
W
O
O
HH
H
W
Lead Nitrate
Table AX5-6.13. Genotoxic/Carcinogenic Effects of Lead - Genotoxicity as it Pertains to
Potential Developmental Effects - Children
Compound Exposure Regimen Species
Co-exposure Effects Reference
Lead chloride Administered in drinking water Male NMRI Cyclophosphamide - Pb did not increase resorptions indicating no dominant lethal Kristensen et al.
1.33 g/L for 6 weeks Mice - 9 weeks old 120 mg/kg b.w. given mutagenic effect. Pb appeared to have a small, but statistically (1993).
i.p 7 days prior to start insignificant reduction in the number of resorptions.
of breeding Cyclophosphamide reduced live implants in female mice.
Lead Nitrate 12.5-75 mg/kg given iv on 9th day of ICR Swiss Webster
gestation for 9 days. Mice - 6-8 week old
None 12.5-50 mg/kg had no effect on resorption or fetal viability. Nayak et al.
75 mg/kg demonstrated some increased resorption though (1989)a
Mothers and fetuses analyzed on
G18.
5 animals per group
Resorptions, fetal viability, and
chromosome damage in the mother
and fetus were examined.
100-200 mg/kg given iv on 9th day
of gestation for 9 days.
Mothers and fetuses analyzed on
G18.
Group size not given.
Resorptions, fetal viability and
chromosome damage, SCE and
NOR in the mother and fetus were
examined. Mother - bone marrow;
fetus liver or lung
3 mothers and fetuses per dose were
analyzed.
ICR Swiss Webster
Mice - 6-8 week old
statistics were not done.
No chromosome damage was seen in untreated controls. A low
level 1-3 and 2-5 aberrations were seen in mothers and fetuses
respectively.
There was no dose response and no statistical analyses. Data
interpretation is also complicated as too few metaphases were
analyzed 20-40 total.
No descriptions of potential effects on maternal health parameters
or fetal weights.
No indication of how many animals included in the chromosomal
analysis.
None Lead levels were found in both mother and fetus indicating no
problems crossing the placenta.
All doses indicated increased resorption and decreased placental
weights. No effects on fetal weight.
Significant increase in SCE in mothers at 150 and 200 mg/kg.
No increase in SCE in fetuses.
Significant decrease in NOR in both mother and fetuses.
No gaps or breaks in mothers or fetuses.
Some weak aneuploidy at lowest dose.
Some karyotypic chromosome damage was seen. No explanation
of how many cells analyzed per animal (3 animals per dose were
analyzed) as only 20- 40 cells were analyzed.
There was no dose response and no statistical analyses for
chromosome damage. No details on how many animals analyzed
for metaphase damage or how many cells per animal.
Data interpretation is also complicated as too few metaphases
were analyzed 10-25 for SCE. Not given for CA.
No detail on potential maternal toxicity.
Nayak et al.
(1989)b
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Table AX5-6.14. Genotoxic/Carcinogenic Effects of Lead - Epigenetic Effects and Mixture Interactions - Animal
to
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Compound
Exposure Regimen
Species
Co-exposure
Effects
Reference
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Lead Acetate Administered as an i.p. injection of
100 nl/kg.
Animals were studied either 24 h
after a daily dose for 3 days or for
various times (5 min-48 h) after a
single dose.
Lead Nitrate
Administered as an i.p. injection of
100 umol/kg.
Animals were studied either 24 h
after a daily dose for 3 days or for
various times (5 min-48 h) after a
single dose.
Lead Nitrate Administered as an i.p. injection of
100 umol/kg. Some rats were
partially hepatectomized.
Animals were studied 48 h after
injection.
Lead Nitrate Administered as an i.v. injection of
20, 50, or 100 umol/kg.
Animals were studied 24 h after
injection.
Male Wistar Rats -
10 weeks old
Male Wistar Rats -
10 weeks old
Male Sprague
Dawley rats
Actinomycin D
(0.8 mg/kg)
administered i.p. for
4 h before a single
dose of lead acetate.
Actinomycin D
(0.8 mg/kg)
administered i.p. for
4 h before a single
dose of lead acetate.
Partial Hepatectomy
Male Fisher 344 rats 2-methoxy-
- 7 weeks old 4-aminobenzene to
induce P4501A2
or
3-methylcholanthrene
to induce 4501A1
Lead acetate induced GST-P, which required the cis element, GPEI Suzuki et al.
(GST-P enhancer I). (1996)
Actinomycin D blocked the effects indicating that regulation was
at the mRNA level.
Lead acetate induced c-jun, which exhibited three peaks of
exposure over 48 h.
Lead acetate was more potent than lead nitrate.
Lead nitrate induced GST-P, which required the cis element, GPEI Suzuki et al.
(GST-P enhancer I). (1996)
Actinomycin D blocked the effects indicating that regulation was
the mRNA level.
Lead nitrate induced c-jun, which exhibited three peaks of
exposure over 48 h.
Lead nitrate was less potent than lead acetate.
Lead nitrate induced GSH and GST 7-7 activity. Dock (1989)
Partial hepatectomy did not induce GSH or GST 7-7.
Lead nitrate selectively inhibited P4501A2 and its induction by 2- Degawa et al
methoxy-4-aminobenzene at the mRNA and protein level in a (1993)
dose-dependent manner.
Lead nitrate had minimal effect on P4501A1 and its induction by
3- methyl cholanthrene.
Lead nitrate did not affect microsomal activity.
Lead nitrate induced GST-P in a dose-dependent manner.
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Table AX5-6.15. Genotoxic/Carcinogenic Effects of Lead - Epigenetic Effects and Mixture Interactions - Human
to
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Assay (Concentration and
Compound Exposure Time)
Lead Acetate Tyrosine aminotransferase
expression and activity
(0.3-10 uM for 24- 48 h)
PKC activity: 10 uM for 48h
Lead Nitrate EROD/MROD activity
(10-100 uM for 24 h)
NAD(P)H: quinone
oxidoreductase activity
(10-100 uM for 24 h)
Glutathione- S -transferase
Ya activity (10-100 uM for 24 h)
Lead Nitrate NAD(P)H: quinone
oxidoreductase activity (25 uM
for 24 h)
Glutathione-S-transferase Ya
activity (25 uM for 24 h)
Abbreviations:
Medium and Components
DMEM = Dulbecco's Minimal Essential Medium;
FBS = Fetal Bovine Serum
FCS = Fetal Calf Serum
Differences between the serum types are unclear as
Cell Type
and Culture
Medium
H4-IIE-C3 -
human
hepatoma cells
in DMEM +
2. 5% FCS
Hepa Iclc?
wild type cells
in DMEM +
10% FBS
C12- AHR-
deficient Hepa
Iclc? cells in
DMEM + 10%
FBS
Co-exposure
Dexamethasone
(0.1 uMfor 16 h),
or calcium chloride
(10 uM) or genistein
(100 uM to block PKC
activity)
TCDD(O.lnM),
3-methyl cholanthrene
(0.25 uM), beta-
naptflavone (10 uM),
benzo(a)pyrene (1 uM)
TCDD (0. 1 nM),
3-methyl cholanthrene
(0.25 uM), beta-
naptflavone (10 uM),
benzo(a)pyrene (1 uM)
Effects
Lead acetate inhibited glucocorticoid -induction of tyrosine
aminotransferase in a time- and dose-dependent manner.
Co-treatment with calcium reduced the effects of lead.
Co-treatment with genistein increased the effects of lead.
Lead acetate decreases PKC activity and its translocation from the
cytosol to the particulate cellular fraction.
Lead did not affect EROD/MROD activity.
Lead reduced CYP1A1 induction by TCDD, 3-methyl cholanthrene,
beta-naptflavone, benzo(a)pyrene.
Lead increased NAD(P)H: quinone oxidoreductase activity
Lead increased NAD(P)H: quinone oxidoreductase activity induction
by TCDD, 3-methyl cholanthrene, beta-naptflavone, benzo(a)pyrene.
10 uM increased Glutathione-S-transferase Ya activity.
25 and 100 uM increased Glutathione-S-transferase Ya activity.
Lead nitrate did not affect Glutathione-S-transferase Ya induction by
TCDD, 3-methyl cholanthrene, beta-naptflavone, benzo(a)pyrene.
Lead nitrate did not increase NAD(P)H: quinone oxidoreductase and
Glutathione-S-transferase Ya activity
Lead increased NAD(P)H: quinone oxidoreductase activity induction
by TCDD, 3-methyl cholanthrene, beta-naptflavone, benzo(a)pyrene.
Lead did not affect Glutathione-S-transferase Ya induction by TCDD,
3-methyl cholanthrene, beta-naptflavone, benzo(a)pyrene.
Reference
Tonner and Heiman
(1997)
Korashy and
El-Kadi (2004)
Korashy and
El Kadi (2004)
insufficient details are provided by authors to distinguish.
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Table AX5-6.16. Genotoxic/Carcinogenic Effects of Lead - Epigenetic Effects and Mixture Interactions -
DNA Repair - Human
to
o
Assay (Concentration and
Compound Exposure Time)
Lead Acetate DNA strand breaks as nucleoid
sedimentation (500 uM for
20-25 h)
Cell Type
and Culture
Medium Co-exposure
HeLa Cells in UV (5 J/m2)
AMEM +
5%FBS
Effects
Lead acetate alone did not induce single strand breaks. UV did induce
strand breaks. Co-exposure of lead and UV cause DNA strand breaks
to persist longer suggesting an inhibition of repair.
Reference
Hartwigetal(1990)
Abbreviations:
Medium and Components
AMEM = Alpha Minimal Essential Medium;
FBS = Fetal Bovine Serum
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Table AX5-6.17. Genotoxic/Carcinogenic Effects of Lead - Epigenetic Effects and Mixture Interactions -
DNA Repair - Animal
Assay (Concentration and
Compound Exposure Time)
Cell Type
and Culture
Medium
Co-exposure
Effects
Reference
>
X
Lead Acetate Cytotoxicity (0.5-5 uM for 24 h)
Mutagenesis- HPRT (0.5-5 uM
for 44h)
SCE(l-10nMfor26h+)
Lead Acetate Mutagenesis - gpt (0.5-1700 mM
for 24 h)
DNA strand breaks as
supercoiled relaxation (1000 mM
for 24 h)
V79in
AMEM +
10%FBS
UV (5 J/m2)
G12-CHV79
cells with 1
copy gpt gene
in Ham's F12
+ 5%FBS
UV (2 J/m2), or MNNG
(0.5 ug/L)
Lead acetate ( 3 and 5 uM) increased UV-induced increased
cytotoxicity with no dose response (plateau). There were no statistical
analyses of these data.
Lead acetate (0.5-5) increased UV mutagenicity though with no dose
response (plateau). There were no statistical analyses of these data
Lead acetate (1-10 uM) increased UV-induced SCE. Significant at
p<0.01. Only 25 cells per treatment analyzed.
Lead acetate was co-mutagenic with UV and MNNG increasing
frequency 2-fold.
Lead acetate does not increase strand breaks induced by UV.
Hartwigetal(1990)
Roy and Rossman
(1992)
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Abbreviations:
G12 - CHV79 are derived from V79;
V79 are a Chinese Hamster Lung Cell Line;
Medium and Components
AMEM = Alpha Minimal Essential Medium;
FBS = Fetal Bovine Serum
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Table AX5-6.18. Genotoxic/Carcinogenic Effects of Lead - Mitogenesis - Animal
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>
X
Compound
Lead Acetate
Lead Nitrate
Exposure Regimen
Administered lead acetate (12.5 mg/kg) i.p.
Animals studied 24 h after injection.
Liver initiation induced by the resistant hepatocyte
model
Species
Male B6 Mice
Male Wistar Rats-
4 per group
Co-exposure
None
Partial Hepatectomy
Effects
Lead acetate induced TNF-alpha in glial and
neuronal cells in the cerebral cortex and
subcortical white matter and on Purkinje cells in
the cerebellum, but did not induced apoptosis in
these areas
Lead nitrate stimulated DNA synthesis and liver
cell proliferation
Reference
Cheng et al. (2002)
Columbano et al.
(1987)
Initiation followed by iv injection of lead nitrate
(100 uM/kg ) or partial hepatectomy
Studied DNA synthesis (30 h after injection) and
preneoplastic nodule formation
(5 weeks after injection)
Lead Nitrate Liver initiation induced by the resistant hepatocyte
model (diethylnitrosamine followed by 2-
acetylaminofluorene plus carbon tetrachloride)
Initiation followed by iv injection of 4 doses of lead
nitrate (100 uM/kg ) given once every 20 days or
partial hepatectomy, ethylene dibromide, or
nafenopine
Animal were evaluated for preneoplastic foci at 75 or
155 days after initiation.
Lead nitrate did not induce preneoplastic nodule.
Partial hepatectomy did.
Male Wistar Rats- Diethylnitrosamine Lead nitrate, partial hepatectomy, ethylene
4 per group dibromide, or nafenopine all stimulated DNA
synthesis and liver cell proliferation
Lead nitrate, ethylene dibromide, or nafenopine
did not induce preneoplastic nodule. Partial
hepatectomy did.
Columbano et al.
(1990)
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Lead Nitrate Liver initiation induced by the orotic acid model
(diethylnitrosamine plus orotic acid)
Initiation followed by iv injection of lead nitrate (100
uM/kg) or partial hepatectomy, or by gavage:
ethylene dibromide, or cyproterone
DNA synthesis was examined at various time
intervals (24 h -5 days) after injection.
Male Wistar Rats- Diethylnitrosamine Lead nitrate, partial hepatectomy, ethylene
4 per group dibromide, or cyproterone all stimulated DNA
synthesis within 30 minutes.
Lead nitrate induced DNA synthesis for 5 days.
Coni et al. (1991)
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Table AX5-6.18 (cont'd). Genotoxic/Carcinogenic Effects of Lead - Mitogenesis - Animal
to
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Compound
Exposure Regimen
Species
Co-exposure
Effects
Reference
>
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o
H
O
O
H
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O
O
HH
H
W
Lead Nitrate Liver initiation induced by the resistant hepatocyte Male Wistar Rats-
model (diethylnitrosamine followed by 2- 4 per group
acetylaminofluorene plus carbon tetrachloride) or the
phenobarbital model (diethylnitrosamine plus orotic
acid), or the orotic acid model (diethylnitrosamine
plus orotic acid)
Initiation followed by iv injection of lead nitrate
(100 uM/kg ) or partial hepatectomy, or carbon
tetrachloride by gavage
Animals were studied 6 weeks after initiation.
Lead Nitrate Liver initiation induced by the resistant hepatocyte Male Wistar Rats-
model (diethylnitrosamine followed by 4 per group
2-acetylaminofluorene plus carbon tetrachloride)
Initiation followed by iv injection of lead nitrate
(100 uM/kg ) or partial hepatectomy, or by gavage:
ethylene dibromide, or cyproterone, or nafenopine
Also tried either 1 or 2 additional iv injections of lead
over 3 day intervals.
Animals were studied at various intervals (1-6 days)
after injection
Lead Nitrate Administered as i.v. injection of lead nitrate Male Wistar Rats-
(100 uM/kg ) or partial hepatectomy, or by gavage: 4 per group
carbon tetrachloride, or ethylene dibromide, or
cyproterone, or nafenopine
Animals were studied at various time intervals
(0.25-24 h) after injection.
Lead Nitrate Administered as i.v. injection of lead nitrate Male Wistar Rats-
(100 uM/kg ) or partial hepatectomy, or nafenopine 4 per group -
by gavage. 8 weeks old
Animals were studied at various time intervals
(24-96 h) after injection.
Lead Nitrate Administered as i.v. injection of lead nitrate Male Wistar rats -
(10 uM/100 g) 4 rats per group
Studies for apoptosis at 12, 24, 36, 48, 72, 96, 120,
168, 336 h after injection
Partial
Hepatectomy,
carbon tetrachloride
Diethylnitrosamine,
2-AAF
Partial
Hepatectomy,
carbon tetrachloride
None
None
Lead nitrate, partial hepatectomy, carbon
tetrachloride all stimulated DNA synthesis and
liver cell proliferation
Lead nitrate, did not induce preneoplastic nodules.
Partial hepatectomy and carbon tetrachloride did.
Ledda-Columbano
etal. (1992)
Coni et al. (1993)
Coni et al. (1993)
This study aimed to determine if mitogens induce
nodules at different time points.
Lead nitrate, ethylene dibromide, cyproterone, or
nafenopine did not induce preneoplastic nodules
at all. Partial hepatectomy did within 3 days.
Multiple injections of lead nitrate did not induce
preneoplastic lesions.
Lead nitrate, ethylene dibromide, cyproterone, or
nafenopine induced c-jun and c-myc but did not
induce c-fos.
Partial hepatectomy and carbon tetrachloride
induced c-jun, c-fos, and c-myc.
Lead nitrate induced a high incidence of
polyploidy and binucleated cells. These changes
were irreversible after 2 weeks. Many of these
cells were the newly synthesized cells.
Partial hepatectomy and carbon tetrachloride
induced tetraploid and octaploid mononucleated
cells.
Liver weight increased until day 5 then returned to Nakajima et al.
control levels. (1995)
DNA synthesis peaked at 36 h
Apoptosis peaked at day 4 and then decreased
gradually.
Melchiorri et al.
(1993)
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Table AX5-6.18 (cont'd). Genotoxic/Carcinogenic Effects of Lead - Mitogenesis - Animal
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Compound
Lead Nitrate
Lead Nitrate
Exposure Regimen
Administered as i.v. injection of lead nitrate
(100 uM/kg) or TNF-alpha.
Animals were studied at various time intervals
(12-48 h) after injection.
Administered after diethylnitrosamine (200 mg/kg
given i.p) as i.v. injection of lead nitrate (100 uM/kg )
Species
Male Wistar Rats-
4 per group - 5-6
weeks old
Male Wistar rats -
4 per group
Co-exposure
None
Carbon
tetrachloride
Effects
Lead nitrate and TNF-alpha induced similar
proliferative responses.
Lead nitrate induced apoptosis affects both newly
synthesized cells and non-replicative cells.
Reference
Shinozuka(1996)
Columbano et al.
(1996)
or instead carbon tetrachloride by gavage
Animals were studied at various time intervals
(3 -21 days) after injection.
Lead Nitrate Administered as i.v. injection of lead nitrate
(100 uM/kg ) or partial hepatectomy, or by gavage:
carbon tetrachloride, or cyproterone, or nafenopine
Animals were studied at various time intervals
(0.5-24h) after injection.
Male Wistar Rats
- 8 weeks old
Partial
Hepatectomy,
ethylene
dibromide,
nafenopine, or
cyproterone
Lead nitrate decreased the number and had no effect
on the size of placental glutathione-S-transferase
lesions. Carbon tetrachloride substantially
increased these lesions both in number and in size.
Lead nitrate induced NF-kB, TNF-alpha and iNOS, Menegazzi et al.
but not AP-1. (1997)
Carbon tetrachloride induced and activated NF-kB,
TNF-alpha iNOS, and AP-1.
Nafenopine and cyproteone did not induce or
activate NF-kB,TNF-alpha iNOS, or AP-1.
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Table AX5-6.19. Genotoxic/Carcinogenic Effects of Lead - Mitogenesis Human and Animal Cell Culture Studies
K> Assay (Concentration and Cell Type and
O Compound Exposure Time) Culture Medium Co-exposure
Effects
Reference
>
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Lead Acetate
Lead Acetate
Lead Acetate
Lead Acetate
Lead Acetate
Lead Acetate
H4-II-C3 - human
hepatoma cells in DMEM
+ 2.5% PCS
Cell Proliferation (0.1-100 uM for
2-6 days)
DNA synthesis (1-100 uM for 72
h)
Tyrosine aminotransferase
expression and activity (0.3-10 uM
for 24- 48 h)
Cell proliferation (10 uM- ImM for REL cells- Rat Epithelial
24 h-7 days) cells in Ham's F10
medium + 10%FBS
Cell growth (0.01-10 uM for
12-72 h)
Expression of genes in cytokine
pathways (0.01-10 uM for 24 h)
Cell proliferation (0.078-320 uM
for 48 h)
Apoptosis (1.25-80 uM)
Cell cycle analysis
DNA synthesis (1-50 uM for 24 h)
Expression of genes in mitogen
activated pathways (1-50 uM for
5 min-4h)
Cell proliferation (1 uM for 24 h)
Cell differentiation
(1 uM for 48 h)
PKC activation (1 uM for 24 h)
U-373MG-human
glioma cell line in DMEM
+ 10 or 20% FBS
Rat-1 fibroblasts in
EMEM +10% FBS
1321N1- human
astrocytoma cells in
DMEM + 0.1%BSA
Primary oligodendrocyte
progenitor cells - in
DMEM + 1% FBS
Dexamethasone Lead acetate inhibited cell growth in a time- and dose-
(0.1 uM for 16 h) dependent manner.
Lead acetate inhibited DNA synthesis in a dose-dependent
manner.
Lead acetate alone did not inhibit tyrosine aminotransferase.
Lead acetate inhibited glucocorticoid -induction of tyrosine
aminotransferase in a time- and dose-dependent manner.
None Lead acetate inhibited cell growth at all concentrations for
24 h - 7 days.
Lead acetate did not affect gap junction capacity, which is
often inhibited by tumor promoters.
None Lead acetate did not inhibit or enhance cell growth.
Lead acetate enhanced the expression of TNF-alpha, but
decreased interleukin- 1 beta, interleukin-6, gamma-
aminobutyric acid transaminase, and glutamine synthetase
under 10% FBS.
Lead acetate further enhanced the expression of TNF-alpha
under 20% serum, but had no effect at all on expression of
the other genes.
None Lead acetate inhibited cell growth at 0.635-320 uM.
Lead acetate induced apoptosis from 2.5-10 uM.
Lead acetate caused GS/M and S-phase arrest.
None Lead acetate induced DNA synthesis.
Lead acetate induced activation of MAPK, ERK1. ERK2,
MEK1 , MEK2, PKC, andpgO^.
Lead acetate did not activate PI3K or p70s*
None Lead acetate inhibited basal and growth factor stimulated
growth.
Lead acetate inhibited cell differentiation in a PHC
dependent-manner.
Lead acetate redistributes PKC from the cytosol to the
membrane, but did not increase PKC activity.
Heiman and
Tonner(1995)
Apostoli et al.
(2000)
Liu et al. (2000)
lavicoli et al.,
2001
Lu et al. (2002)
Deng and Portez
(2002)
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Table AX5-6.19 (cont'd). Genotoxic/Carcinogenic Effects of Lead - Mitogenesis Human and Animal Cell Culture Studies
to
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Compound
Lead Acetate
Lead Chloride
Lead Oxide
Lead Sulfate
Lead Chromate
Lead Chromate
Lead Chromate
Lead
Glutamate
Lead
Glutamate
Assay (Concentration and
Exposure Time)
Expression of TNF-alpha
(0.1-10 uM for 24 h)
Cell proliferation
(10 uM-lmM for 24 -48 h)
Cell proliferation
(10 uM-lmM for 24 h -7 days)
Cell proliferation
(10 uM-lmM for 24 -48 h)
Apoptosis (350 uM for 24 h)
Apoptosis (0.4-2 ug/cm2 for 24 h)
Growth Curve (0.5-5 ug/cm2 24 h)
Growth Curve
(250-1,000 uM for 24 h)
Mitotic Index
(250-2,000 uM for 24 h)
Growth Curve
(250-2,000 uM for 24 h)
Cell cycle Analysis
(250-2,000 uM for 24 h)
Cell Type and
Culture Medium
U-373MG - human
glioma cell line in
DMEM + 20% FBS
REL cells- Rat Epithelial
cells in Ham's F10
medium + 10% FBS
REL cells- Rat Epithelial
cells in Ham's F10
medium + 10% FBS
REL cells- Rat Epithelial
cells in Ham's F10
medium + 10% FBS
Chinese Hamster Ovary
AA8 cells in AMEM +
10%FBS
Primary Human Small
Airway Cells in Clonetics
growth medium
WTHBF-6 -human lung
cells with hTERT in
DMEM/F12 + 10%CCS
WTHBF-6 -human lung
cells with hTERT in
DMEM/F12 + 10%CCS
WTHBF-6 -human lung
cells with hTERT in
DMEM/F12 + 10%CCS
Co-exposure Effects
None Lead acetate did not induce apoptosis.
Lead acetate increased the expression of TNF-alpha in a
dose-dependent manner.
TNF-alpha was not involved in lead-induced apoptosis.
None Lead chloride inhibited cell growth at all concentrations for
24-48 h.
Lead chloride did not affect gap junction capacity, which is
often inhibited by tumor promoters.
None Lead oxide inhibited cell growth at all concentrations for 24
h - 7 days.
Lead oxide did not affect gap junction capacity, which is
often inhibited by tumor promoters.
None Lead sulfate inhibited cell growth at all concentrations for
24-48 h.
Lead sulfate did not affect gap junction capacity, which is
often inhibited by tumor promoters.
None Lead chromate induced apoptosis.
This study was focused on chromate.
None Lead chromate induced apoptosis in a concentration-
dependent manner.
None Lead chromate inhibited cell growth.
None Lead glutamate had no effect on growth.
None Lead glutamate induced a concentration-dependent increase
in intracellular lead ions.
Lead glutamate increased the mitotic index, but either had
no effect or inhibited growth and induced mitotic arrest.
Reference
Cheng et al.
(2002)
Apostoli et al.
(2000)
Apostoli et al.
(2000)
Apostoli et al.
(2000)
Blankenship
etal. (1997)
Singh et al.
(1999)
Holmes et al.
(2005)
Wise et al. (2005)
Wise et al. (2005)
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Table AX5-6.19 (cont'd). Genotoxic/Carcinogenic Effects of Lead - Mitogenesis Human and Animal Cell Culture Studies
Compound
Lead Nitrate
Lead Nitrate
Lead Nitrate
Assay (Concentration and
Exposure Time)
Mitotic Index
(3-30 uM for 2h +16 h recovery)
Mitotic Index
(0.05- 1 uM for 3-12 h)
Apoptosis (15-240 uM for 3 h)
Cell Type and
Culture Medium
CHO cells in EMEM +
10%FBS
CHO AA8 in DMEM
+10%NCS
Rat Alveolar
Macrophages in DMEM
+ 10% FBS
Co-exposure
None
Crown ethers to modify
effect through chelation
and uptake
None
Effects
Lower concentrations (1 and 3 uM) of lead nitrate
significantly increased the mitotic index. Higher
concentrations (10 and 30 uM) had no effect.
Lead nitrate dramatically reduced the mitotic index at 1 uM
though this was not statistically analyzed. There was no
effect on mitotic index at lower concentrations. Crown
ethers had no modifying effect.
Lead nitrate induced apoptosis in a dose-dependent manner.
Reference
Linetal. (1994)
Cai and Arenaz
(1998)
Shabani and
Rabbani (2000)
>
X
H
6
o
Abbreviations:
G12 - CHV79 are derived from V79;
V79 are a Chinese Hamster Lung Cell Line;
hTERT = hTERT is the catalytic subunit of human telomerase
Medium and Components
AMEM = Alpha Minimal Essential Medium;
DMEM = Dulbecco's Minimal Essential Medium;
DMEM/F12 = Dulbecco's Minimal Essential Medium/Ham's F12;
EMEM = Eagle's Minimal Essential Medium;
BSA = Bovine Serum Albumin
CCS = Cosmic Calf Serum
FBS = Fetal Bovine Serum
FCS = Fetal Calf Serum
NCS = Newborn Calf Serum
Differences between the serum types are unclear as insufficient details are provided by authors to distinguish.
O
H
O
c
o
H
W
O
V
O
HH
H
W
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Table AX5-6.20. Genotoxic/Carcinogenic Effects of Lead - Mitogenesis Other
>
X
H
O
O
o
H
O
O
H
W
O
O
HH
H
W
Compound
Lead Acetate
Lead Acetate
Lead Acetate
Lead Chloride
Lead Chloride
Assay (Concentration and
Exposure Time)
Production of reactive oxygen
species (1 mM for 180 min)
Glutathione levels
(1 mM for 0-1 80 min)
Catalase Activity
(500-2,000 uM for 24 h)
Thiol Levels
(100 uM for 30 min- 4 h)
Oxidative Metabolism
(0.1-100 uM for 20 h)
Phagocytosis (0.1-100 uM for
20 h)
Oxidative Enzyme Levels
(0.1-1 uMfor Ih)
Cell Type and
Culture Medium
SH-SY5Y- Human
neuroblastoma cells in
DMEM + 7% FCS
Human Foreskin
Fibroblasts (Chinese) in
DMEM +10% FCS
HeLa in HEPES/glucose
buffer
Macrophages from NMRI
mice in EMEM (serum
not given)
AS52-CHO-gpt, lackhprt
in HBSS followed by
Ham'sF12 + 5%FBS
Co-exposure
Glutamate (1 mM)
or PKC inhibitor
(IjiM)
3-aminotriazole
(3-AT) (80 mM to
inhibit catalase)
Buthionine
sulfoximine
(BSO) to deplete
cells of thiols
Zymosan and
latex particles as
substrates for
phagocytosis
Allopurinol
(50 uM) to inhibit
xanthine oxidase
Effects
Lead acetate alone did not produce reactive oxygen species.
Glutamate alone did.
Lead acetate plus glutamate increase glutamate induced increases in
reactive oxygen species.
Lead acetate alone did not deplete glutathione. Glutamate alone did.
Lead acetate plus glutamate decreased glumate- induced decrease in
glutathione.
Lead acetate had no effect on catalase activity.
Lead acetate only lowered thiols marginally
Lead inhibited oxidative metabolism.
Lead inhibited phagocytosis, but only significantly at the highest
dose.
Lead chloride at low concentrations produced H2O2 at 1 h and not at
24 h. Lead chloride at high concentrations produced no change at 1
h and increased H2O2 at 24 h. Allopurinol inhibited H2O2 formation
at high lead concentrations.
Lead chloride had no effect on catalase, glutathione peroxidase,
glutathione reductase. Lead chloride inhibited glutathione-S-
transferase, CuZn-superoxide dismutase, and xanthine oxidase.
Reference
Naarala et al.
(1995)
Hwua and
Yang (1998)
Snyder and
Lachmann
(1989)
Hilbertz et al.
(1986)
Ariza et al.
(1998)
Abbreviations:
AS 52 are derived from CHO;
CHO are a Chinese Hamster Ovary Cell Line;
Medium and Components
DMEM = Dulbecco's Minimal Essential Medium;
EMEM = Eagle's Minimal Essential Medium;
HBSS = Hank's Balanced Salt Solution
FBS = Fetal Bovine Serum
FCS = Fetal Calf Serum
Differences between the serum types are unclear as insufficient details are provided by authors to distinguish.
-------
ANNEX TABLES AX5-7
May 2006 AX5-111 DRAFT-DO NOT QUOTE OR CITE
-------
Table AX5-7.1. Light Microscopic, Ultrastructural, and Functional Changes
to
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Author
Animal Species
Lead Dosage
Blood Lead
Findings
>
X
Khalil-Manesh Sprague-Dawley 0.5% Pb acetate in drinking water Max 125.4 ug/dL
etal. (1992a) rat for 12 mo. Mean55ug/dL
Khalil-Manesh
etal. (1992b)
Khalil-Manesh
etal. (1993a)
Sprague-Dawley
rat
0.5% Pb discontinued after 6 mo
0.01% Pb discontinued after 6 mo
DMSA 0.5% used in 1/2
Sprague-Dawley 0.01% Pb acetate for 12 mo.
rat
Hi Pb @12 mo
Disc 30.4 ug/dL
Disc + DMSA 19.1 ug/dL
Ctrl 3.1 ng/dL
Lo Pb@12mo
Disc 6.9 ug/dL
DMSA5.5 ug/dL
Max 29.4 ug/dL
Range 9-34 ug/dL
Hyperfiltration at 3 mo. Decreased filtration at 12 mo.
NAG and GST elevated.
Nuclear inclusion bodies at all times.
Tubulointerstitial scarring from 6 mo.
No arterial or arteriolar pathology.
High Pb:
Nuclear inclusion bodies prominent.
Tubulointerstitial disease severe but less than 12 mo
continuous DMSA caused reduction in nuclear inclusion
bodies and tubuloint decrease, and an increase in GFR.
LowPb:
Neg pathology and increase in GFR with DMSA
GFR increased at 1 and 3 mo.
NAG increased but GST normal.
Pathology neg except at 12 mo-mild tubular atrophy and
interstitial fibrosis seen.
H
6
o
o
H
O
O
H
W
O
O
HH
H
W
Sanchez-
Fructuoso et al.
(2002a)
Wistar rat
Sanchez- Wistar rat
Fructuoso et al.
(2002b)
Papaioannou et al. Dogs
(1998)
500 ppm (0.05%) Pb acetate for 2
mo, then EDTA
500 ppm (0.05%) Pb acetate for 2
mo, then EDTA
12 mg Pb acetate i.p. x 10
Max 52.9 ug/dL
Day 90: 33.2
Day 137:22.8
Ctrl: 5.90 ug/dL
Max 52.9 ug/dL
Day 90: 33.2
Day 137:22.8
Ctrl: 5.90 ug/dL
Rats given Pb to day 90, then treated with EDTA or untreated
today 137.
Marked decrease in kidney, liver, and brain Pb with EDTA but
no change in femur Pb
Hypertrophy and vacuolization of medium and small arteries,
mucoid edema and muscular hypertrophy of arterioles, include
bodies and fibrosis.
EDTA slowed progression.
Lead includes bodies intracytoplasmically in mesothelial and
giant cells of peritoneum and in interstitial connective tissue
cells of kidney. None in prox tubules of kidney.
-------
Table AX5-7.1 (cont'd). Light Microscopic, Ultrastructural, and Functional Changes
to
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X
OJ
DRAFT-DO I
^
0
H
O
O
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W
O
O
H
W
Author
Vyskocil et al.
(1989)
Vyskocil et al.
(1995)
Vyskocil and
Cizkova(1996)
Sanchez et al.
(2001)
Herak-
Kramberger et al.
(2001)
Fujiwaraet al.
(1995) and Kaji
etal. (1995)
Animal Species Lead Dosage Blood Lead Findings
Wistar rat 0.5%,1%, and 2% Pb acetate for 2-3 0.5%-105 ug/dL 0.5%-no morphologic or functional changes
mo. 1%- 196 ug/dL 1%-Incr in P-2 microglobulin excretion.
2%-320 ug/dL 2%-Incr in P2micr, glucose, protein, lysozyme, and LDH.
Hyperplasia and include bodies of prox tubules seen in both
I%and2%
Wistar rat 1% or 0. 1% Pb acetate for 2-4 mo. 1%- 173 ug/dL 1% caused increase in P-2 microglobulin excretion and injury
0.1%-37.6 ug/dL to proximal tubule.
0.1% caused no changes.
Wistar rats Unleaded petrol vapor (4mg/m3) B-2 microglobulin excretion increased at 60 days
8 hrs/day for 60 days
Sprague-Dawley 0.06% Pb acetate for 4 mo. 13.9 ug/dL vs. <0.5 ug/dL Decrease in expression of laminin-1 and increase in expression
rat in ctrl of fibronectin in kidneys.
Rat brush border 500 uM Pb 58% loss of sealed brush border membrane vesicles. Lower
membranes loss of sealed basolateral membrane vesicles.
Bovine cultured 0.5 - 10 uMPb nitrate Stimulated proliferation in smooth muscle cells. Reduced
vascular smooth proliferation in endothelial cells No leakage of LDH.
muscle and
endothelial cells
-------
Table AX5-7.2. Lead and Free Radicals
to
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!>
X
^
£
U
!>
-n
H
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!2
O
H
/C)
r*^S
O
H
W
O
H
W
Author
Pereira et al.
(1992)
Somashekaraiah
etal. (1992)
Bondy and Guo
(1996)
Blazka et al.
(1994)
Quinn and Harris
(1995)
Ercal etal. (1996)
Vaziri and co-
workers (1997-
2004)
Farmand et al.
(2005)
Gurer et al.
(1999)
Animal Species
Rats
Chick embryos
Sprague-Dawley
rat cerebral
synapto somes
Mouse brain
micro vascular
endothelial cell
culture
Rat cerebellum
homogenates
C57BL/6 mice
Sprague-Dawley
rats
Sprague-Dawley
rats
Fischer 344 rats
Lead Dosage
ALA-treated (40 mg/kg every
2 days for 15 days)
1.25 and 2.5 umol/kg of Pb acetate
0. 5 mMPb acetate
10, 100, and 1,000 nM Pb acetate
17-80 nMPb nitrate
1300 ppm Pb acetate for 5 weeks.
Nac, 5.5 mmol/kg, or DMSA,
1 mmol/kg, given in 6th week.
See Section 5.5 for details
100 ppm Pb acetate for 3 months
1 100 ppm Pb acetate for 5 wks.
Captoprilfor6thwk
Blood Lead Findings
Fatigued earlier than controls.
Increase of CuZn SOD in brain, muscle and liver
Lipoperoxides maximal at 9 hrs and returned to normal at 72
hrs.
GSH depleted. GST, SOD and catalase increased in liver,
brain and heart at 72 hrs
Generation of ROS not increased by Pb alone but increased
when 50 uM iron sulfate added.
Constitutive production of nitrite, but not inducible, decreased
by Pb. Extracellular calcium abolishes this effect.
—
Constitutive NOS activity inhibited 50% by 17 nM Pb and
— 100% by 80 nM Pb. Reversed by increasing Ca concentration.
36.5 ug/dL in Pb-treated; Liver and brain GSH depleted by Pb and MDA increased.
13.7 ug/dL in Pb+ Both were restored by either DMSA or NAC. However,
DMSA-treated. DMSA reduced blood, liver, and brain Pb levels while NAC
did not.
Variable. See section 5.5 for details.
CuZnSOD activity increased in kidney. CuZnSOD activity
increased in aorta whereas protein abundance unchanged.
Guanylate cyclase protein abundance in aorta decreased.
24.6ug/dL in Pb-treated. MDA in liver, brain, and kidney increased by Pb. GSH
23.8 ug/dL in Pb + decreased. Captopril reversed these findings.
Captopril-treated
-------
Table AX5-7.2 (cont'd). Lead and Free Radicals
to
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o
Author
Animal Species
Lead Dosage
Blood Lead
Findings
Acharya and
Acharya(1997)
Swiss mice
200 mg/kg Pb acetate i.p. x 1
MDA-TBA increased x 4 in liver, brain, kidney, and testis by
end of 1st wk and persisted for 4 wks.
Upasani et al.
(2001)
Rats
100 ppm Pb acetate for 30 days.
Groups given vit C, vit E, or algae
MDA, conj dienes, and H2C>2 increased in liver, lung, and
kidney by Pb. Treatment with vit C, vit E, or Blue Green algae
reversed these findings.
Pande et al. Wistar rats Lead nitrate 50 mg/kg i.p. x 5
(2001) Lead + DMSA,MiADMSA,NAC,
DMSA + NACTJMSA +
MiADMSA
DMSA most effective in blocking inhib of ALAD, elev of
ZPP, and inhib of GSH. Combined DMSA+NAC most
effective when given during or post-exposure.
>
X
Pande and Flora Wistar rats
(2002)
2000 ppm Pb acetate x 4 wks.
DMSA,MiADMSA, DMSA +
LA,MiADMSA + LA x 5 days
Lead caused decrease in ALAD, GSH, and increase ZPP.
Lipoic acid (LA) did not chelate Pb in contrast to DMSA, but
both agents increased ALAD and GSH
Flora et al. (2002) Wistar rats
1000 ppm Pb acetate x 3 mo.
DMSA or MiADMSA + vit C or
vit E x 5 days
13.3 ng/dL
lead Rx
3 ng/dL DMSA Rx
-------
Table AX5-7.2 (cont'd). Lead and Free Radicals
N>
O
Author
Sivaprasad et al.
(2002)
Animal Species Lead Dosage
Wistar rats 2000 ppm Pb acetate x 5 wks.
LA and DMSA during 6th week.
Blood Lead Findings
Lead caused red in kidney GGT & NAG, decline in GSH,
catalase, SOD, GPx and Glut reductase, and increased MDA.
Lipoic acid
+DMSA restored these changes
Senapati et al. Rats
(2000)
Patra et al. (2001) IVRI 2CQ rats
1% sol of 5mg/kg Pb acetate x43 d.
Thiamine 25 mg/kg
1 mg/kg Pb acetate for 4 wks.
Vit E, vit C or methionine in 5th wk.
VitE + EDTA.
6.8 ng/dL Pb-Rx
6.3 ng/dL
lead, vit E +EDTA
Thiamine reduced Pb content and MDA levels of both liver
and kidney and improved pathology.
Lead in liver, kidney and brain reduced by vit E + EDTA
treatment. MDA increased by Pb in all 3 organs but decreased
by vit E + EDTA.
>
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McGowan and Chicks
Donaldson (1987)
2000 ppm Pb acetate x 3 wks.
GSH, non-protein SH, lysine and methionine increased in liver
and non-prot SH, glycine, cysteine and cystathionine in
kidney. Cysteine reduced in plasma.
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HH
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Table AX5-7.3. Chelation with DMSA
to
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Author
Cory-Slechta
(1988)
Pappas et al.
(1995)
Animal Species
Rats
Sprague-Dawley
rats
Lead Dosage
50 ppm Pb acetate for 3-4 mo.
550-1 100 ppm Pb acetate for 35
days
Blood Lead
20 ng/dL-lead
1 (ig/dL-lead+25 mg/kg
DMSA
52 ng/dL @ 550 ppm Pb
Findings
DMSA 25-50 mg/kg i.p. for 1-5 d mobilized Pb from blood,
brain, kidney and liver, but not femur.
DMSA @16-240 mg/kg/day p.o. for 21 days given with and
without concurrent Pb exposure. Rats showed dose-related
Smith and Flegal Wistar rats
(1992)
lead
206Pb 210 ng/mL for l.Sdays.DMSA 5.1 ng/g-ctrl
20 mg/kg i.p. 3.0 ng/g-DMSA
reduction in Pb content of blood, brain, femur, kidney, and
liver with or without concur Pb.
Rats on low Pb diet given DMSA decreased soft tissue but not
skeletal Pb. Lead redistributed to skeleton.
!> Varnai et al.
X (2001)
3
Wistar rats
(suckling)
2 mg/kg/d for 8 d
DMSA 0.5 mmol/kg 6x/d on dl-3
and 6-8
DMSA reduced Pb concentration in carcass, liver, kidneys, and
_ brain by ~ 50%.
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O
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Table AX5-7.4. Effect of Chelator Combinations
o
0-1 Author
Flora et al.
(2004)
Jones et al.
(1994)
Animal Species Lead Dosage
Wistar rats 1000 ppm Pb acetate for 4 mo
Mice 10 i.p. injections of Pb acetate, 5.0
mg/kg
Blood Lead
46 ng/dL-lead
12.8 ng/dL-combined Rx
Findings
5 days Rx with DMSA. CaNa2EDTA, or DMSA +
CaNa2EDTA. Comb Rx resulted in increased ALAD &
decreased Pb in blood, liver, brain, and femur.
Mice Rx'ed with DMSA, CaNa2EDTA, ZnNa2EDTA, or
ZnNa3DTPA 1 .0 mmol/kg/d 4-8 d. CaNa2EDTA most
effective in removing brain Pb; DMSA in removing kidney
and bone Pb.
>
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Table AX5-7.5. Effect of Other Metals on Lead
to
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Author
Animal Species
Lead Dosage
Blood Lead
Findings
>
X
H
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o
H
O
O
H
W
O
O
HH
H
W
Maldonado-Vega Wistar rats
etal. (1996) (pregnant &
non-pregnant)
Olivi et al. (2002) MDCK canine
kidney cells
Bogden et al.
(1991)
Wistar rats
Skoczynska et al. Buffalo rats
(1994)
Othman & Albino rats
Missiry (1998)
Tandon et al.
(1992)
Albino rats
Flora et al. (1989) Albino rats
Flora et al. (1994) Wistar albino rats
100 ppm Pb acetate for
144-158 days
0,1,100 ppm Pb for 31 wks
0.2% or 4.0% Ca diet
Pb 70 mg/kg 2x/wk for 7 wks Cd
20 mg/kg Ix/wk for 7 wks.
All intragastric.
Pb acetate 100 (jmol/kg I.M. x 1
Se 10 (imol/kg I.M. 2 hrs before Pb
Pb acetate 10 mg/kg/d p.o. x 6 wks.
EDTA or DTPA given for 5 d w or
w/o Se
Pb acetate 10 mg/kg/d p.o. x 6 wks
Thiamine, Zn or thiamine + Zn x 6
wks
Pb acetate 10 mg/kg/d x 56 d p.o.
EDTA or EDTA + Zn x 5 days p.o.
5.2 (ctrl) to 27.3 ng/dL in
Pb-exposed
8 (non-preg)to 17 ng/dL
in rats exposed only
during lactation
1.9 to 39.1 |ig/dLonlow
Ca diet and 2.0 to 53.3
Hg/dL on high Ca diet
5.1 to 29.6 ng/dL in Pb-
exposed. 37.4 ng/dL in
Pb + Cd
17tol38|ig/dLafterPb
58 ng/dL after EDTA.
50 ng/dL after EDTA + Se
6.2 to 120.9 ng/dL after
Pb
44.1 ng/dL after thiamine
+ Zn
4.6to43.0|ig/dLinPb.
22.5|ig/dLinEDTA
16.5|ig/dLinEDTA +
Zn.
Lead administered to period before lactation (144 d) or to mid-
lactation (158 d).Lead in blood, kidney, liver, and bone
increased.
ALAD decreased and FEP incr. Lactation increased
Blood Pb from 24.7 to 31.2 ng/dL and decreased bone Pb from
83.4 to 65.2 nmol/g.
In response to agonists ADP or bradykinin levels of
intracellular Ca increased
3-fold and 2-fold. Lead inhibited the response.
At 100 ppm Pb high Ca diet produced higher BP and more
renal cancers than low Ca diet and higher levels of Pb in brain,
liver, bone, heart, and testis but lower levels in kidney. Serum
Ca on high Ca diet was 13.2 mg/dL.
Simultaneous Pb and Cd administration increased blood Pb but
decreased Pb in liver and kidneys as compared to Pb
administration alone.
Sodium selenite (Se is a well-known anti-oxidant) prevented
lipid peroxidation (TEARS) and reduction in GSH caused by
Pb. SOD & glut reductase also normalized.
Selenium had no additional benefit over chelators except for
higher ALAD and lower ZPP in blood, lower Pb in liver and
kidney.
Thiamine given as 25 mg/kg/d and Zn sulfate as 25 mg/kg/d.
ALAD restored by combined Rx. Liver and kidney Pb affected
to a minor degree but brain Pb not affected.
CaNa2EDTA given as 0.3 mmol/kg/d i.p. and Zn sulfate as 10
or 50mg/kg/d. ALAD partially restored after EDTA + Zn but
not after EDTA. EDTA reduced Pb in bone, kidney, and liver
but not in brain. Zn cone increased in blood, kidney, & brain
by 50 mg Zn dosage.
-------
Table AX5-7.5 (cont'd). Effect of Other Metals on Lead
to
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>
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ANNEX TABLES AX5-8
May 2006 AX5-5-121 DO-NOT QUOTE OR CITE
-------
Table AX5-8.1. Bone Growth in Lead-exposed Animals
to
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Compound
Dose/Concentration
Duration Exposure
Route
Species
Effects
Blood Level
Reference
>
X
to
to
Lead Acetate
41.7mgPb/l
83.3 mg Pb/1
166.6 mgPb/1
12 to 16 weeks
Drinking water
Lead aerosol
77,249,orl546ug/m3
for 50 to70 days
Inhalation
Rat Lead level in bone of control animals
Wk 0 = 1.3 ± 0.83 ug Pb/g; Wk4 = 1.2 ± 0.99 ng Pb/g;
Wk 8 = 1.3 ± 1.08 ug Pb/g; Wk 12 = 0.8 ± 0.13 ug Pb/g;
Wkl6 = 1.3±0.95ngPb/g
Lead level in bone of animals receiving 41.7 mg Pb/1
Wk 0 = 1.0 ± 0.50 ng Pb/g; Wk4 = 5.9* ± 1.76 ng Pb/g;
Wk 8 = 2.9* ±1.15 ug Pb/g; Wk 12 = 6.2* ±1.01 ^g Pb/g;
Wk 16 = 6.0* ± 0.75 ug Pb/g
Lead level in bone of animals receiving 83.3 mg Pb/1
Wk 0 = 2.0 ± 0.97 ug Pb/g; Wk4 = 11.7* ± 3.56 ug Pb/g;
Wk 8 = 8.8* ± 3.37 ug Pb/g; Wk 12 = 14.3* ± 4.29 ng Pb/g
Lead level in bone of animals receiving 166.6 mg Pb/1
Wk 0 = 0.9 ± 0.23 ng Pb/g; Wk 4 = 17.0* ± 3.89 ug Pb/g;
Wk 8 = 35.7* ± 3.64 ug Pb/g; Wk 12 = 21.7* ± 5.11 ug Pb/g;
* significantly higher than control animals at corresponding time point
Rat 16.9 ± 6.6 ug Pb/g bone taken up in animals exposed to 77 ug/m3 for
70 days versus 0.2 ± 0.2 ug Pb/g in control animals;
15.9 ± 4.3 ug Pb/g bone in rats exposed to 249 ug/m3 for 50 days;
158 ± 21 ug Pb/g bone in rats exposed to 1546 ug/m3 for 50 days
Not given
Hac and Kruchniak
(1996)
Control: 2.6 ug/dL
77 ug/m3: 11.5 ug/dL
249 ug/m3: 24.1 ug/dL
1546 ug/m3: 61.2 ug/dL
Grobleretal. (1991)
H
6
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o
H
O
O
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W
O
O
HH
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W
Lead acetate
250 ppm or 1000 ppm
7 weeks to females
prior to mating,
continuing through
gestation and lactation
Drinking water
Rat Offspring body weight was depressed relative to controls during
suckling (Day 11) and after weaning (Day 24) in high dose and
continuously lead-exposed groups.
Continuous lead exposure caused a greater decrease in offspring body
weight than lead exposure only prior to or after parturition.
Decreased tail length growth suggested possible effects of lead on tail
vertebral bone growth.
Dams prior to mating:
Control = 2.7 ± 0.6 ug/dL
250 ppm =39.9 ±3.5 ug/dL
1000 ppm = 73.5 ±9.3 ug/dL
Hamilton and
O'Flaherty(1994)
-------
Table AX5-8.1 (cont'd). Bone Growth in Lead-exposed Animals
to
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Compound
Dose/Concentration
Duration Exposure
Route
Species
Effects
Blood Level
Reference
>
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to
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O
O
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O
O
HH
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Lead acetate
Hi Pb animals 5000
ppm for 6 months,
reduced to 1000 ppm;
Lo Pb animals 100 ppm
Drinking Water
Rat
Lead acetate
17 mg per kg of feed
50 days
In diet
Lead acetate
17 mg per kg of feed
50 days
In diet
Rat
Rat
In male rats exposed to 100 ppm lead in drinking water and a low
calcium diet for up to one year, bone density was significantly
decreased after 12 months, while rats exposed to 5000 ppm lead had
significantly decreased bone density after 3 months. Lead content of
femurs was significantly elevated over the content of control rats at all
time points (1, 3, 6, 9, 12 months). Trabecular bone from the low dose
animals was significantly decreased from 3 months forward.
No differences in the length of the femurs, but the mean length of the
5th lumbar vertebra was significantly decreased. The mean length of
the femur growth plate cartilage was also significantly decreased in
lead-exposed animals.
No differences in the length of the femurs, but the mean length of the
5th lumbar vertebra was significantly decreased. The mean length of
the femur growth plate cartilage was also significantly decreased in
lead-exposed animals.
LowPb(|ig%):
1 month
Control = 2 ± 1; Exp = 19 ± 10*
3 months
Control = 2 ± 1; Exp = 29 ± 4*
6 months
Control = 3 ± 1; Exp = 18 ± 2*
9 months
Control = 1 ± 1; Exp = 17 ± 3*
12 months
Control = 3 ± 1; Exp = 21 ± 3*
Hi Pb (ng%):
1 month
Control = 3 ± 1; Exp = 45 ± 13*
3 months
Control = 3 ± 1; Exp = 90 ± 15*
6 months
Control = 4 ± 1; Exp = 126 ± 10*
9 months
Control = 4 ± 1; Exp = 80 ± 39*
12 months
Control = 3 ± 1; Exp = 59 ± 18*
*p< 0.001
Not given
Not given
Gruberetal. (1997)
Gonzalez-Riola et al.
(1997)
Escribano et al. (1997)
-------
Table AX5-8.1 (cont'd). Bone Growth in Lead-exposed Animals
to
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o
Compound
Dose/Concentration
Duration Exposure
Route
Species
Effects
Blood Level
Reference
>
X
to
Lead acetate
0.6%
GD 5 to Adulthood
(various)
In drinking water
Rat Early bone growth was significantly depressed in a dose-dependent
fashion in pups of lead-exposed pups, with growth suppression in male
offspring considerably greater than females. Significant decreases in
plasma insulin-like growth factor and plasma sex steroids and
increased pituitary growth hormone were also observed.
Groups:
DDW = Dams and pups received distilled deionized water entire study
Ac/Ac = Dams and pups received acetic acid solution entire study
Preg = Dams received 0.6% lead water from GD 5 to parturition
Lact = Dams received 0.6% lead water during lactation only
P + L = Dams received 0.6% lead water from GD 5 through lactation
Postnatal = Dams and pups received 0.6% lead water from parturition
through adulthood
Pb/Pb = Dams and pups received 0.6% lead water from GD 5 through
adulthood
Whole blood lead (ug/dL) in
male/female offspring at age Day
85:
DDW =5.5 ±2.0/6.8 ±1.5;
Ac/Ac = 1.9 ±0.2/1.4 ±0.3;
Preg = 9.1±0.7*/11.6±4.6*;
Lact =3.3 ±0.4/3.4 ±0.8;
P + L= 16.1 ± 2.3*710.4 ± 1.8*;
Postnatal = 226.0 ± 29.0*/292.0 ±
53.0*;
Pb/Pb = 316.0 ±53.0*/264.0 ±
21.0*
*p< 0.05 compared to Ac/Ac
group.
Ronisetal. (1998a)
H
6
o
o
H
O
O
H
W
O
O
HH
H
W
Lead acetate Rat
0.05% to 0.45%
GD 5 through sacrifice
of pups at 21, 35, 55,
and 85 days
In drinking water
Lead nitrate Rat
0.02% (125 ppm)
GD5 to 1 day before
sacrifice
In drinking water
Early bone growth was significantly depressed in a dose-dependent
fashion in pups of all lead-exposed groups, with growth suppression in
male offspring considerably greater than females. Significant
decreases in plasma insulin-like growth factor and plasma sex steroids
and increased pituitary growth hormone were also observed.
Between age 57 and 85 days growth rates were similar in control and
lead-exposed pups, suggesting exposure at critical growth periods such
as puberty and gender may account for differences in growth reported
by various investigators.
Exposure to 0.02% lead nitrate (125 ppm lead) did not significantly
affect growth, though males weighed significantly less than females.
Offspring:
0.05% Pb = 49 ± 6 ug/dL; 0.15%
Pb = 126 ± 16 ug/dL; 0.45% Pb =
263 ± 28 ug/dL
Ronisetal. (1998b)
Rat Pups
5 days old: 43.3 ±2.7 ug/dL
49 days old: 18.9 ± 0.7 ug/dL
(females: 19.94 ±0.8 ug/dL;
males: 17.00 ±1.1 ug/dL)
Camoratto et al. (1993)
-------
Table AX5-8.1 (cont'd). Bone Growth in Lead-exposed Animals
to
o
o
Compound
Dose/Concentration
Duration Exposure
Route
Species
Effects
Blood Level
Reference
>
X
to
H
6
o
o
H
O
O
H
W
O
O
HH
H
W
Lead acetate
0.15% or 0.45%
GD 4 until Day 55
In drinking water
Lead acetate
lOOOppm
22-26 days
In drinking water
Rat A dose-dependent decrease in load to failure in tibia from lead-exposed
(0.15% and 0.45% lead acetate in drinking water) male pups only.
Hormone treatments (L-dopa, testosterone or dihydrotestosterone in
males, or estradiol in females) failed to attenuate lead deficits during
the pubertal period. Distraction osteogenesis experiments performed
after stabilization of endocrine parameters (at 100 days of age) found
decreased new endosteal bone formation and gap x-ray density in the
distraction gaps of lead-exposed animals.
Rat Lead disrupted mineralization during growth in demineralized bone
matrix implanted subcutaneously into male rats. In the matrix that
contained 200 micrograms lead/g of plaque tissue, alkaline phosphatase
activity and cartilage mineralization were absent, though calcium
deposition was enhanced. Separate experiments found enhanced
calcification and decreased alkaline phosphatase activity in rats
implanted with a control (no lead) matrix and given 1000 ppm lead in
drinking water for 26 days.
Offspring:
0.15%Pb = 67-192 ng/dL; 0.45%
Pb = 120-388 ng/dL
Ronisetal. (2001)
Blood Pb(ngML)
Control:
Implantation Day 0 = 1.3 ± 0.6;
Day 8 = 2.2 ± 0.9; Day 12 = 2.1 ±
0.7.
Lead added to matrix:
Implantation Day 0 = 1.5 ± 0.8;
Day 8 = 5.7 ± 0.8a'b; Day 12 = 9.5
± 0.5a'b.
Lead in drinking water:
Implantation Day 0 = 129.8 ± 6.7
a; Day 8= 100.6 ± 6.8 a'b; Day 12
= 96.4±5.3a'b.
a Significant (p<0.05) difference
from control.
b Significance (p<0.05) difference
from corresponding value at
implantation (Day 0).
Hamilton and
O'Flaherty(1995)
Abbreviations
Mg-
ppm-
GD-
Pb-
g-
ng%-
milligram
microgram
parts per million
gestational day
lead
gram
microgram percent
Exp
wk-
dL-
liter
cubic meter
experimental group
week
deciliter
percent
-------
Table AX5-8.2. Regulation of Bone Cell Function in Animals - Systemic Effects of Lead
to
o
o
Compound
Dose/Concentration
Duration Exposure
Route
Species
Effects
Blood Level
Reference
>
X
to
Oi
H
6
o
o
H
O
O
H
W
O
O
HH
H
W
Lead acetate
30 mg/kg
Single IV injection
Lead acetate
0.82%
1 week
In diet
Rat
Rat
Lead acetate
0.15% or 0.45%
GD 4 until Day 55
In drinking water
Rat
Groups of male rats were killed 0.5, 5,15, and 30 min and 1, 2, 6, and 12 h after the single
lead injection. Serum calcium and phosphorus levels both initially increased after lead
injection with serum phosphorus reaching a maximum value (13.5 mg%) after 30 min and
calcium (17 mg%) after 1 h. Calcium and phosphorus levels decreased after 1 h and
returned to baseline levels after 12 h.
Ingestion of 0.82% lead in male rats fed either a low phosphorus or low calcium diet
reduced plasma levels of 1,25-(OH)2CC, while lead had no effect in rats fed either a high
calcium diet or a normal phosphorus diet.
Effect of lead on serum 1 ,25-(OLTbCC levels in rats fed low P or normal P diet
Dietary Phosphorus
0.1%
0.1%
0.1%
0.3%
0.3%
0.3%
Supplement
Control
Cholecalciferol
0.82% Pb+Cholecalciferol
Control
Cholecalciferol
0.82% Pb+Cholecalciferol
Serum 1,25-(OH)2CC
<10 pg/mL
248 ± 7 pg/mL
94 ± 13pg/mL
<10 pg/mL
285 ± 44 pg/mL
245 ± 46 pg/mL
Effect of lead on serum 1,25-(OLTbCC levels in rats fed low Ca or high Ca diet
Dietary Calcium
0.02%
0.02%
0.02%
1.2%
1.2%
1.2%
Supplement
Control
Cholecalciferol (50ng/day)
0.82% Pb+Cholecalciferol
Control
Cholecalciferol (50ng/day)
0.82% Pb+Cholecalciferol
Serum 1,25-(OH)2CC
<10 pg/mL
754±18pg/mL
443 ± 79 pg/mL
<10 pg/mL
285 ± 44 pg/mL
245 ± 46 pg/mL
No effects of lead on plasma concentrations of vitamin D metabolites, 25-OH D3 or 1,25-
(OH2)D3, in pubertal male rats exposed to either 0.15% or 0.45% lead acetate in drinking
water and maintained on an adequate diet.
Not given
Kato et al.
(1977)
Smith et al.
(1981)
ug/lOOmL
3±1
9±8
352 ± 40
<3
<3
284 ± 36
ug/lOOmL
<3
<3
284 ± 36
Offspring:
0.15%Pb = 67-192
ug/dL; 0.45% Pb =
120-388 ug/dL
Ronisetal. (2001)
-------
Table AX5-8.2 (cont'd). Regulation of Bone Cell Function in Animals - Systemic Effects of Lead
to
o
o
>
X
to
Compound
Dose/Concentration
Duration Exposure
Route
PbCl2
0,0.2, or 0.8%
1 or 2 weeks
In diet
PbCl2
0,0.2, or 0.8%
1 or 2 weeks
In diet
Lead acetate
l%for 10 weeks or
0.00 1-1% for 24 weeks
In drinking water
Species Effects
Chicks Compared with control animals, lead exposure significantly increased intestinal calbindin
protein and mRNA levels in addition to plasma 1,25-dihydroxyvitamin D concentration.
The effect was present after 1 week of exposure and continued through the second week.
In calcium-deficient animals increased plasma 1,25-dihydroxyvitamin D and calbindin
protein and mRNA were significantly (p < 0.05) inhibited by lead exposure in a dose
dependent fashion over the 2 week experimental period.
Chicks Dose dependent increases in serum 1,25-(OH2)D3 levels (and Calbindin-D protein and
mRNA) with increasing dietary lead exposure (0. 1% to 0.8%) in experiments performed on
Leghorn cockerel chicks fed an adequate calcium diet.
Rat Short term administration of 1% lead resulted in significant increases in bone lead. Total
serum calcium and ionized serum calcium were significantly decreased, as compared to
controls. Circulating levels of 1 ,25-(OH2)D3 were also decreased, though the rats were
maintained on a normal calcium diet (0.95%). In the long term study, a dose-dependent
increase in parathyroid weight occurred with increasing exposure to lead in drinking water.
Blood Level
None given
None given
Short term (10
week) study:
Control:
< 0.02 ng/1
Lead-exposed:
> 5ng/l
Reference
Fullmer (1995)
Fullmer (1996)
Szaboetal. (1991)
H
6
o
o
H
O
O
H
W
O
O
HH
H
W
Short term (10 weeks) exposure
Serum Calcium (mM) Ionized Calcium
(mM)l,25(OH)2D3(pM)
Parathyroid Weight (jig/gland)
*p<0.01
Long term (24 weeks) exposure
Pb in water
Controls
2.42 ±0.03
1.25 ±0.03
232 ±18.9
96 ±34
Normalized Parathyroid
Weight (|ig/g body wt)
Lead-exposed
2.32 ±0.02*
1.15±0.03*
177 ±10.8*
178 ±25*
l,25(OH)2D3(pM)
0%
0.001%
0.01%
0.1%
1.0%
p<0.01
0.50 ±0.06
0.72 ± 0.25
0.81 ±0.28
0.94 ±0.27
0.81 ±0.29*
241 ± 32
188 ±27
163 ±17
206 ± 24
144 ± 33*
-------
Table AX5-8.2 (cont'd). Regulation of Bone Cell Function in Animals - Systemic Effects of Lead
to
o
o
>
X
to
oo
H
6
o
o
H
O
O
H
W
O
O
HH
H
W
Compound
Dose/Concentration
Duration Exposure
Route Species
Lead nitrate Rat
0.02% (125 ppm)
GD5 to 1 day before
sacrifice
In drinking water
Lead acetate Rat
0.05% to 0.45%
GD 5 through sacrifice
of pups at 21, 35, 55,
and 85 days
In drinking water
Abbreviations
mg - milligram
h - hour
Effects
Basal release of growth hormone from control and lead-exposed pups at age 49 days was
not significantly different. Growth hormone releasing factor-stimulated release of growth
hormone from pituitaries of lead-exposed pups was smaller than the stimulated release of
growth hormone from pituitaries of control animals (75% increase over baseline vs. 171%
increase, respectively), but the difference did not achieve significance (P = 0.08). Growth
hormone content of the pituitary glands was also not influenced by lead exposure.
Pituitary GH content (ug/mg) at postnatal day 55:
Control Male Pups = 56.6 ± 8.0; Female Pups = 85.6 ± 9.3
0.05% Pb Male Pups = 107.2 ± 10.5*; Female Pups = 116.2 ±9.1
0.15% Pb Male Pups = 96.8 ± 5.0*; Female Pups = 105.1 ± 7.3
0.45% Pb Male Pups = 106.0 ± 9.8*; Female Pups = 157.0 ± 9.9*
*significantly different from control, p < 0.05
GD - gestational day
mM millimolar
Blood Level Reference
Rat Pups Camoratto et al.
5 days old: 43.3 ± (1993)
2.7 ug/dL
49 days old: 18. 9 ±
0.7 ug/dL
(females: 19.94±
0.8 ug/dL; males:
17.00 ±1.1 ug/dL)
Offspring: Ronis et al.
0.05%Pb = 49±6 (1998b)
ug/dL; 0.1 5% Pb =
126 ±16 ug/dL;
0.45% Pb = 263 ±
28 ug/dL
1,25- (OH)2CC-l,25-dihydroxycholecalcigerol Pb - lead
ug - microgram
pM - picomolar
25 - OH D3- 25-hydroxycholecalciferol IV - intravenous
PbCl2 lead chloride
1 ,25 - (OH)2 D3 - vitamin D3
kg - kilogram
mg% - milligram percent
pg - picogram
% - percent
mL - milliliter
dL deciliter
mRNA - messenger ribonucleic acid
ppm - parts per million
GH - growth hormone
min - minute
-------
Table AX5-8.3. Bone Cell Cultures Utilized to Test Effects of Lead
to
o
o
>
X
to
VO
H
6
o
o
H
O
O
H
W
O
O
HH
H
W
Compound
Dose/Concentration
Duration Exposure
Route
210Pb nitrate
5 uM
20 hours
In medium
Lead acetate
0 to 50 uM
20 h
In medium
Species
Effects
Blood Level
Reference
Stable "Pb"
5 mg/mL in drinking
water given during
gestation.
On GDI 8, SOuCi
210PbgivenIVto
pregnant dams
210Pb nitrate
6.5 to 65 uM
5 min to 2 h
In medium
Rat
(Fetal Bone
Organ
Culture)
Mice
(bone cell
isolation from
calvaria)
PTH (3885 lU/mg bone) enhanced cell-mediated release of 210Pb from bone. Release Not given/Not
of 210Pb was accompanied by proportional loss of stable lead and calcium from treated applicable
bones.
Time: Release of 210Pb (EM/CM ratio)
Omin 1.00
10 min 0.82 ±0.05
2hr 1.12 ±0.04
6hr 1.59 ±0.08*
24 hr 3.69 ±0.15*
48 hr 3.75 ±0.09*
48 hr 0.78 ± 0. 14* (in presence of 30 mU/mL salmon calcitonin)
*Different from 1 .00, p < 0.01 .
Uptake of 210Pb by OC cells rapid. Not applicable
OC cells have greater avidity for lead compared to OB cells.
OC cell uptake of lead almost linear vs. little increase in lead uptake by OB cells with
increasing Pb concentrations in media.
Rosen and Wexler
(1977)
Rosen (1983)
Mice
(osteoclastic
bone cell
isolation from
calvaria)
Mice
(osteoclastic
bone cell
isolation from
calvaria)
15-30% release of 210Pb label occurred in OC cells over 2 h time period.
Physiological concentrations of PTH resulted in marked increase in 210Pb and 45Ca
uptake by OC cells. 210Pb uptake linear over PTH concentrations of 50 to 250 ng/mL).
Media concentrations of lead > 26 uM enhanced calcium uptake by cells.
Three readily exchangeable kinetic pools of intracellular lead identified, with the
majority (approximately 78%) associated with the mitochondrial complex.
Cultures were labeled with Ca (25 uCi/mL) for 2 or 24 h and kinetic parameters
were examined by analysis of 45Ca washout curves.
In kinetic analysis using dual-label (1-2 uCi/mL 210Pb and 25 uCi/mL 45Ca) wash out
curves, the Ca:Pb ratios of the rate constants were approximately 1:1, suggesting
similar cellular metabolism.
Not applicable
Not applicable
Pounds and Rosen
(1986)
Rosen and Pounds
(1988)
-------
Table AX5-8.3 (cont'd). Bone Cell Cultures Utilized to Test Effects of Lead
to
o
o
>
X
H
6
o
o
H
O
O
H
W
O
O
HH
H
W
Compound
Dose/Concentration
Duration Exposure
Route
Lead acetate and 210Pb
label
0-100 uM
20 hours
In medium
Lead acetate
5 or 25 uM
Up to 5 hours
In medium
Lead nitrate
5 uM
20 minutes
In medium
Pb2+
5 or 12.5 uM
Up to 100 minutes
In medium
Species
Mice
(osteoclastic
bone cell
isolation from
calvaria) and
Rat
Osteosarcoma
Cells
(ROS 17/2.8)
Rat
Osteosarcoma
Cells
(ROS 17/2.8)
Rat
(osteoblastic
bone cell
isolation from
calvaria)
Rat
(osteoblastic
bone cell
isolation from
calvaria)
Effects Blood Level
Concentrations as high as 100 uM did not cause toxicity in either cell culture. There Not applicable
was a slight decrease in growth of ROS cells at 5 uM lead concentration and a 50%
decrease in growth at 25 uM lead at day 9.
210Pb washout experiments with both cell cultures indicated similar steady-state lead
kinetics and intracellular lead metabolism. Both cell cultures exhibited one large,
slowly exchanging pool of lead, indicative of the mitochondrial pool.
Used 19F NMR in combination with 1 ,2-bis(2-amino-5-fluorophenoxy )ethane- Not applicable
N,N,N',N'-tetraacetic acid (5F-BAPTA) to distinguish and measure concentrations
of Pb2+ and Ca2+ in aqueous solution.
Basal concentration of [Ca +]i was 128 ± 24 nM. Treatment of cells with 5 and 25
uMPb2+ produced sustained 50% and 120% increases in [Ca2+];, respectively, over a
5 hour exposure period.
At a medium concentration of 25 uM Pb2+ a measurable entry of Pb2+into the cells
([Pb2+], of 29 ± 8 pM) was noted.
Lead (5 uM) linearly raised the emission ratio of FURA-2 loaded cells 2-fold within Not applicable
20 minutes of application, most likely due to increase in [Pb +]i rather than increase
in [Ca2+]r
Intracellular calcium increased even in the absence of extracellular calcium.
5 or 12.5 uM Pb2+ applied simultaneously with re-added calcium reduced immediate Not applicable
CRAC to 70% or 37% of control value, respectively.
During CRAC a large influx of Pb + occurred, leading to a 2.7-fold faster increase in
the FURA-2 excitation ratio. These effects were exclusive of any inhibitory action
of Pb2+on calcium ATPase activity.
Reference
Longetal. (1990a)
Schanne et al.
(1989)
Schirrmacher et al.
(1998)
Wiemann et al.
(1999)
-------
Table AX5-8.3 (cont'd). Bone Cell Cultures Utilized to Test Effects of Lead
to
o
o
Compound
Dose/Concentration
Duration Exposure
Route
Species
Effects
Blood Level
Reference
Lead nitrate
0-150 uM
Up to 72 hours
In medium
Mice (bone
cell isolation
from parietal
bones)
Pb2+ concentrations of 50 |iM and above stimulated release of hydroxyproline and
previously incorporated 45Ca from organ culture. This did not occur in bone
inactivated by freezing and thawing. Eel calcitonin, bafilomycin AI, and scopadulcic
acid B significantly inhibited Pb mediated 45Ca release. There was a high correlation
between 45Ca and PGE2
release (p < 0.001), inferring Pb-induced bone resorption mediated by PGE2. This
was further supported by the significant depression of Pb-stimulated 45Ca release that
occurred with concurrent exposure to 10 uM of either indomethacin or flurbiprofen,
both inhibitors of cyclooxygenase.
Not applicable
Miyahara et al.
(1995)
>
X
Lead acetate
0-25 uM
48 hours
In medium
Rat Osteocalcin production in cells treated with 100 pg 1,25-dihydroxyvitamin D3/mL of
Osteosarcoma medium and 0 uMPb2+for 16, 24, or 36 h was 20.1 ±2.1,23.5 ± 3.4, 26.1 ±2.5 in cell
Cells digests, and 87.2 ± 3.3, 91.6 ± 6.7, 95.1 ± 5.2 in the medium, respectively. The
(ROS 17/2.8) presence of 25 uM Pb2+in the medium, reduced osteocalcin levels to as low as 30% of
control levels.
Cells treated with 0, 5, 10, or 25 uM lead acetate for 24 h, followed by an additional
24 h exposure to 0 or 100 pg of 1,25-dihydroxy vitamin D3 and continued Pb2+
exposure, resulted in a concentration-dependent reduction of 1,25-dihydroxy vitamin
D3-stimulated osteocalcin secretion. 10 uM Pb resulted in medium osteocalcin levels
similar to control levels, however, 25 uM Pb resulted in about a 30% decrease.
Cellular osteocalcin levels were unaffected.
Not applicable
Longetal. (1990b)
H
6
o
o
H
O
O
H
W
O
O
HH
H
W
Lead glutamate
4.5X10-5to4.5X10-
M
2, 4, or 6 days
In medium
Rat In the presence of serum in the cultures, concentrations of Pb2+ less than 4.5 X 10"5 M Not applicable
Osteosarcoma had no effect on cell proliferation. In the absence of serum, 4.5 X 10"7 M Pb2+
Cells increased proliferation at Day 4 and 4.5 X 10"6 M Pb2+ inhibited proliferation at Day 6.
(ROS 17/2.8) Lead exposure for 48 h (4.5 X 10'6 M) significantly (p < 0.01) increased total protein
production in cells and media of cultures labeled with [ H] proline, but did not
increase collagen production. Protein synthesis and osteonectin were enhanced in
cells following Pb + exposure.
Sauketal. (1992)
-------
Table AX5-8.3 (cont'd). Bone Cell Cultures Utilized to Test Effects of Lead
to
o
o
>
X
to
H
6
o
o
H
O
O
H
W
O
O
HH
H
W
Compound
Dose/Concentration
Duration Exposure
Route
Lead glutamate
4.5xlO'5M-10'7M
1,3, or 5 days
incubation
In medium
Lead glutamate
5-20 \iM
48 hours
In medium
Lead
0.5 to 5 uM
40min
In medium
Lead nitrate
5X10'4to5X10-15M
24 h
In medium
Species
Human
Dental Pulp
Cells
Rat
Osteo sarcoma
Cells
(ROS 17/2.8)
Rat
Osteo sarcoma
Cells
(ROS 17/2.8)
Rat
Osteo sarcoma
Cells
(ROS 17/2.8)
Effects Blood Level
All concentrations significantly increased cell proliferation on Day 1, 3 and 5 of Not applicable
exposure in serum free conditions. Lead exposure resulted in dose-dependent
decrease in intracellular protein and procollagen I production over 5 days. In presence
of serum only, 4.5 x 10" M Pb + significantly increased protein production, however, at
that same concentration lead significantly decreased osteocalcin production (i.e.
reduced the level of osteocalcin by 55% at 12 hours).
Cells treated with 0, 5, 10, or 20 uM lead acetate for 24 h, followed by an additional Not applicable
24 h exposure to 0 or 100 pg of 1,25-dihydroxy vitamin D3 and continuted Pb +
exposure, resulted in a significant (p < 0.05 or less) reduction of osteocalcin secretion,
both in the presence and absence of 1 ,25-dihydroxy vitamin D3 at all Pb2+
concentrations. This effect is not mediated by PKC.
1 and 5 |iM Pb2+ significantly increased [Ca2+]; in the absence of 1,25- Not applicable
dihydroxyvitamin D3 and significantly reduced the peak elevation in [Ca2+]j induced
by 1,25-dihydroxy vitamin D3.
Simultaneous treatment of previously unexposed cells to Pb2+and 1,25-
dihydroxy vitamin D3 produced little reduction in the 1,25-dihydroxy vitamin D3-
induced 45Ca uptake, while 40 min of treatment with Pb2+ before addition of 1,25-
dihydroxy vitamin D3 significantly reduced the 1,25-dihydroxy vitamin D3-induced
increase in 45Ca influx.
Osteocalcin secretion significantly reduced below control values by culture with 1 uM Not applicable
Pb2+ in the presence or absence of added 1 ,25-dihydroxy vitamin D3 or 1 ,25-
dihydroxy vitamin D3 and IGF-I. Inhibition of osteocalcin secretion was almost
complete in either hormone- stimulated or basal cultures with the addition of 100 uM
Pb +. Cellular alkaline phosphatase activity paralleled those of osteocalcin, though
there was no response to IGF-I alone or in combination with 1,25-dihydroxy vitamin
D3. Pb2+at 10"15, 10"12, and 10"9 to 10"7M did not influence DNA contents of cell
cultures, but 1 uM significantly (p < 0.05) inhibited basal cultures and those with IGF-
I + D3. Cell cultures exposed to 1 ,25-dihydroxyvitamin D3 and Pb2+ were inhibited at
10uMPb2+.
Reference
Thaweboon et al.
(2002)
Guity et al. (2002)
Schanne et al.
(1992)
Angle etal. (1990)
-------
Table AX5-8.3 (cont'd). Bone Cell Cultures Utilized to Test Effects of Lead
to
o
o
>
X
H
6
o
o
H
O
O
H
W
O
O
HH
H
W
Compound
Dose/Concentration
Duration Exposure
Route
Lead acetate
2 to 200 MM
72 h
In medium
Unidentified Pb2+
Various incubation
times
Not applicable
Unidentified Pb2+
Various incubation
times
Not applicable
Lead acetate
10 MM
2h
In medium
Lead acetate
5 or 25 MM
Up to 24 h
In medium
Species
Rat
Osteo sarcoma
Cells
(ROS 17/2.8)
Bovine
(Bovine-
derived
osteocalcin)
Bovine
(Bovine-
derived
osteocalcin)
Rat
Osteo sarcoma
Cells
(ROS 17/2.8)
Rat
Osteo sarcoma
Cells
(ROS 17/2.8)
Effects Blood Level
Lead (2 to 200 |iM) had no effect on cell number or DNA and protein synthesis. Not applicable
Alkaline phosphatase activity was significantly reduced (p < 0.001) by lead in a dose-
and time-dependent manner.
Pb Concentration. Alkaline Phosphatase Inhibition
2 MM. 10.0 ±1.1%
20 MM. 22.0 ± 6.4%
200 nM. 57.8 ±8.8%
Reductions in alkaline phosphatase mRNA levels mirrored Pb2+ -induced inhibition of
enzyme activity.
Binding studies of Ca + to osteocalcin suggested a single binding site with a Not applicable
dissociation constant (Kd) of 7 ± 2 MM for Ca-osteocalcin. Competitive displacement
experiments by addition of Pb + indicated the Kd for Pb-osteocalcin is 1 .6 ± 0.42 nM,
approximately 3 orders of magnitude higher.
Circular dichroism indicated Pb2+ binding induced a structural change in osteocalcin Not applicable
similar to that found in Ca2+ binding, but at 2 orders of magnitude lower concentration.
Pb + has 4 orders of magnitude tighter binding to osteocalcin (Kd = 0.085 MM) than
Ca2+ (Kd = 1 .25 mM). Hydroxyapatite binding assays showed similar increased
adsorption of Pb + and Ca +to hydroxyapatite, but Pb + adsorption occurred at a
concentration 2-3 orders lower than Ca2+.
Pb2+ treatment reduced the unidirectional rate of ATP synthesis (P; to ATP) by a factor Not applicable
of 6 or more (AM/H,: Control = 0.18 ± 0.04, Pb2+< 0.03). Intracellular free Mg2+
concentration decreased 21% after 2 h of 10 MM Pb + treatment (0.29 ± 0.02 mM prior
to Pb2+ treatment and 0.23 ± 0.02 mM after 2 h of Pb2+ treatment, p < 0.05).
5 MM Pb2+ significantly altered effect of EOF on intracellular calcium metabolism. In Not applicable
cells treated with 5 MM Pb + and 50 ng/mL EGF, there was a 50% increase in total cell
calcium over cells treated with 50 ng/mL EGF alone.
Reference
Klein and Wiren
(1993)
Dowdetal. (1994)
Dowdetal. (2001)
Dowdetal. (1990)
Long and Rosen
(1992)
-------
Table AX5-8.3 (cont'd). Bone Cell Cultures Utilized to Test Effects of Lead
to
o
o
>
X
H
6
o
o
H
O
O
H
W
O
O
HH
H
W
Compound
Dose/Concentration
Duration Exposure
Route
Lead acetate
5 or 25 uM
20 h
In medium
Lead acetate
10-utolO-7M
3 min
In medium
Lead acetate
0.5 to 60 uM
24 to 48 h
In medium
Species
Rat
Osteo sarcoma
Cells
(ROS 17/2.8)
Rat
Osteo sarcoma
Cells
(ROS 17/2.8)
Human
Osteo sarcoma
Cells (HOS
TE85)
and
Rat
Osteo sarcoma
Cells
(ROS 17/2.8)
Effects Blood Level
Treatment with 400 ng/mL culture medium for 1 h or with 25 uM Pb2+ for 20 h Not applicable
increased total cell calcium:
Treatment Cell Calcium
Control 7.56 ± 1 .05 nmol/mg protein
PTH (400 ng/mL) 23.28 ± 1 .40* nmol/mg protein
Pb (25 uM) 1 1 .37 ± 0.57* nmol/mg protein
PTH + Pb 37.88 ± 4.21 * nmol/mg protein
* p < 0.05 from control
Treatment of ROS cells with Pb at 1 or 5 uM concentrations produced a rise in [Ca2+]; Not applicable
to 170 nM and 230 nM, respectively, over the basal level of 125 nM. An elevation in
[Ca2+]i to 210 nM occurred during treatment with an activator of PKC, phorbol 12-
myristate 13-acetate (10 uM). Pretreatment with a selective inhibitor of PKC,
calphostin C, did not change basal [Ca2+]j, but prevented the Pb-induced rise in [Ca2+];.
Free Pb2+activated PKC in a range from 10"11 to 10"7 M, with a Kcat (activation
constant) of 1 . 1 X 10"10M and a maximum velocity (Vmax) of 1.08 nmol/mg/min
compared with Ca activation of PKC over a range of 10"8 to 10"3 M, with a Kcat of
3.6 X 10" M, and a Vmax of 1.12 nmol/mg/min.
HOS TE 85 Cells Not applicable
Inhibition of proliferation (IC50) = 4 uM lead
Cytotoxicity = 20 uM lead
ROS 17/2.8 Cells
Inhibition of proliferation (IC50) = 6 uM lead
Cytotoxicity = 20 uM lead
Highest lead concentration in both cell types found in mitochondrial fraction.
Reference
Long etal. (1992)
Schanne et al.
(1997)
Angle etal. (1993)
-------
Table AX5-8.3 (cont'd). Bone Cell Cultures Utilized to Test Effects of Lead
to
o
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Compound
Dose/Concentration
Duration Exposure
Route
Species
Effects
Blood Level
Reference
>
X
Lead acetate or lead
chloride
O.lto200 uM
24 h to 6 d
In medium
Lead acetate
0.1 to 30 uM
24 h
In medium
Chick growth Growth plate chondrocytes were exposed to 3 or 30 uM for up to 6 days. Maximal Not applicable
plate inhibition of cell proliferation as measured by thymidine incorporation occurred after a
chondrocytes 3-day exposure to lead. A similar 40% inhibition was found at both concentrations.
Higher concentrations (up to 100 uM) did not produce further inhibition.
In cultures treated for 24 h, lead produced a dose-dependent inhibition of alkaline
phosphatase, with 10 uM producing maximal inhibition (40-50% inhibition). Effects
of lead on proteoglycan synthesis were not found until after 48 h of exposure, with
maximal effect after 72 h of exposure (twofold, 30 uM). Lead exposure (10 to
200 uM) for 24 h produced a dose-dependent inhibition of both type II and type X
collagen synthesis.
Chicken A dose-dependent inhibition of thymidine incorporation into growth plate Not applicable
growth plate chondrocytes was found with exposure to 1-30 uM lead for 24 h. A maximal 60%
and sternal reduction occurred at 30 uM. Lead blunted the stimulatory effects on thymidine
chondrocytes incorporation produced by TGF-pl (24% reduction) and PTHrP (19% reduction),
however, this effect was less than with lead alone. Lead (1 and 10 uM) increased type
X collagen in growth plate chondrocytes approximately 5.0-fold and 6.0-fold in TGF-
pl treated cultures and 4.2-fold and 5.1-fold in PTHrP treated cultures when compared
with controls, respectively. Lead exposure alone reduced type X collagen expression
by 70-80%.
Hicks etal. (1996)
Zuscik et al.
(2002)
H
6
o
o
H
O
O
H
W
O
O
HH
H
W
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Table AX5-8.3 (cont'd). Bone Cell Cultures Utilized to Test Effects of Lead
to
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o
Compound
Dose/Concentration
Duration Exposure
Route
Species
Effects
Blood Level
Reference
Abbreviations
>
X
Oi
H
6
o
o
H
O
O
H
W
O
O
HH
H
W
Pb - lead
uCi - microCurie
IU - international units
hr - hour
OB - osteoblast
5F-BAPTA - l,2-bis(2-amino-5-£luorophenoxy)ethane-N,N,N',N'-tetraacetic acid
[Pb2+]i-free intracellular lead
FLTRA-2 - l-[6-Amino-2-(5-carboxy-2-oxazolyl)-5-benzofuranyloxy]-2-(2-amino-5-methylphenoxy)
ethane-N,N,N',N'-tetraacetic acid
M - molar
DNA - deoxyribonucleic acid
AM - decrease in magnetization of intracellular P; upon prolonged saturation of gamma-phosphate of ATP
mM - millimolar
Kcat - activation constant
IC50 - inhibitory concentration 50%
mg - milligram
210Pb - lead-210 radionuclide
EM - experimental medium
mU - milliunits
45Ca - calcium-45 radionuclide
CRAC - calcium release activated calcium reflux
pg - picogram
mRNA - messenger ribonucleic acid
ng - nanogram
Vmax - maximum velocity
TGF-p1! - transforming growth factor-beta 1
mL - milliliter
IV - intravenous
CM - control medium
uM - micromolar
ng - nanogram
[Ca2+]i -free intracellular calcium
PGE2 - prostaglandin E2
PKC - protein kinase C
Kd - dissociation constant
nmol - nanomole
HOS TE 85 cells - human osteosarcoma cells
PTHrP - parathyroid hormone-related protein
GD - gestational day
PTH - parathyroid hormone
min - minute
OC - osteoclast
ROS 17/2.8 -rat osteosarcoma cells
nM - nanomolar
h - hour
IGF-I- insulin growth factor -1
ATP - adenosine triphosphate
EGF - epidermal growth factor
-------
Table AX5-8.4. Bone Lead as a Potential Source of Toxicity in Altered Metabolic Conditions
to
o
o
Compound
Dose/Concentration
Duration Exposure
Route
Species
Effects
Blood Level
Reference
>
X
H
6
o
o
H
O
O
H
W
O
O
HH
H
W
Lead acetate
200 ug/mL
105 days prior to mating
or 105 days prior to
mating and during
gestation and lactation
(160 days)
In drinking water
Lead acetate
12 mM
8 weeks prior to mating
and during gestation
In drinking water
Lead acetate
100 ppm
(A) Exposure for 158 ±
2 days from 21 days of
age to midlactation; (B)
Exposure 144 ± 2 days
from day 21 up to
delivery; (C) Exposure
only during lactation;
(D, E, and F) groups of
non-pregnant rats
exposed for periods
equivalent to groups A,
B and C, respectively.
In drinking water
Mice Results suggested very little lead was transferred from mother to fetus during
gestation, however, lead transferred in milk and retained by the pups
accounted for 3% of the maternal body burden of those mice exposed to lead
prior to mating only. The amount of lead retained in these pups exceeded that
retained in the mothers, suggesting lactation effectively transfers lead burden
from mother to suckling offspring. Transfer of lead from mothers was
significantly higher when lead was supplied continuously in drinking water,
rather than terminated prior to mating.
Not given
Rat
Rat
Considerably higher lactational transfer of lead from rat dams compared to
placental transfer was reported. Continuous exposure of rat dams to lead until
day 15 of lactation resulted in milk lead levels 2.5 times higher than in whole
blood, while termination of maternal lead exposure at parturition yielded
equivalent blood and milk levels of lead, principally from lead mobilized from
maternal bone.
In rats exposed to lead 144 days prior to lactation (B), the process of lactation
itself elevated blood lead and decreased bone lead, indicating mobilization of
lead from bone as there was no external source of lead during the lactation
process. Rats exposed to lead for 158 days (A)(144 days prior to lactation and
14 days during lactation) also experienced elevated blood lead levels and loss
of lead from bone. Lead exposure only during the 14 days of lactation was
found to significantly increase intestinal absorption and deposition (17 fold
increase) of lead into bone compared to non-pregnant rats, suggesting
enhanced absorption of lead takes place during lactation. The highest
concentration of lead in bone was found in non-pregnant, non-lactating control
animals, with significantly decreased bone lead in lactating rats secondary to
bone mobilization and transfer via milk to suckling offspring.
Concentration (ug/1) in
whole blood at day 15 of
lactation:
Controls = 14 ± 4; Lead-
exposed until parturition =
320 ± 55; Lead-exposed
until day 15 of lactation =
1260 ±171*
*p < 0.001 compared with
dams at parturition.
Concentration (ug/dL) in
whole blood at day 14 of
lactation or equivalent:
Group A =31.2 ±1.1;
Group B = 28.0 ± 1.7;
Group D = 27.3 ±2.2;
Group E = 24.7 ±1.2
Keller and Doherty
(1980a)
Palminger Hallen
etal. (1995)
Maldonado-Vega
etal. (1996)
-------
Table AX5-8.4 (cont'd). Bone Lead as a Potential Source of Toxicity in Altered Metabolic Conditions
to
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o
Compound
Dose/Concentration
Duration Exposure
Route
Species
Effects
Blood Level
Reference
>
X
oo
H
6
o
o
H
O
O
H
W
O
O
HH
H
W
Lead acetate
100 ppm
(A) Exposure for 158 ±
2 days from 21 days
through 14 days of
lactation; (B)
Nonpregnant control
Group A; (C) Exposure
144 ±2 days from day
21 up to delivery; (D)
Nonpregnant control
Group C; (E) Lactating
rats not exposed to Pb;
(F) Nonpregnant rats
not exposed to Pb.
In drinking water
Lead acetate
250 mg/mL
Beginning at 5 weeks of
age, rats exposed to lead
for 5 weeks, followed
by no additional
exposure.
In drinking water
Rat When dietary calcium was reduced from the normal 1% to 0.05%, bone
calcium concentration decreased by 15% and bone lead concentration
decreased by 30% during the first 14 days of lactation. In non-lactating rats on
the 0.05% calcium diet, there were also decreases in bone calcium, but no
incremental bone resorption nor lead efflux from bone, suggesting the efflux
from bone during lactation was related to bone resorption. Enhancement of
calcium (2.5%) in the diet of lactating rats increased calcium concentration in
bone by 21%, but did not decrease bone resorption, resulting in a 28%
decrease in bone lead concentration and concomitant rise in systemic toxicity.
Rat Demonstrated adverse effects in rat offspring bom to females whose exposure
to lead ended well before pregnancy. Five week-old-female rats given lead
acetate in drinking water (250 mg/mL) for five weeks, followed by a one
month period without lead exposure before mating. To test the influence of
dietary calcium on lead absorption and accumulation, some pregnant rats were
fed diets deficient in calcium (0.1%) while others were maintained on a normal
calcium (0.5%) diet. All lead-exposed dams and pups had elevated blood lead
levels, however pups bom to dams fed the diet deficient in calcium during
pregnancy had higher blood and organ lead concentrations compared to pups
from dams fed the normal diet. Pups born to lead-exposed dams had lower
mean birth weights and birth lengths than pups born to non-lead-exposed
control dams (p < 0.0001), even after confounders such as litter size, pup sex,
and dam weight gain were taken into account.
Concentration (ng/dL) in
whole blood at day 14 of
lactation or equivalent:
Group B = 26.1 ±2.1,
Group A =32.2 ±2.7*;
Group D = 23.8 ±2.1,
Group C = 28.2 ± 2.2*;
Groups E and F = 5.1 ±
0.4.
* p < 0.01, compared to
appropriate control
Blood lead concentration
of pups (nM): Low
calcium/no Pb = 0.137 ±
0.030°; Low calcium/Pb =
1.160 ±0.053A; Normal
calcium/No Pb = 0.032 ±
0.003°; Normal
calcium/Pb = 0.771 ±
0.056B.
Values that are not
marked by the same letter
are significantly different
(p<0.05).
Maldonado-Vega
et al. (2002)
Han et al. (2000)
-------
Table AX5-8.4 (cont'd). Bone Lead as a Potential Source of Toxicity in Altered Metabolic Conditions
to
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o
Compound
Dose/Concentration
Duration Exposure
Route
Species
Effects
Blood Level
Reference
>
X
vo
Lead acetate
1500 ug/Common
Pb/kg/day
approximately 10 years,
replaced by a 204Pb-
enriched dose (50 days),
then 206Pb-enriched dose
(50 days), and finally a
207Pb-enriched dose (50
days, with reduced
concentration)
Orally, in gelatin
capsule
Nonhuman Sequential doses of lead mixes enriched in stable isotopes (204Pb, 206Pb,
Primate and 207Pb) were administered to a female cynomolgus monkey (Macaco
fascicularis) that had been chronically administered a common lead isotope
mix. The stable isotope mixes served as a marker of recent, exogenous lead
exposure, while the chronically administered common lead served as a marker
of endogenous (principally bone) lead. From thermal ionization mass
spectrometry analysis of the lead isotopic ratios of blood and bone biopsies
collected at each isotope change, and using end-member unmixing equations,
it was determined that administration of the first isotope label allowed
measurement of the contribution of historic bone stores to blood lead.
Exposure to subsequent isotopic labels allowed measurements of the
contribution from historic bone lead stores and the recently administered
enriched isotopes that incorporated into bone. In general the contribution from
the historic bone lead (common lead) to blood lead level was constant
(approximately 20%), accentuated with spikes in total blood lead due to the
current administration of the stable isotopes. After cessation of each
sequential administration, the concentration of the signature dose rapidly
decreased.
Total blood lead range:
31.2 to 62.3 ug/lOOg.
Inskipetal. (1996)
H
6
o
o
H
O
O
H
W
O
O
HH
H
W
Lead acetate
1300 to 1500
ug/Common Pb/kg/day
approximately 10 years,
replaced by a 204Pb-
enriched dose (47 or
281 days), then 206Pb-
enriched dose (50 or
105 days), and finally a
207Pb-enriched dose
(50 days, with 650 ug
concentration in only
one primate)
Orally, in gelatin
capsule
Nonhuman Initial attempts to apply a single bone physiologically based model of lead
Primate kinetics were unsuccessful until adequate explanation of these rapid drops in
stable isotopes in the blood were incorporated. Revisions were added to
account for rapid turnover of the trabecular bone compartment and slower
turnover rates of cortical bone compartment, an acceptable model evolved.
From this model it was reported that historic bone lead from 11 years of
continuous exposure contributes approximately 17% of the blood lead
concentration at lead concentration over 50 ug/dL, reinforcing the concept that
the length of lead exposure and the rates of past and current lead exposures
help determine the fractional contribution of bone lead to total blood lead
levels. The turnover rate for cortical (approximately 88% of total bone by
volume) bone in the adult cynomolgus monkey was estimated by the model to
be approximately 4.5% per year, while the turnover rate for trabecular bone
was estimated to be 33% per year.
Various
O'Flaherty et al.
(1998)
-------
Table AX5-8.4 (cont'd). Bone Lead as a Potential Source of Toxicity in Altered Metabolic Conditions
to
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Compound
Dose/Concentration
Duration Exposure
Route
Species
Effects
Blood Level
Reference
>
X
Lead acetate
1100 to 1300
ug/Common Pb/kg/day
approximately 14 years,
replaced by a 204Pb-
enriched dose, 206Pb-
enriched dose, and/or
finally a 207Pb-enriched
dose of varied durations
and concentrations.
Orally, in gelatin
capsule
Nonhuman Using the method of sequential stable isotope administration examined flux of
Primate lead from maternal bone during pregnancy of 5 female cynomolgus monkeys.
Blood lead levels in maternal blood attributable to lead from mobilized bone
were reported to drop 29 to 56% below prepregnancy baseline levels during
the first trimester of pregnancy. This was ascribed to the known increase in
maternal fluid volume, specific organ enlargement (e.g. mammary glands,
uterus, placenta), and increased metabolic activity that occurs during
pregnancy. During the second and third trimesters, when there is a rapid
growth in the fetal skeleton and compensatory demand for calcium from the
maternal blood, the lead levels increased up to 44% over pre-pregnancy levels.
With the exception of one monkey, blood lead concentrations in the fetus
corresponded to those found in the mothers, both in total lead concentration
and proportion of lead attributable to each isotopic signature dose (common =
22.1% vs. 23.7%, 204Pb = 6.9% vs. 7.4%, and 206Pb = 71.0% vs. 68.9%,
respectively). Between 7 and 25% of lead found in fetal bone originated from
maternal bone, with the balance derived from oral dosing of the mothers with
isotope during pregnancy. In offspring from a low lead exposure control
monkey (blood lead <5 ug/100 g) approximately 39% of lead found in fetal
bone was of maternal origin, suggesting enhanced transfer and retention of
lead under low lead conditions
Various, with total blood
lead as high as
approximately 65 ug/lOOg
Franklin et al.
(1997)
H
6
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o
H
O
O
H
W
O
O
HH
H
W
Lead acetate
250 mg/1
Exposure began either
at 5, 10, or 15 weeks of
age and continued for a
total of 5 weeks.
Drinking water
Rat Exposed rats for five weeks to 250 mg/1 lead as acetate in drinking water
beginning at 5 weeks of age (young child), 10 weeks of age (mid-adolescence),
or 15 weeks of age (young adult), followed by a 4 week period of without lead
exposure. An additional group of rats were exposed to lead beginning at 5
weeks, but examined following an 8 or 20 week period after cessation of lead.
Significantly lower blood and bone lead concentrations were associated with
greater age at the start of lead exposure and increased interval since the end of
exposure. Young rats beginning exposure to lead at 5 weeks and examined 20
weeks after cessation of exposure still, however, had bone lead concentrations
higher than those found in older rats only 4 weeks after cessation of exposure.
Lead concentration (uM)
4 weeks after cessation of
lead exposure:
Exposure started at 5
weeks of age = 1.39 ±
0.09; Exposure started at
10 weeks of age =1.18 ±
0.12;
Exposure started at 15
weeks of age = 0.82 ±
0.05.
Han etal. (1997)
-------
Table AX5-8.4 (cont'd). Bone Lead as a Potential Source of Toxicity in Altered Metabolic Conditions
to
o
o
>
X
H
6
o
o
H
O
O
H
W
O
O
HH
H
W
Compound
Dose/Concentration
Duration Exposure
Route
Lead acetate
50 ppm
1 1 months
Drinking water
Lead acetate
0, 2, or 10 mg/kg/day
9.5 months
Drinking water
Lead acetate
7 years total
Drinking water
Species Effects
Rat Studied differences in tissue distribution of lead in adult and old rats. Adult
(8 months old) and old (16 months old) rats were exposed to 50 ppm lead
acetate in drinking water for 1 1 months at which time the experiment was
completed. Bone (femur) lead levels in older rats were found to be less than
those in younger rats, however, blood lead levels were higher in the older rats.
Brain lead concentration in the older rats exposed to lead were significantly
higher, and brain weight significantly less than the brain lead concentration
and weights of unexposed older control rats or adult rats exposed to lead,
suggesting a potential detrimental effect. Authors suggested that a possibility
for the observed differences in tissue concentrations of lead was due to
changes in the capacity of bone to store lead with advanced age.
Rat Examined kinetic and biochemical responses of young (21 day old), adult
(8 months old), and old (16 months old) rats exposed to lead at 0, 2, or 10 mg
lead acetate/kg/day over a 9.5 month experimental period. Results suggested
older rats may have increased vulnerability to lead due to increased exposure
of tissues to lead and greater sensitivity of the tissues to the effects of lead.
Nonhuman In studies of bone lead metabolism in a geriatric, female nonhuman primates
primate exposed to lead approximately 10 years previously, there were no significant
changes in bone lead level over a 10 month observation period as measured by
Blood Level
Approximate median
values after 6 months of
exposure: Adult rats : 23
ug/dL
Old rats: 31 ug/dL
After 1 1 months of
exposure:
Adult rats: 16 ug/dL
Old rats: 31 ug/dL
Various from
approximately 1 ug/dL up
to 45 ug/dL
Historic concentrations
during exposure: 44 to 89
ug/lOOmL.
Reference
Cory-Slechta et al.
(1989)
Cory-Slechta
(1990)
McNeill et al.
(1997)
Lead (type unidentified)
occurring naturally in
diet (0.258 ng/mg dry
wt) and water (5.45
ppb).
Exposure from age 1
month up to 958 days.
Drinking water and diet
Mice
109CD K X-ray fluorescence. The mean half-life of lead in bone of these
animals was found to be 3.0 ± 1.0 years, consistent with data found in humans,
while the endogenous exposure level due to mobilized lead was 0.09 ± 0.02
ug/dL blood.
The lead content of femurs increased by 83% (values ranged from 0.192 to
1.78 ng Pb/mg dry wt), no significant relationship was found between lead and
bone density, bone collagen, or loss of calcium from bone. The results suggest
against low levels of bone lead contributing to the osteopenia observed
normally in C57BL/6J mice.
None given
Massie and Aiello
(1992)
-------
Table AX5-8.4. (cont'd). Bone Lead as a Potential Source of Toxicity in Altered Metabolic Conditions
to
o
o
Compound
Dose/Concentration
Duration Exposure
Route
Species
Effects
Blood Level
Reference
>
X
to
Lead acetate
250 mg/1
Exposure for 5 weeks
Drinking water
Rat Rats were exposed to lead for 5 weeks, followed by a 4 week washout period
without lead to allow primarily accumulation in the skeleton. Rats were then
randomly assigned to a weight maintenance group (WM), a moderate weight
loss (MWL) group (70% of maintenance diet), or a substantial weight loss
(SWL) group (40% of maintenance diet) for a four week period. At the end of
this experimental period the blood and bone levels of lead did not differ
between groups, however, the amount and concentration of lead in the liver
increased significantly.
Treatment Group Lead (nmol/g)
Femur WM 826 ± 70
MWL 735 ±53
SWL 935 ± 84
Spinal Column Bone
WM
MWL
SWL
702 ± 67
643 ± 59
796 ± 59
WM=1.25±0.10uM;
MWL = 1.16±0.10uM;
WM=1.32±0.10uM;
Han etal. (1996)
H
6
o
o
H
O
O
H
W
O
O
HH
H
W
Lead acetate
250 ng/1
14 days
Drinking water
Rat Study was undertaken to determine the effect of weight loss and exercise on
the distribution of lead. Weight loss secondary to dietary restriction was a
critical factor elevating organ lead levels and, contrary to prior study (Han
et al. (1996)), elevated blood levels of lead. No significant difference in organ
or blood lead concentrations were reported between the exercise vs. no
exercise groups.
Graphs indicate
concentrations ranging
from 0.20 to 2.00 uM.
Han etal. (1999)
Abbreviation
ug - microgram
mL - milliliter
% - percent
mM - millimolar
1 - liter
ppm - parts per million
dL - deciliter
mg - milligram
uM - micromolar
Pb - lead
kg - kilogram
g - gram
204r>K 206
'Pb, 206Pb, 207Pb - Stable isotopes of lead 204, 206, 207, respectively
wt - weight
ppb - parts per billion
-------
Table AX5-8.5. Uptake of Lead by Teeth
to
o
o
>
X
Compound
Dose/Concentration
Duration Exposure
Route
Species
Effects
Blood Level
Reference
Lead acetate
1 Hg/kg body weight
Single IP injection
Rat Uptake of lead label into incisors of suckling rats:
0.7% of injected dose in 4 incisors of suckling rat after 24 h,
1 .43% after 1 92h. 0.6% of injected dose in 4 incisors of adult after 24 h,
0.88% after 192h.
Mean percent of dose after
time:
Suckling:
3.04%after24h
Momcilovic and
Kostial(1974)
1.71% after 72h
1.52% after 144h
1.18%afterl92h
Adult:
6.40%after24h
3.41% after 24h
1.92% after 24h
1.04% after 72h
0.72% after 144h
0.48% after 192h
H
6
o
o
H
O
O
H
W
O
O
HH
H
W
Lead aerosol
77,249,orl546ng/m3
for 50 to70 days
Inhalation
Lead acetate
0, 3, orlOppm
During pregnancy and
21 days of lactation
Drinking water
1 1 micrograms Pb/g incisor taken up in animals exposed to 77 ng/m3 for 70
days versus 0.8|ig Pb/g in control animals
13.8 ng Pb/g incisor in rats exposed to 249 ng/m3 for 50 days
g Pb/g incisor in rats exposed to 1546 ng/m for 50 days
Rat
Rat Lead concentration in teeth of offspring:
0 ppm group - Incisors (1 .3 ppm), 1st molars (0.3 ppm)
3 ppm group - Incisors (1 .4 ppm), 1st molars (2.7 ppm)
10 ppm group - Incisors (13.3 ppm), 1st molars (1 1.4 ppm)
Control: 2.6 ng/dL
77|ig/m3: ll.Sjjg/dL
249 ng/m3: 24.1 ng/dL
1546ng/m3:61.2|ig/dL
Not given
Grobleretal. (1991)
Grobleretal. (1985)
Abbreviations
Hg - microgram
kg - kilogram
IP - intraperitoneal
% - percent
h - hour
m - cubic meter
Pb - lead
g - gram
ppm - parts per million
-------
Table AX5-8.6. Effects of Lead on Enamel and Dentin Formation
to
o
o
Compound
Dose/Concentration
Duration Exposure
Route
Species
Effects
Blood Level
Reference
>
X
H
6
o
o
H
O
O
H
W
O
O
HH
H
W
Pb "salt" Rat
0.075 mM/lOOg, 0.15
mM/100gorl.5
mM/lOOg
Single, SC injection
Pb acetate Rat
30 mg/kg
Single, IV injection
Pb acetate Rat
3 mg/kg
Single, IV injection
Pb acetate Rat
Omg/1, 34mg/l, or 170
mg/1
70 days
Drinking water
Pb acetate Rat
40 mg/kg
Single, IP injection
0.075 mM dose, no disruption of dentin and enamel.
0.15 mM dose, mild mineralization disruption of dentin and enamel.
1.5 mM dose, mild to moderate disruption of dentin and enamel.
Rapid rise in serum calcium and phosphorus after injection. Formation of a
"lead line" in growing dentin within 6 hours after injection.
Production of a hypomineralized band in dentin
Increased in relative amount of protein in enamel matrix.
Significant (p<0.05) decrease in microhardness values of groups exposed to
lead in regions of maturation enamel, but not fully mature enamel.
Delay in enamel mineralization in animals exposed to lead.
Significantly (p<0.05) reduced eruption rates at various time points (days 8,
14,16, 22, 24, 28) under hypo functional conditions.
Not given
Not given
Not given
0 mg/1 group: 0 ppm
34 mg/1 group: 18.1 ppm
170 mg/1 group: 113.3
ppm
Days after injection
0 d: 48 ug/dL
10 d: 37 ug/dL
20 d: 28 ug/dL
30 d: 16 ug/dL
(Values estimated from
graph)
Eisenmann and Yaeger
(1969)
Appleton(1991)
Appleton(1992)
Gerlach et al. (2002)
Gerlach et al. (2000b)
Abbreviations
Pb - lead
mM - millimolar
g-gram
SC - subcutaneous
mg - milligram
kg - kilogram
IV - intravenous
1 - liter
ppm - parts per million
IP - intraperitoneal
ug - microgram
dL - deciliter
-------
Table AX5-8.7. Effects of Lead on Dental Pulp Cells
to
o
o
Compound
Dose/Concentration
Duration Exposure
Route
Species
Effects
Blood Level
Reference
Pb glutamate
4.5xlO'5M-10'7M
1,3, or 5 days
incubation
Human All concentrations significantly increased cell proliferation on Day 1, 3 and
Dental 5 of exposure in serum free conditions. Lead exposure resulted in dose-
Pulp Cells dependent decrease in intracellular protein and procollagen I production
over 5 days. In presence of serum only 4.5 x 10"5M significantly increased
protein production. Lead significantly decreased osteocalcin production.
Not applicable
Thaweboon et al. (2002)
>
X
Abbreviations
Pb - lead
M - molar
H
6
o
o
H
O
O
H
W
O
O
HH
H
W
-------
Table AX5-8.8. Effects of Lead on Teeth - Dental Caries
^C Compound
to Dose/Concentration
O Duration Exposure
Route
Lead acetate
0.5 mEq
84 d males
98 d females
Drinking water
Lead acetate
34 ppm
Pre- and perinatal
Drinking water
Lead acetate
10 or 25 ppm lead
3 weeks
Drinking water
Species Effects
Hamster Significant increase in dental caries in male hamsters only (85
mean molar caries score control vs. 118 for lead exposed).
No significant difference in dental caries in female hamsters (68
mean molar caries score control vs. 85 for lead exposed).
Rat Lead exposure resulted in an almost 40% increase in the
prevalence of caries and nearly 30% decrease in stimulated
parotid salivary gland function.
Rat When 1 5 ppm fluoride was concurrently given in diet, lead did
not increase prevalence of caries.
Blood Level
Not given
Control: < 5 ng/dL
34 ppm Pb: 48 ±13
Hg/dL
Not given
Reference
Wisotzky and Hein (1958)
Watson etal. (1997)
Tabchouryetal. (1999)
>
X
^ Abbreviations
mEq - milliequivalents
d - days
ppm - parts per million
O % - percent
^ Hg - microgram
(^ dL - deciliter
H
O
O
2
O
H
O
c
o
H
W
O
V
O
HH
H
W
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ANNEX TABLES AX5-9
May 2006 AX5-5-147 DO-NOT QUOTE OR CITE
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Table AX5-9.1. Studies on Lead Exposure and Immune Effects in Humans
to
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[>
H
6
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o
H
O
o
H
W
O
O
1 — I
H
W
Nature of
Exposure
Environmental
Occupational
Environmental
Environmental
Occupational
Environmental
Occupational
Occupational
Dose or Blood Lead Levels
(BLLs)
10.1-48.2 ug/L (BLL)
22 ug/dL: <30 years old
23.0 ug/dL: 30-39 years old
24. 1 ug/dL: > 40 years old
3.47 -49. 19 ug/dL
2.56-43.69 ug/dL (mean
9.52 ug/dL)
Range of 10.0-400.9 ug/dL
Mean=88.3 ug/dL
Controls all below 10 ug/dL
2.56 - 43.69 ug/dL
(BLL mean of 9.52 ug/dL)
10-20 year exposure (original
BLL mean 60 ug/dL; at time of
study BLL mean = 30 ug/dL)
74.8 ± 17.8 ug/dL vs. 16.7 ±
5.0 ug/dL for controls
Sample Population
2ntl grade children living near
industrial waste incinerator or other
industries causing pollution
Employees of lead storage battery
factories in Korea
554 Men
52 Women
Children 6- 1 1 years of age
30 girls
35 boys
Proximity to smelter
38 preschool children (3-6 years of
age);
35 controls
Male lead-exposed workers
96 females
121 males
(3-6 years old)
30 lead workers from battery
manufacturing plant (43 males and
21 females)
25 male storage battery workers
exposed >6 months; age 33 ± 8.5
years
Reported Effects
Increased blood lead concentration associated w/ increased IgE, especially above
28 ug/dL
Also decreased T-cells, cytotoxic T-cells, and B-cells (non-linear relation)
Serum IgE higher when BLL >30 ug/dL - Correlation of BLL with serum IgE
For employees less than 30 years old, IL-4 was lower when BLL >30 ug/dL
Indirect (PHA) macrophage activation
NO production negatively associated with BLL
With proximity closest to smelter monocytes had increased superoxide anion
production by indirect and direct activation (positive correlation with BLL -
stronger for boys than girls)
Percent of CD4+ and CD4+CD+ cells decreased while CD8+ increased
PHA-mitogen response decreased; and IFN-gamma production increased.
No effect on NK cytotox.
IgG and IgM lower in high BLL group (>9.52 ug/dL)
IgE greater in high BLL group (P < 0. 10)
No difference among males but females exposed to higher lead had significant
decreases in IgG and IgM and increased in IgE
Correlation of BLL and serum IgE r = 0.48; P = 0.002
Increased percentage of monocytes while percentage of B-cells, numbers of
lymphocytes, monocytes, and granulocytes decreased
Decreased blood hemoglobin, TCD4+ cells, IgG, IgM, C3 and C4 compliment
proteins.
Increased zinc protoporphyrin.
Impaired neutrophil chemotaxis and random migration
Reference
Karmaus et al.
(2005)
Heo et al.
(2004)
Pineda-
Zavaleta et al.
(2004)
Zhao et al.
(2004)
Mishra et al.
(2003)
Sun et al.
(2003)
Sune et al
(2003)
Basaran and
Undeger
(2000)
-------
Table AX5-9.1 (cont'd). Studies on Lead Exposure and Immune Effects in Humans
to
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o
Oi
J>
X
1
^
VO
O
>
\T\
H
6
o
2!
-^— i
0
H
/O
r*^
O
w
o
73
o
HH
— ]
w
Nature of
Exposure
Environmental
Epidemiological
study
Occupational
Occupational
Occupational
Occupational
Occupational
Occupational
Occupational
In Vitro
Dose or Blood Lead Levels
(BLLs)
1.7-16.1 ug/dL
(Range in <3 yr old group)
Blood leads from 1-45 ug/dL
BLL=39
Range 15-55 ug/dL
Lead workers with BLL
between 7-50 ug/dL; mean
19 ug/dL
Exposed— Range of 38-100
ug/dL mean = 74. 8 ug/dL;
Controls 1 1-30 ug/dL mean =
16.7 ug/dL (high controls!)
BLL 12-80.0 ug/dL
Males
high BLL > 25 ug/dL
lower BLL <25 ug/dL
control BLL < 10 ug/dL
>60 ug/dL for group showing
best IgE effect
BLL 14.8-91.4 ug/dL
207-1035 ug/L
Sample Population
1561 children and adults in high
lead community
480 controls
Urban Children population in
Missouri; 56% male
279 children 9 months-6 yr. of age
145 lead exposed workers
84 controls
71 male chemical plant workers vs.
29 controls
25 Male battery plant workers vs.
25 controls
33 male workers in a storage
battery plant
5 1 Firearms instructors (high and
lower) vs. controls
2 groups of male workers
occupationally exposed
39 male workers of storage battery
plant
(4 year mean exposure)
Human lymphocytes from adults
25-44 years of age
Reported Effects
6-35 months:
increased IgA, IgG, IgM, number and proportion of B lymphocytes
decreased proportion of T-lymphocytes especially true when BLL > 1 5 ug/dL
>3 years of age - no differences
Correlation of blood lead levels and serum IgE levels in Missouri children
No major effects; only subtle effects
Elev. B cells elevated CD4+/CD45RA+ cells
Deer. Serum IgG
T cell populations,
Naive T cells correlated positively with PBB levels. Memory T cells reduced
with lead.
Absolute and relative numbers of CD4+ T cells reduced in exposed group.
IgG, IgM C3 and C4 serum levels all lower in workers.
No changes in serum Igs of PHA response of PBMC
T cell phenotypes and response — lead reduced relative CD3+ cells and relative
and absolute CD4+ cells also reduced PHA
(high lead)and PWM mitogen responses, reduced MLR also(high lead)
IgE positively correlated with BLL
Impaired neutrophil migration
Impaired nitroblue tetrazolium positive neutrophils
Greater for those exposed up to 1 year than those with longer exposure "safe"
levels of lead can still cause immunosuppression
Lead associated with greater IgG production after stimulation with PWM - not
dose dependent
Reference
Sarasua et al.
(2000)
Lutz, et al.
(1999)
Pinkerton
etal. (1998)
Sata et al.
(1998)
Undeger et al.
(1996)
Queiroz et al.
(1994)
Fischbein
etal. (1993)
Horiguchi
etal. (1992)
Queiroz et al.
(1993)
Borella and
Giardino
(1991)
-------
Table AX5-9.1 (cont'd). Studies on Lead Exposure and Immune Effects in Humans
to
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i
01
O
O
>
H
6
o
0
H
O
o
H
W
O
O
HH
H
W
Nature of
Exposure
Occupational
Occupational
Occupational
Occupational
Environmental
Occupational
Environmental
Dose or Blood Lead
Levels (BLLs) Sample Population
33.2 ug/dL in lead-exposed 10 Male workers in scrap metal
group refinery vs. 10 controls
2.7 ug/dL in controls
10 lead exposure workers vs.
controls
worker BLLs of 41-50 ug/dL
no controls >19 ug/dL
Blood leads 64 ug/dL 39 male workers in lead exposed
Range 2 1-90 group
Comparison of workers with Workers exposure to lead
25-53 ug/dL vs. controls with
8-17ug/dL
Near smelter BLLs varied Boys and girls —11.5 years old
seasonally 25-45 ug/dL living near lead smelting plant
Control area BLLs varied
seasonally 10-22 ug/dL
Workers (18-85.85.2 ug/dL 73 workers vs. 53 controls
BLL)
controls (6.6-20.8 ug/dL BLL)
12 Afr.- American children 12 African American preschool
BLLs 41-51|ig/dL; children vs. 7 controls
7 controls BLLs 14-30 ug/dL
Reported Effects
PMN chemotaxis reduced to 2 different chemoattractants
ConA-generated suppressor cell production — increased,
Some other cellular parameters unchanged
PHA response of lymphocytes from workers decreased
No change in serum Ig levels PHA response of cells or NK activity
Higher BLL associated with: decreased A-amino levulinic acid dehydrogetase
Decreased IgM and secretory IgA
Inversely related to IgG
Negative correlation of BLL and serum IgG and C3.
Positive correlation of BLL and saliva IgA
No difference in anti-tetanus antibody levels or in complement levels
Reference
Valentino
etal. (1991)
Cohen et al.
(1989)
Alomran and
Shleamoon
(1988)
Kimber et al.
(1986b)
Wagnerova
et al. (1986)
Ewers et al.
(1982)
Reigart and
Graber(1976)
-------
Table AX5-9.2. Effect of Lead on Antibody Forming Cells (AFC) (In Vitro Stimulation)
to
o
Oi
X
i
^
O
H
6
o
0
H
O
O
H
W
O
O
HH
H
W
Species Strain/Gender Age
Mouse Various Adult
Mouse Various Adult
Mouse CBA/J females Adult
Mouse BDFi females Adult
Mouse CBA/J females Adult
Mouse CBA/J Adult
Mouse CBA/J females Adult
Mouse Swiss males Adult
Mouse Swiss Adult
Rat SD Neonate-
Juvenile
Mouse Swiss Adult
Mouse Swiss Adult
In Vivo/ Ex Lead Dose/
Effect Vivo Concentration
|AFC No 10 uM
AFC No change Yes 10 mM in water
| AFC primary response No 100 uM
|AFC - T dependent antigen 50 ug lead acetate in water
AFC - T independent antigen, no
change
|AFC Yes 0.08 mM and
0.4 mM
|AFC No 10'5 M
|AFC No 10"4 M
4 AFC Yes 0.5 ppm tetraethyl lead
|AFC Yes 1300 ppm
|AFC (IgM) Yes 25 ppm and 50 ppm
|AFC - IgM Yes 4 mg i.p. or oral
|AFC - IgG
|AFC-IgM Yes 13.75 ppm- 1,375 ppm
|AFC - IgG
Duration
of Exposure
5 days
8 weeks
5 days
3 weeks
4 weeks
5 days
1 hr preincubation
3 weeks
10 weeks
3 weeks prenatal
and 6 weeks
postnatal
Single dose
8 weeks
Reference
McCabe and
Lawrence (1991)
Mudziuski et al.
(1986)
Warner and
Lawrence (1986)
Blakley and
Archer (1981)
Lawrence (1981a)
Lawrence (1981b)
Lawrence (1981c)
Blakley et al.
(1980)
Koller and Roan
(1980)
Luster et al ( 1978)
Koller et al.
(1976)
Koller and
Kovacic (1974)
-------
Table AX5-9.3. Studies Reporting Lead-Induced Suppression of Delayed Type Hypersensitivity and Related Responses
to
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Oi
>
X
1
01
to
o
Prj
H
6
o
2|
0
H
O
O
H
W
O
O
HH
H
W
Species
Rat
Chicken
Rat
Rat
Rat
Chicken
Mouse
Rat
Rat
Goat
Rat
Mouse
Rat
Mouse
Age
Embryo
Embryo
Fetal
Embryo - fetal
Embryo - fetal
Embryo
Adult
Embryo- fetal
Embryo- fetal
Adult
Adult
Adult
Neonatal/ Juvenile
Adult
Strain/Gender Route
SD females Oral to Dam
Cornell K females in ovo
CD females Oral to Dam
F344 and CD females Oral to Dam
F344 females Oral to Dam
Cornell K females in ovo
BALB/c females Oral
F344 females Oral to Dam
F344 females Oral to Dam
Females Gastric intubation
Wistar males Oral
Swiss s.c.
CD females Oral
BALB/c i.p.
Lowest Effective Dose
250 ppm (BLL at 4 wk = 6.75 ug/dL)
200 ug
500 ppm
250 ppm
250 ppm (BLL = 34.8ug/dL at birth)
200 ug(BLL= 11 ug/dL)
512 ppm (BLL = 87 ug/dL)
250 ppm lead acetate
250 ppm lead acetate
50 mg/kg lead acetate
6.3 mmol kg "'
0.5 mg/kg/day
25 ppm lead acetate (BLL= 29.3 ug/dL)
0.025 mg lead acetate
Duration
of Exposure
5 weeks
Single injection E12
6 days
3 weeks
3 weeks
Single injection E12
3 weeks
5 weeks (2 before, 3
during gestation)
5 weeks (2 before, 3
during gestation)
6 weeks
8 weeks
Shortest = 3 days
just prior to
challenge
6 weeks
30 days
Reference
Chen et al. (2004)
Lee et al. (2002)
Bunn et al.
(200 Ic)
Bunn et al.
(200 Ib)
Bunn et al.
(200 la)
Lee etal. (2001)
McCabe et al.
(1999)
Chen et al. (1999)
Miller et al.
(1998)
Haneef et al.
(1995)
Kumar et al.
(1994)
Laschi-Loquerie
et al. (1984)
Faith etal. (1979)
Muller et al.
(1977)
-------
to
o
o
Species
Table AX5-9.4. Effect of Lead on Allogeneic and Syngeneic Mixed Lymphocyte Responses (MLR)
Strain/Gender
Age
Proliferation
Effects
In Vivo/
Ex Vivo
Lead Dose/Concentration
Duration
of Exposure
References
>
X
Mouse
Rat
Mouse
Mouse
Mouse
C57B1/6 and
BALB/c
Lewis males
CBA/J females
CBA/J females
DBA/2J males
Adult fAllo-MLR No
Adult fAllo-MLR No
fSyngeneic-MLR
Adult fAllo-MLR No
Adult fAllo-MLR Yes
Adult Allo-MLR no Yes
significant change
0.1 nM
50 ppm lead acetate
10'6-10-4M
0.08 mM and 0.4 mM
13, 130 and 1300 ppm
4 days McCabe et al. (2001)
4 days Razani-Boroujerdi et al.
(1999)
5 days Lawrence (1981b)
4 weeks Lawrence (1981a)
10 weeks Roller and Roan (1980a)
H
6
o
o
H
O
O
H
W
O
O
HH
H
W
-------
Table AX5-9.5. Effect of Lead on Mitogen-Induced Lymphoid Proliferation
to
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Oi
^
X
1
£
o
H
6
o
0
H
O
O
H
W
O
O
HH
H
W
In Vivo/
Species Strain/ Gender Age Proliferation Effects Ex Vivo Lead Dose/ Concentration
Human - Adult |PHA Yes Not available
Mouse TO males Adult |ConA Yes 1 mg/kg daily
|LPS
Mouse Several Adult PHA stimulation No No 25 uM
change
Rat Lewis and F344 Adult |ConA No 25 ppm
males TLps
Mouse CBA/J Adult LPS No change No 10 uM
Rat AP strain males Adult PHA No change Yes 100 ppm and 1,000 ppm
Mouse CBA/J females Adult ConA No 10'4 M
LPS
No change
Mouse BDF1 females Adult |ConA Yes 0-1,000 ppm
TPHA
fStaph A enterotoxin
LPS no change
Rat SD males Adult |ConA Yes 1% lead acetate in diet
|PHA
TLPS
Mouse CBA/J females Adult PHA no change Yes 10 mM
|LPS (high doses only)
Mouse CBA/J females Adult ConA, PHA no change No 10'6 - 10^ M
TLPS
Mouse CBA/J females Adult ConA, PHA no change No 10'6 - 10'4 M
TLPS
Duration
of Exposure
Occupational
2 weeks
3 days
3 days
3 days
2-20 weeks
2 days
3 weeks
2 weeks
4 weeks
2.5 days
2-5 days
References
Mishra et al. (2003)
Fernandez - Carbezudo et al.
(2003)
McCabeetal.,2001
Razamni - Boroujerdi et al.
(1999)
McCabe and Lawrence (1990)
Kimber et al. (1986)a
Warner and Lawrence (1986)
Blakley and Archer (1982)
Bendichetal. (1981)
Lawrence (1981a)
Lawrence (1981)b
Lawrence (1 98 l)c
-------
Table AX5-9.5 (cont'd). Effect of Lead on Mitogen-Induced Lymphoid Proliferation
to
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o
Oi
X
1
DRAFT-DO I
^
0
H
O
O
H
W
O
O
HH
H
W
In Vivo/
Species Strain/ Gender Age Proliferation Effects Ex Vivo Lead Dose/ Concentration
Mouse C57B1/6 males Adult |PHA Yes 1,300 ppm
|ConA
LPS No change
Rat SD females Neonatal- |PHA Yes 25 ppm
Juveniles iConA
Mouse BALB/c Adult |LPS No 10'5 - 10'3 M
Mouse Swiss males Adult |PHA Yes 2,000 ppm
|PWM
Mouse Swiss males Adult |PHA No 0. 1 mM -1.0 mM
fPWM
Mouse CBA/J Adult |LPS Yes 13 ppm
Mouse BALB/c Adult fLPS No 10-5-10-3M
Duration
of Exposure References
8 weeks Neilan et al. (1980)
6 weeks Faith et al. (1979)
3 days Gallagher et al. (1979)
30 days Gaworski and Sharma (1978)
2-3 days Gaworski and Sharma
(1978)
1 8 months Roller et al. ( 1 977)
2-3 days Shenker et al. ( 1 977)
-------
Table AX5.9.6. Pattern of Lead-Induced Macrophage Immunotoxicity
to
o
o
ON
Species
Strain/
Gender Age
Function
In Vivo/ Lowest Effective Duration
Ex Vivo Dose of Exposure
References
Nitric Oxide
X
1
ON
O
H
6
o
0
H
O
O
H
W
O
O
HH
H
W
Human
Rat
Chicken
Mouse
Chicken
Mouse
Mouse
Reactive
Human
Rat
Mouse
Rabbit
Both genders Juvenile
CD males Embryo
Cornell K Embryo
strain females
BALB/c Adult
females
HD-llcell
line
CBA/J Adult
females
CBA/J Adult
females
Oxygen Intermediates
Associated in Juvenile
males
Not indicated NK
BALB/c Adult
females
New Zealand Adult
white males
|NO
|NO
|NO
|NO
|NO
|NO
|NO
tROI
tROI
tROI
tROI
Yes NK
Yes 500 ppm 6 days
Yes 10 \ig One injection
(E5)
No 20 ng/mL 2 hrs
one lower dose tNO
No 4.5 \ig 18 hrs
No 1 .0 \ig 4 days
No 0.625 |iM 4 days
Yes NK NK
No 240 |iM 3 hrs
Yes l.Smg/kgdiet 30 days
Yes 31 jig/m3 inhaled 3 days
Pineda-Zavaleta et al.
(2004)
Bunnetal. (2001c)
Lee etal. (2001)
Krocova et al. (2000)
Chen etal. (1997)
Tian & Lawrence (1996)
Tian & Lawrence (1995)
Pineda-Zavaleta et al.
(2004)
Shabani & Rabbani (2000)
Baykov etal. (1996)
Zelikoff etal. (1993)
-------
ANNEX TABLES AX5-10
May 2006 AX5-157 DO-NOT QUOTE OR CITE
-------
Table AX5-10.1. Hepatic Drug Metabolism
to
o
ON
i
(^
oo
o
!>
-n
H
1
O
o
0
H
O
o
H
W
O
O
HH
H
W
Concentration
Triethyl Pb chloride,
0-3.0mg/kgb. wt.
In vitro, 0.0-3.0 mM
triethyl Pb
5 or 10 umol/lOOgb.
wt. Pb nitrate; i.v.
5, 10, 50 mg Pb
acetate kg"1 b. wt.
100 umol/kg; i.v. Pb
acetate
100 umol/kg
Pb nitrate; i.v.
100 umol/kg; i.v. Pb
acetate
100 umoles /kg; i.v.
Duration Species
2 days In vitro, rat
microsomes
Not specified In vivo, rat
microsomes
36 h Male Fischer 344
rats
Multiple durations Female albino rats
(15 days,
2 and 3 months)
24 h Male Fischer 344
rats
9 h before or 6 h Male Fischer 344
after rats
2-methoxy-4-
aminoazobenzene
(2-Meo-AAB)
24 h Male F 344 rats
Multiple durations Male Wistar rats
(3, 6, 12, 24, and
36 h)
Blood Lead Effects11
— Triethyl Pb increased microsomal N-oxygenation in vivo and decreased
microsomal C oxygenation by in vitro treatment. Either treatment thus gave
rise to an increase in the N-oxygenation/C-oxygenation ratio, which may
lead to tumor potentiation.
— Lead decreases phase I components (liver microsomal cyt.P-450), and
increases Phase II components (GST, DT diaphorase etc). Liver cytosol in
treated animals had a polypeptide that cross-reacted with GSTP.
— Over all induction of cyt-p - 450 and b5 in liver, long-term increase in liver
GST and GSH.
— Decrease in total CYP amount, selective inhibition of CYP1A2 and decrease
in the expression at m-RNA and protein level, induction of placental form of
glutathione s-transferase (GST-P).
— Male fisher rats treated with different metal ions — Pb nitrate, nickel
chloride, cobalt chloride or cadmium chloride exhibited decreased total CYP
amount in liver microsomes. However, only Pb reduced the levels of the
mRNA and protein of CYP 1A2 induced with 2-methoxy-4-
aminoazobenzene (2-Meo-AAB) and decreased the microsomal activity (Per
CYP), Pb also induced placental form of Glutathione, a marker enzyme for
preneoplastic lesion.
— Inhibition of CYP 1 A mRNA(s) by Pb nitrate is by aromatic amines, not by
aryl hydrocarbons.
— Stimulation of TNFa preceding hepatocyte DNA synthesis indicates a role
for it in liver cell proliferation.
Lead nitrate enhances sensitivity to bacterial LPS, in hepatocytes.
Reference
Odenbro and
Arhenius (1984)
Roomietal. (1986)
Nehru and Kaushal
(1992)
Degawa et al.
(1994)
Degawa et al.
(1995)
Degawa et al.
(1996)
Shinozuka et al.
(1994)
-------
Table AX5-10.1 (cont'd). Hepatic Drug Metabolism
to
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o
>
X
H
6
o
o
H
O
O
H
W
O
O
HH
H
W
Concentration Duration Species
Single 0.33 mg/kg-1 Multiple durations Male Wistar rats
Pb nitrate
Lead acetate, 75 mg Multiple analyses C57BL/6 male
ofPb2+/kg, up to 30 h mice
intraperitonial
0-10~6MPb nitrate 3 days Fishhepatoma
cell line PLHC-1
DT Diaphorase 24 h - 120 h Male Wistar rats
activity 0-125 mg/kg
Pb acetate, Pb nitrate
Time course
experiments
100 mg/kg Pb acetate,
i.p.
Cell viability assays 24 h in general and Primary human
0-30 uM, for all other for EROD assays by hepatocytes
As , Pb, Hg, 5 uM, PAHs 24 -72 h
Cd, 1 uM
Blood Lead Effects11
— Lead confers protection against the CCL4 induced hepatotoxicity as evident
by marked reduction in serum Alanine aminotransferase (AST) and
aspartate aminotransferase (AST) and this protection is not associated with
the mitotic response of Pb.
— The decrease in P-450 as a result of Pb poisoning occurs at two levels. (1)
A mechanism unrelated to heme, where Pb interferes with P-450 in 2 ways.
(2) A mechanism dependent on heme, in which Pb inhibits heme synthesis.
— Effect of heavy metals Cu(II), Cd(II), Co(II), Ni(II), Pb(II), and Zn(II), on
Cytochrome induction (CYP1A) induction response and Ethoxy resorufin-o-
deethylase (EROD) activity.
All metals had a more pronounced effect on EROD activity than Cyp 1 A
protein. The rank order of the metal inhibition on EROD is Cd(II) > Ni(II)
> Cu(II) > Co(II) = Zn(II), Pb(II), Cd(II) and (Cu).
May affect Cypl A system of the fish liver at low concentrations through
the direct inhibition of CYP 1A enzyme activity.
— Lead acetate and nitrate induce DT diaphorase activity which is inhibited
significantly by Dil a calcium antagonist, showing that these changes are
mediated by intracellular calcium changes. Lead acetate induces DT
diaphorase activity with out thymus atrophy and hence was suggested to be
a monofunctional inducer as against the Methyl cholanthrene induced DT
diaphorase activity
— The effect of metals on PAH induced CYP1A and 1A2 as probed by Ethoxy
resorufin o- deethylase activity has demonstrated, metals -Arsenic, Pb,
mercury and cadmium decreased CYP1A1/ A2 expression by polycyclic
aromatic hydrocarbons depending on the dose, metal and the PAH. Arsenic
was most effective, followed by Pb, cadmium, and mercury. Cell viability
was decreased by 20-28% by metals.
Reference
Calabrese et al.
(1995)
Jover (1996)
Bruschweiler et al.
(1996)
Arizono et al.
(1996)
Vakharia(2001)
-------
Table AX5-10.1 (cont'd). Hepatic Drug Metabolism
to
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o
>
X
H
6
o
o
H
O
O
H
W
O
O
HH
H
W
Concentration
10-100 uM,
in vitro
5 and 10 umoles/100
g of b.wt, single i.p
100umoles/100gb.
wt. Pb nitrate, single
injection, i.v.
100 mg/kg i.p., single
exposure
100 umol/kg body wt,
i.v
100 umol/kg b. wt,
intra cardiac
10 umol/lOOgb. wt.,
Pb nitrate, i.v. Single
dose
Duration
24 h
—
Animals were
sacrificed at
1, 2, 3, 4, and 15
days
Multiple analyses 0-
96 h
70 h
Multiple time point
analyses starting
6 days to 5 months.
Analyses at multiple
time points 0-10
days
Species
Murine hepatoma
cell line.
Wistar Rat
Wistar rats
Male DDY strain
mice
Male Sprague
Dawley rats
Male and female
Wistar rats
Male Fischer 344
young adult rats.
Blood Lead Effects11
— Effect of heavy metals on Aryl hydrocarbon regulated genes -metals alone
did not induce a significant change in the cyplal activity and protein levels
but increased its m-RNA expression. AHR ligand - mediated induction of
cyplal activity and protein was observed by all the metals. Pre and post
translational modulation in this regulation have been implicated. These
results demonstrate that the heavy metals differentially modulate the
constitutive and the inducible expression of AHR regulated genes.
— Lead nitrate induced the expression of Placental form of Glutathione
transferase along with liver cell proliferation. The biochemical lesions
induced by Pb under these conditions were similar to that of hepatic
nodules.
— Acute Pb treatment results in induced activity of Gamma- glutamyl
transpeptidase, induced GSTP, atypical marker of pre neoplastic lesion in
most hepatocytes. Lead also inhibited liver adenylate cyclase activity 24 h
post exposure.
— Lead decreased Glutathione content and decreased Glutathione s-transferase
activity that is independent of Glutathione levels.
— Acute Pb nitrate treatment caused a significant increase of GST activity in
liver and kidney. While in liver the activity increase is mainly due to
isozyme GST 7-7, in kidney it is through the induction of all the isozymes.
— Intracardiac administration of Pb acetate results in elevation of glutathione
transferase (GST) in Kupffer cells, the early response to GST.Yp was
observed in sinusoidal cells and had a later patchy response in the
expression of GST.Yp Yp in hepatocytes
— Glutathione transferase Pl-1 is induced significantly by a single intravenous
dose of Pb nitrate through increased transcription and modulations at post
transcription and translational levels.
Reference
Korashy and
El-Kadi (2004)
Roomietal. (1987)
Columbano et al.
(1988)
Nakagawa et al.
(1991)
planas-Bhone and
Elizalde (1992)
Boyce and Mantel
(1993)
Kooetal. (1994)
-------
Table AX5-10.1 (cont'd). Hepatic Drug Metabolism
N>
O
ON
i
ON
O
w
j>
H
1
O
o
0
H
O
o
H
W
O
O
HH
H
W
Concentration Duration Species
Lead nitrate, 48 h Transgenic rats
100 urn/kg i.p, with 5 different
3 times every 24 h constructs having
GST-P and/or
Lead acetate 0.5-24 h chloromphenical
lOOuM/kg. acetyltransferase
coding sequence.
10 nM Pb nitrate 24 h before NRK Kidney
transfection with fibroblasts
ECAT deletion
mutant, every 24 h
there after till 48 h
after transfection
10 mg Triethyl Pb, Analyses at multiple Fischer 344 rats
i.p. single dose durations (3, 4, 7,
10, or 14 days)
1 14 mg Pb acetate/kg Single (0.5-12 h Sprague Dawley
b. wt. i.p group) or multiple
(72 h and 7 d group)
exposure
A. 1.5-3.0 mg/kgwt 2 exposures for 48 h Female Wistar
Triethyl Pb (TEL) i.p.
B. 0.05-0.5, TEL to 30 min incubations Liver microsomes
liver microsomal from female
fractions Wistar rats
„, , T , Effects" Reference
Blood Lead
— GSTP (placental GST), is regulated by Pb at transcriptional level. GST-P Suzuki etal. (1996)
enhancer (GPEI), is an essential cis- element required for the activation of
the GST-P gene by Pb and is involved in the activation regardless of the
trans-activators involved. GPEI element consists of two AP-1 binding sites.
Activation of GST-P gene by Pb is mediated in major part by enhancer
GPEI, which may involve AP- 1 activation partially.
— Lead induces GST-P in NRK normal rat kidney fibroblast cell line.
— Decreased liver Glutathione s-transferase (GST) activity and lower levels of Daggett et al.
several hepatic GST (1997)
Increase in quinone reductase activity by day 14 in liver.
— Pb exposure resulted in hepatic Glutathione (GSH) depletion and increased Dagget et al. (1998)
malondialdehyde (MDA) production.
— Pretreatment of rats did not affect the liver microsomal Oestradiol-17p Odenbro and Rafter
metabolism or the content of cytochrome P-450 and cytochrome b5. (1988)
— TEL at 0.05 mM significantly reduced 17p-hydroxy steroid oxidation and at
concentration of 0.05 mM decreased 16a-hydroxylation
-------
Table AX5-10.1 (cont'd). Hepatic Drug Metabolism
to
o
o
>
X
to
Concentration
50 mg/kg, intragastric
b. wt.
CYP
GSH
GSSG
TEL
CCL
GSTP
MDA
ALA
PAH
LPS
Duration Species
8 weeks Male Albino
Wistar rats
body weight
Cytochrome P-450
Glutathione
Oxidized glutathione
Triethyl lead
Carbon tetrachloride
Placental glutathione transf erase
Malondialdehyde
Alanine aminotransferase
Polycyclic aromatic hydrocarbons
Lipopolysaccharides
Blood Lead
—
Cu
Cd
Al
Zn
Pb
Ni
Effects"
Accentuation of liver membrane lipid peroxidation. significant inhibition of
liver antioxidant enzymes. Reduced ratio of reduced glutathione(GSH) to
oxidized glutathione (GSSG),
Copper TNFa Tumor necrosis factor
Cadmium
Aluminum
Zinc
Lead
Nickel
Reference
Sandhir and Gill
(1995)
H
6
o
o
H
O
O
H
W
O
O
HH
H
W
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Table AX5-10.2. Biochemical and Molecular Perturbations in Lead-induced Liver Tissue
to
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>
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H
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O
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O
O
HH
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Concentration Duration Species
Lead - diethyl 0.5-20h Primary hepatocytes
dithiocarbomate
complex, Pb (DTC) 2,
or lead acetate 0.033-
10 uM
— — Primary rat
hepatocytes
— — DBA and C57 mice
100 umol/kg b. wt. i.v Single dose, Male Wistar Albino
single dose analyses performed Rats
12, 24, 48, 72, 96
and 168 h
Blood Lead Effects11
— Effect of interactions between lead and diethyl dithiocarbomate (DTC) on
the enzyme 8 amino levulinic acid dehydratase in primary hepatocytes.
Lipophilic Pb (DTC)2 caused a more rapid and stronger inhibition of ALAD
activity than lead acetate. Lead uptake is higher and more rapid with Pb
(DTC) 2 than lead acetate. This increased inhibition of ALAD activity by
Pb (DTC) 2 might be due to facilitated cellular transport in the complexed
form resulting in higher cellular concentrations of lead.
— DTC decreases cellular effects of Pb and Cd despite unchanged/ even
slightly increased concentrations of the metals. Hepatic ALAD was
significantly inhibited in cells treated with Pb Ac and Pb (DTC)2.
— DBA mice(with a duplication of the ALAD gene accumulated twice the
amount of lead in their blood and had higher lead levels in liver and kidney
than mice with the single copy of the gene (C57), exposed to the same oral
doses of the lead during adult hood. Blood Zinc protoporphyrin (ZPP)
increased with lead exposure in C57 mice and were not affected in DBA
mice
— First in vivo report showing association between lead induced liver
hyperplasia, Glucose - 6 - phosphate levels, and cholesterol synthesis.
Lead treatment increased hepatic de novo synthesis of cholesterol as evident
by increased cholesterol esters and increase of G-6-PD to possibly supply
Reference
Oskarsson and
Lindahl (1989)
Hellstrom- Lindahl
and Oskarsson
(1990)
Claudio et al.
(1996)
Dessi et al. (1984)
Lead nitrate, Single
dose
100 umol/kgb. wt.
Lead nitrate
0-168h
Male Wistar rats
Wistar rats
the reduced equivalents for de novo synthesis of cholesterol. Changes in
these biochemical parameters were accompanied by liver hyperplasia.
Lead nitrate induces hepatic cell proliferation followed by reabsorption of Pani et al. (1984)
excess tissue with in 10-14 days. The proliferation was correlated with
hepatic denovo synthesis of cholesterol, stimulation of hexose
monophosphate shunt pathway and alterations in serum lipo proteins.
Lead nitrate induces multiple molecular forms of Glucose-6- phosphate Batetta et al. (1990)
dehydrogenase with an increase of band 3 and a concomitant increase of
band 1, shifting from the pattern induced by fasting with an increase in
band 1.
-------
Table AX5-10.2 (cont'd). Biochemical and Molecular Perturbations in Lead-induced Liver Tissue
to
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X
ON
H
6
o
Concentration
Lead nitrate, single
i.v.
10uM/100gb. wt.
—
10 or 20 mg/kg as
lead acetate,
subcutaneous
100 uM/kg b. wt lead
nitrate, i.v
2000 ppm lead acetate
in diet.
0-4000 ppm
lead acetate, oral
Duration Species
Multiple time points Male Wistar rats
24-72 h and 20 days
— Rats
Once a wk for Occupationally
5 wks. exposed workers
Rats
36 h post exposure Male Wistar Albino
rats
3 wks Arbor Acres male
Chicks
2 1 days Arbor Acre broiler
chicks
Blood Lead
—
—
Lead-exposed
workers:
0.24-30 nM/mL
Control rats:
0.1 8 nM/mL
10 mg Pb/kg:
2.42 nM/mL:
20 mg Pb/kg:
3.82 nM/mL
—
—
—
Effects"
Lead nitrate exposure results in complete loss of liver glycogen between 24
and 48 h, which was replenished and was found in excess in treated liver
hepatocytes by 20 days. Glycogen synthase and glycogen phosphorylase
activities were diminished by 24 h and return to normal values by day 20.
The pentose phosphate enzymes were upregulated, which coincided highly
with the increase in mitotic rate. Overall lead nitrate induces drastic
alterations in hepatic carbohydrate metabolism along with increased hepatic
cell proliferation.
Lead acetate induced mitotic response much more effectively in renal
epithelial cells than liver cells (675 fold less).
Lead induces lipid peroxidation in serum of manual workers, while blood
superoxide dismutase (SOD) activity decreased. Similar phenomenon was
observed with rats that were subcutaneously injected with lead acetate. At
higher than 20 uM concentration, lead in untreated microsomes increased
NADPH dependent lipid peroxidation.
Endogenous source of newly synthesized cholesterol together with increase
of HMP shunt enzyme activities is essential for hepatic cell proliferation by
lead nitrate
Liver non protein sulphahydryl (NPSH) and glutathione (GSH) were
increased upon lead exposure. The concentrations of liver glutamate,
glycine, and methionine were also elevated upon lead exposure.
Lead increases tissue peroxidation via a relative increase of 20:4 fatty acids.
Decrease in the hepatic ratio of 18:2/20:4 might be specific to lead toxicity
Reference
Hacker et al. (1990)
Calabrese and
Baldwin et al.
(1992)
Ito et al. (1985)
Dessi et al. (1990)
Me Gowan and
Donaldson et al.
(1987)
Donald and
Leeming (1984)
O
H
O
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O
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HH
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W
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Table AX5-10.2 (cont'd). Biochemical and Molecular Perturbations in Lead-induced Liver Tissue
to
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Concentration Duration
Sodium Vanadate, Acute studies,
30 mg/kg 24 h
subcutaneous in mice
30 mg/kg b.wt, i.p. in
rats
0.5 mM
Vanadium sulphate in Chronic studies
drinking water for lOwks
Species
Male Swiss-
Webster mice
Male Sprague
Dawley Rat
Blood Lead Effects"
— Sodium orthovandate increases lipid peroxidation in kidneys of mice and
rats. Malondialdehyde (MDA) formation increased 100%, with in 1 h.
after injection.
In both rat and mice, no significant increase in lipid peroxidation was
observed in brain, heart, lung, and spleen.
Chronic exposure to vanadium, through maternal milk and drinking water
for 10 weeks increased MDA formation and lipid peroxidation in kidneys.
—
Reference
Donaldson et al.
(1985)
chronic treatment
>
X
250-2000 ppm lead
acetate in diet
19 days
1.25-20.00 mg/L lead 30 days
nitrate, oral
Arbor Acre broiler
chicks
Fresh water fish
Dietary Pb consistently increased liver arachidonic acid, the Knowles and
arachidonate/linoleate ratio and hepatic non-protein sulfhydryl Donaldson et al.
concentration. Hepatic microsomal fatty acid elongation activity was (1990)
decreased by Pb. over all these results demonstrate that changes in the
precursors and mechanisms involved with eicosanoid metabolism are not
always reflected in tissue concentrations of leukotriens and prostaglandin.
Lead accumulation in the liver and other tissue increased in a dose Tulasi et al. (1992)
dependent manner up to 5mg/L, exposure to sublethal concentration
(5 ppm) of lead reduced the total lipids, phospholipids, and cholesterol
levels in the liver and ovary. Lead nitrate may affect the fecundity of fish
by altered lipid metabolism.
k
frj
H
6
o
NW'
i — j
z.
—I
^J
/o
r*^S
O
H
W
O
O
HH
H
W
250 mg/L of lead as
lead acetate, oral
35-70 mg, lead intra
gastric
CYP
ALAD
GSH
5 weeks of exposure Weanling female
followed by SD rats
4 weeks of recovery
One or two times a Male Buffalo rats
wk/7 wks.
Cytochrome P-450
Reduced Glutathione
Aminolevulinic acid dehydratase
—
Control: 4.6 ug/dL
Lead 35 mg/kg:
16.8 ug/dL
Lead 70 mg/kg:
32.4 ug/dL
ZPP
HMP
b.wt.
Effect of weight loss on body burden of lead - Weight loss increases the
quantity and concentration of lead in the liver even in the absence of
continued exposure
Decrease in plasma cholesterol, & HDL fraction, increase in serum
triglyceride, atrophy of the elastic fibers in the aorta.
Zinc protoporphyrin
Hexose monophosphate shunt pathway
body weight
Han et al. (1996)
Skoczynska et al.
(1993)
-------
Table AX5-10.3. Effect of Lead Exposure on Hepatic Cholesterol Metabolism
to
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X
Concentration
100 umol/kg body wt,
i.v. lead nitrate
100 umol/kg body wt,
i.v. lead nitrate
0.05 mg/kg body
wt/day. lead acetate,
subcutaneous, with or
without cadmium
acetate 0.025 mg lead
acetate/kg body
wt/day
300 mg/L lead
acetate, oral
Duration
Multiple durations
0, 3, 6, 12, 24, and
48 h
Multiple durations
0, 1,3,6, 12, 18,24,
48, and 72 h
preexposure for
5-7 days, gestation
through lactation.
Gestation through
lactation analyses
done at day 12 and
day 21 postnatal.
Species
Male Sprague
Dwaley (SD)
Rats.
Male Sprague
Dwaley (SD) rats
female Charles
Foster rats
Female Wistar
Rats
Blood Lead Effects"
— Lead nitrate, activates the expression of the SREBP-2 and CYP 51 gene
with out decreasing the serum cholesterol level.
— Lead nitrate effects on hepatic enzymes involved in cholesterol
homeostasis— Demonstrated for the first time sterol independent gene
regulation of cholesterol synthesis in lead nitrate treatment
— Lead and cadmium accumulated in the livers of metal treated pregnant and
lactating rats. Hepatic steroid metabolizing enzyme 17-p-hydroxy steroid
oxidoreductase and UDP glucaronyl transferase were decreased and the
hepatic Cytochrome P-450 content was reduced by the metal exposure.
Lead and cadmium alter liver biochemical parameters, however, combined
exposure had no intensifying effect on liver parameters. When
administered together on similar concentration basis, the major effects are
mediated by cadmium.
Control: 1.13 ug/dL In neonates, decrease in liver Hb, iron, alkaline and acid phosphatase
Lead-exposed levels. Protein, DNA and lipid total amounts were reduced and hepatic
12 d PN: 22.01 ug/dL glycogen content was reduced. Lead intoxication of mothers in gestation
21d PN: 22.77 ug/dL and lactation results in alterations in the hepatic system in neonates and
pups.
Reference
Kojima et al. (2002)
Kojima et al. (2004)
Pillai and Gupta
(2004)
Corpas et al. (2002)
" CYP = Cytochrome P-450
b wt. = body weight
H
6
o
o
H
O
O
H
W
O
O
HH
H
W
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Table AX5-10.4. Lead, Oxidative Stress, and Chelation Therapy
to
o
ON
i
ON
^
Concentration and
Compound Duration
Lead acetate, 8 wks
50 mg/kg b.wt,
intragastric
2,000 ppm, lead 5 wks
acetate, Diet
0.1-1.0 nM
Species
Male Albino
Wistar rats
Male Fisher 344
rats, young and
old
Rat liver
hepatocytes.
Normal and LAN
loaded
Blood Lead
—
Young control:
<1 ug/dL
Young lead-exposed:
38.8 ug/dL
Old control:
<1 ug/dL
Old lead-exposed:
2 1.7 ug/dL
—
Effects Reference
Lead induces accentuation of membrane lipid peroxidation in liver by the Sandir and Gill
changes (decrease) in the activities of several antioxidant enzymes such as (1995)
SOD, Catalase, GPx and Glutathione reductase. Lead exposure also
caused a reduction in GSH/GSSG ratio (reduced to oxidized Glutathione).
Effect of lead on lipid peroxidation in young vs. adult rats- Liver GSSG Aykin-Burns et al.
and malondialdehydehyde levels were significantly higher in young rats (2003)
than adult rats. Blood lead levels were higher in young exposed animals as
compared to adults. In young, lead exposed animals, lead induced
oxidative stress was more pronounced particularly in liver tissue.
Lipid peroxidation as indicated by Malondialdehyde accumulation upon Furono et al. (1996)
exposure to various redox-sensitive metals in cultured rat hepatocytes and
hepatocytes loaded with a-linolenic acid indicated that - Al, Cr and
Manganese, Ni, lead and tin did not effectively induce lipid peroxidation
in these cells.
-. The induction was the highest in ferrous iron treated cells compared to
other metals (Cu, Cd, V, Ni).
FeSO4, VC13, CuSO4,
CdCl2, CoCl2, A1C13,
CrCl3, MnCl2, NiSO4,
Pb(NO3)2, SnCl2,
culture medium
9 h
H
6
o
o
H
O
O
H
W
O
O
HH
H
W
LAN - bovine serum
complex 0.8 mM in
culture medium
5 mgkg-1, lead
acetate, i.p., single
dose followed by
therapy with chelating
agents
Additional 12 h
incubation
Analyses after
6 days of treatment
DMSA, Mi-
ADMSA at multiple
times (0.5, 24 hr,
4th and 5th day
after lead treat
Wistar 6 day old
suckling rats
With any metal, the induction was higher in a-linolenic acid treated cells.
Iron and V induced cell injury in LAN loaded cells was prevented by
addition of DPPD. Cd was a weak inducer of lipid peroxidation under
these conditions
Treatment with DMSA and Mi- ADMSA showed Mi-ADMSA to be more
effective in reducing the skeletal, kidney and brain content of lead.
However there was no difference in reducing the liver lead content
between the two compounds.
Blanusha et al.
(1995)
-------
Table AX5-10.4 (cont'd). Lead, Oxidative Stress, and Chelation Therapy
to
o
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Concentration and
Compound
550 ppm lead acetate,
oral DMSA treatment.
Duration Species
(A) Pb for 35 + 6-7 Wk old male
21 days Sprague-Dawley
(B) Pb 35 days and rats
Pb& DMSA for
21 days
(C) Pb 35 days and
DMSA for
21 days
Blood Lead
Lead-exposed:
50 ug/dL
Lead 35 days:
Ranged from
5-20 ug/dL + DMSA
from 0-240 ug/kg/day
Effects
DMSA reversed the hematological effects of Pb, decreased the blood,
brain , bone, kidney and liver concentration and produced marked lead
diuresis, even when challenged with ongoing Pb exposure.
Reference
Pappasetal. (1995)
(D) Acedified Di H2O
for 35 days and
Di water for
21 days
>
X
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Table AX5-10.4 (cont'd). Lead, Oxidative Stress, and Chelation Therapy
to
o
o
Concentration and
Compound
Duration
Species
Blood Lead
Effects
Reference
>
X
20 mg/kg lead acetate,
i.p.
3 days treatment
Male Albino rats
1.
1.
2.
3.
4
5.
6.
5 mg/per bird /day 30 days
Lead acetate 5 d/wk/8wk
10mg/mL/kg(a)
Ethanol 1 g/
4 mL/kg (b)
a + b = (c)
a + zinc 10 mg/
4 mL/kg + lysine
25 mg/4 mL/kg
(d)
b + 2n + lysine as
in d, oral
a + b + Zn +
lysine as in d
Broiler chicken
—
Male Albino rats Control: 1.75 ug/dL
1.
2.
3.
4.
5.
6.
47.23 ug/dL
2.08 ug/dL
45.37 ug/dL
34. 19 ug/dL
1.84 ug/dL
46.69 ug/dL
1300 ppm lead acetate
in drinking water
5 weeks
C57BL/6 mice
Significant lead induced inhibition of hepatic heme synthesis associated
with decline of mixed function oxidases, depletion in anti oxidants such as
vitamin C. Oral supplementation with vitamin C confers protection
against toxic insult by reversing these parameters
Lead - induced inhibition of 5' mono deiodinase (5'- D) activity in
chickens appeared to be mediated through the lipid peroxidative process.
Influence of lysine and zinc administration on the lead-sensitive
biochemical parameters and the accumulation of lead during exposure to
lead. (1) Lead exposure inhibited blood ALAD activity. Serum enzymes
increased blood and tissue lead levels. (2) Decreased blood and hepatic
glutathione. Some of these effects were enhanced with co-exposure to
ethanol. Simultaneous administration of lysine and zinc reduced tissue
accumulation of lead and most of the lead-induced biochemical alterations
irrespective of exposure to lead alone or lead and ethanol.
Pb treatment resulted in depletion of GSH, increased GSSG and promoted
Malondialdehyde (MDA) production in both liver and brain samples.
DMSA or N- acetyl cysteine (NAC) treatment resulted in reversion of
these observations. DMSA treatment resulted in reduced lead levels in
blood, liver and brain, where as treatment with NAC did not reduce these
levels.
Vij etal. (1998)
Chaurasia et al.
(1998)
Tandon et al. (1997)
Ercal (1996)
H
6
o
0
H
O
CH
o
— ]
w
o
2000 ppm of lead
acetate in drinking
water
500 uM lead acetate
in cells
2000 ppm of lead
acetate in drinking
water
5 weeks followed
by treatment with
succimer DMSA, or
thiol agent NAC
Cells-20 h
Animals 5 weeks
followed by
treatment with a-
lipoic acid
Fisher 344 male
rats
Male fisher rats
and Chinese
hamster ovary
cells
— Lead induces oxidative stress in RBC and these biochemical alterations
are reversed by both a thiol antioxidant (NAC) as well as a chelating agent
DMSA.
— Lead induces oxidative stress, a-lipoic acid (LA)treatment significantly
increased thiol capacity of cells and animals via. increasing glutathione
levels and reducing Malondialdehyde levels, increased cell survival. LA
was not effective against reducing blood or tissue lead levels.
Gurer etal. (1998)
Gurer etal. (1999)
O
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Table AX5-10.4 (cont'd). Lead, Oxidative Stress, and Chelation Therapy
to
o
o
>
X
H
6
o
o
H
O
O
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W
O
O
HH
H
W
Concentration and
Compound
0-500 uM lead acetate
2000 ppm of lead
acetate in drinking
water for 5 weeks
Taurine
1 . 1 kg/day
1 mg Pb2+/kg B.wt ,
i.p. lead acetate
Lead as acetate,
400 mg Pb2+/mL,
drinking water
0.5 mg/mL
L-methionine
100 uM/kgb.wt. lead
acetate,
intramuscular, single
Duration
6h
5 weeks
6th week
4 wks, treatment
with various
antioxidant in the
5th wk
10 days
4 wks post-lead
exposure
3 and 24 h
Species Blood Lead Effects
CHO cells and — Antioxidant Taurine reversed the abnormalities associated with lipid
peroxidation parameters such as increased. Malondialdehyde formation
Fischer 344 rats. Controls: 0.43 ug/dL and decreased Glutathione and enhanced CHO cell survival. However,
Lead-exposed' was no* £Sective in reducing cell and tissue lead burden in CHO cells and
36 4 ug/dL 'ead exPosed Fischer rats.
Lead + Taurine:
33.8 ug/dL
IVRI 2 CQ rats — Lead exposure resulted in increased lipid peroxidation, with tissue specific
changes in liver. Treatment of exposed rats with ascorbic acid and a-
tocopherol lowered the lipid peroxidation.
Kunming mice — L- methionine has an ameliorative effect on lead toxicity-Methionine
reduced the decrease in Hb content and depressed body growth caused by
lead. Treatment with dietary methione along with lead decreased the
MDA formation as opposed to lead, moderately reversed the decreased
iron content of the organs and decreased organ lead content.
Male Albino rats — Lead exposure resulted in significant increases in acid and alkaline
phosphatases, serum GOT and GPT, elevated liver and kidney lipid
peroxidation and decreased antioxidant enzymes at 3 and 24 h after
Reference
Gureretal. (2001)
Patraetal. (2001)
Xieetal. (2001)
Othman and El
Missiry (1998)
100 ug/ lead acetate,
intra gastric, oral and
intraperitoneal,
treated with or with
out thiamin (25,
50 mg/kg b.wt) and or
Ca EDTA (50 mg/kg
B.wt
3 days
CD-I mice
exposure. Selenium administration prior to lead exposure produced
pronounced prophylactic effects against lead exposure by enhancing
endogenous anti oxidant capacity.
Two times more whole body lead was retained by intraperitoneal injection
as compared to intragastric administration. Thiamin treatment increased
the whole body retention of both intragastric and intraperitoneal lead by
about 10%. Calcium EDTA either alone or in combination with thiamin
reduced the whole body retention of lead by about 14% regardless of the
route of exposure. Regardless of the route Ca EDTA in the combined
treatment reduced the relative retention of lead in both in liver and kidney.
These studies indicate the combination treatment with thiamin and Ca
EDTA alters the distribution and retention of lead in a manner which
might have therapeutic application.
Kim et al. (1992)
-------
Table AX5-10.4 (cont'd). Lead, Oxidative Stress, and Chelation Therapy
to
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O
Concentration and
Compound
2000 ppm lead acetate,
oral
I chelators
LA, DMSA,
MiADMSA
LA + DMSA + LA+
MiADMSA
0.1% lead as acetate in
drinking water
DMSA - 50 mg/kg,
i.p./day
MiADMSA 50 mg/kg,
i.p./day
Vitamin E 5 mg/kg
and vitamin C
25 mg/kg/ day, i.v.
and oral
500 mg/kg lead
acetate daily, oral
treatment with
chelators
Lead as acetate 0.2%
in drinking water
LA 25 mg/kg b.wt and
DMSA 20 mg/kg b.wt
b.wt.
aCYP
SOD
GSH
GSH/GSSG Ratio
MDA
Al
As
Duration Species
4 wks, 5 days of Male Wistar
treatment with albino rats
antioxidant or
chelators
3 months Male Wistar rats
5 days post-lead
exposure
Multiple durations Male Albino rats
(2, 4, and 6 wks)
5 wks followed by Male Albino rats
a 6th wk
administration of
LA and or DMSA
body weight
Cytochrome P-450
Super oxide dismutase
Glutathione
Reduced Glutathione/Oxidized Glutathione
Malondialdehyde
Aluminum
Arsenic
Blood Lead
Normal: 1.42 ug/dL
Lead: 40.93 ug/dL
Lead + chelators:
38. 5-4.27 ug/dL
—
—
Control: 0.32 ug/dL
Lead-exposed:
0.48-0.56 ug/dL
Lead + chelators:
0.32-0.36 ug/dL
—
Cr
V
Pb
NAC
FeSO4
A1C13
VC13
CdCl2,
Effects
Treatment with all the chelators reduced hepatic GSH and reduced GSSG
levels. Significant beneficial role of Alpha-lipoic acid (LA), in recovering
the altered biochemical parameters, however showed no chelating
properties in lessening body lead burden either from blood, liver, or
kidney. Most beneficial effects against lead poisoning was observed with
combined treatment of lipoic acid and either DMSA (meso 2,3 -
dimercaptosuccinic acid) or MiADMSA (Mono isoamyl DMSA).
Single or combined administration of vitamin C, a-tocopherol and the
chelators DMSA and Mi ADMSA against the Parameters of lead induced
oxidative stress- thiol chelators and the vitamins could bring the blood
ALAD to normal levels, most significantly by combined administration of
Mi ADMSA with vitamin C. Vitamin C and E were effective against
reducing oxidized glutathione ( GSSG), and thibarbituric acid reactive
substance(TBARS) and increasing catalase activity. MiADMSA and
DMSA with vitamin C were effective in increasing hepatic GSH levels.
In summary combined treatment regimens with thiol chelators and
vitamins seem very effective in reducing the lead induced Oxidative
stress.
Impact of combined administration of vitamin C and Sylimarin on lead
toxicity. Combined treatment of lead-exposed animals with vitamin C and
Silymarin showed marked improvement of the adverse biochemical,
molecular and histopathological signs associated with lead toxicity.
Lead treatment for 5 weeks resulted hepatic enzymes alanine
transaminase, aspartate transaminase, and alkaline phosphatase, increased
lipid peroxidation, and decreased hepatic anti oxidant enzymes. LA or
DMSA alone, partially abrogated these effects, however, in combination
completely reversed the lipid oxidative damage.
Cromium CuSO4 Copper sulphate
Vanadium CrCl3 Cromium chloride
Lead MnCl2 Manganese chloride
N acetyl cysteine NiSO4 Nickel sulphate
Ferrous sulphate CoCl2 Cobalt chloride
Aluminum chloride LAN a Linolenic acid
Reference
Pande and Flora
(2002)
Flora et al. (2003)
Shalan et al. (2005)
Sivaprasad et al.
(2004)
Vanadium chloride DPPD DPPD^V-W Diphenyl -p-phynylene-diamine
Cadmium chloride LA Lipoic acid
DMSA Monoisoamyl DMSA
MiDMSA Mi monoisoamyl DMSA
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Table AX5-10.5. Lead-induced Liver Hyperplasia: Mediators and Molecular Mechanisms
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Concentration
Duration
Species
Blood Lead
Effects"
Reference
>
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Rat
Apoptosis plays a major role in the regressive phase of lead nitrate
induced hepatic hyperplasia as detected by the apoptotic bodies by in situ
end labeling and H&E sections of the hepatic tissue. H&E scores mostly
cells in apoptosis phase II, ISEL (in situ end labeling) scores for cells in
phase I. Combination of these two methods is suggested for the better
understanding of the extent and nature of apoptotic process in liver cells
treated with chemicals.
Nakajima et al.
(1995)
A. Lead nitrate, 3 and 15 days Male Wistar
100 uM/kg b.vrt, Albino rats
intra-gastric
B. Diethyl
nitrosoamine
200 mg/kg b.vrt, i.p.
0-100uMPb Multiple time points REL liver cells
sulphate, Pb ranging from 24 h
monoxide, Pb up to 7 days.
chloride and Pb
acetate up to 1 mM,
culture media.
Choline Ig/kg/day in 0, 20 and 24 h Male and female
drinking water rats , partial
hepatectomy
Lead nitrate, 6 h - 4 wks Adult male Albino
75 uM/kg b.wt, single rats
i.v.
75 umol/kg b. vrt. Analyses at 72 h Male Wistar
Lead nitrate in adult Albino Rats.
and 20 ug/mL in the
young, i.v.; single
dose
— Mitogenic stimuli (3 days lead nitrate treatment) and complete regression
(15 days after the treatment), affected the apoptosis differentially.
Influence of apoptosis Vs necrosis on the growth of hepatocytes initiated
by diethyl nitrosamine followed by lead nitrate treatment indicated that the
regenerative response elicited by a necrogenic dose of CCL4 promoted
GSTP (Placental glutathione), a pre-neoplastic marker positive cells as
against the lead nitrate that induced the apoptosis.
— Lead compounds showed a dose and time related effect on REL liver cell
proliferation with varying potencies specific to the different lead salts. Pb
acetate was the most effective and Pb monoxide, the least effective. On 1
hr treatment none of the compounds tested affected the intracellular
communication.
— PKC isozymes during liver cell regeneration — PKC 8 showed a
pronounced increase 20h after partial hepatectomy. a, p, and Zeta at 24 h
corresponding with S-phase. Sexual dimorphism matching with sexual
differences in DNA synthesis was evident. Administration of choline was
able to modulate the protein kinase C isozyme pattern in females in
relation to DNA synthesis and c-myc expression. Taken together the data
positively implicates a, p, and Zeta in growth after partial hepatectomy
and 8 in negative regulation.
— Lead induced significant increase in liver weight. Increased 3H
Thymidine incorporation. Lead induces extensive hypomethylation in
treated rat livers. Site-specific effect on methylation was confirmed at
Hpa II, Msp I, Hae III.
— Effect of lead nitrate on the 5- methyl deoxy cytidine (5-mdcyd) content
and the Hpall, MSPI, Hae III restriction patterns of hepatic DNA from
young, middle aged and senescent rats. The results indicated that the
methylation pattern of genomic DNA changed significantly with age and
the methylation patterns were differentially affected in all the three
populations.
Columbano et al.
(1996)
Apostoli et al.
(2000)
Tessitore et al.
(1995)
Kanducetal. (1991)
Kanduc & Frisco
(1992)
-------
Table AX5-10.5 (cont'd). Lead-induced Liver Hyperplasia: Mediators and Molecular Mechanisms
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Concentration
10 umol/lOOgbody
weight lead nitrate,
i.v.
Duration
Multiple analyses
up to 40 h
Species
Male Wistar Rats,
hepatocytes from
partial
hepatectomy and
lead nitrate
treatment.
Blood Level Effects"
— The kinetics of DNA synthesis and expression of Proto oncogenes in
partially hepatectamized liver cells and lead nitrate treated hepatic cells
indicated peak DNA synthesis after 24 h in the formal and after 36 h in the
later case. Both proliferative stimuli induced c-fos, c-myc and c-Ha Ras
expression. Induced c-myc expression persisted for up to 40h during the
lead nitrate- induced liver cell proliferation. Lead induces hepatic
hyperplasia through changes in proto-oncogene expression.
Reference
Coni et al. (1989)
100 umol/kg, b. wt.
lead nitrate, i.v.
>
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O
100 umol/kgb. wt.
Analyses at multiple
time points 0.25 -
24 h
8h
Male Wistar
Albino Rats
100 umol/kgb. wt.,
i.v.
100 umol/kgb. wt.,
i.v. lead nitrate
Multiple analyses
time points,
1-120 h
Analyses at multiple
time points 8 h to
15 days
Male Sprague
Dawley rats
Male Wistar rats
Male Wistar rats
proliferative stimuli by means of lead nitrate exposure resulted in
increased expression of c-jun m-RNA where as compensatory Coni et al. (1993)
regeneration in partially hepatectamized cells occurred through increased
expression of c-fos and c-jun. Different mitogenic stimuli induced
differential expression of these protooncogenes, in addition had a different
profile than cells from partial hepatectomy despite the cell cycle timings
being the same in some cases.
In rat liver, in addition to a few hepatocytes four types of non parenchymal Rijhsinghani et al.
cells namely, fibroblasts, macrophages, bile ducts and periductular cells (1992)
proliferate in response to lead nitrate treatment. This growth is not related
to adaptive response secondary to parenchymal enlargement. However,
such growth in parenchymal cells seems dormant and does not play a
functional role in adult liver epithelial growth.
Both mRNA levels and enzyme activity of DNA polymerase p markedly Menegazzi et al.
increased before and/or during DNA synthesis in proliferating hepatocytes (1992)
in lead nitrate treated and partially hepatectomized rats. 5 fold increase in
the enzyme activity was observed 8 h after lead nitrate administration. In
the regenerative liver cells a 3 fold increase was observed 24-48h after
partial hepatectomy.
Lead nitrate induced Poly (ADP-ribose) polymerase mRNA 24 hr after Menegazzi et al.
exposure. A 2 fold increase in the mRNA levels of the enzyme occurred (1990)
two days after the exposure. Such changes were also observed in hepatic
cells from livers of partial hepatectomy. These changes preceded the
increase in DNA synthesis and remained high during the time of extensive
DNA synthesis.
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Table AX5-10.5 (cont'd). Lead-induced Liver Hyperplasia: Mediators and Molecular Mechanisms
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2
o
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W
Concentration
30 mg/kg b. wt. lead
nitrate
30 mg/kg b. wt. lead
nitrate
100 umol/kgb. wt.
lead nitrate, i.v.,
single dose.
A. Mitosis -
Lead nitrate-
100 uM/kg, i.v.
Ethylene
dibromide
100 mg/kg , intra
gastric
Cyproterone
acetate, 60 mg/kg
intra gastric.
B. Hepatocyte
nodules diethyl
nitrosamine
200 mg/kg
Lead nitrate, single
i.v. 100 urn/ kg b.wt
LPS- 12.5 ug/rat, post
Pb nitrate treatment.
Duration Species Blood Lead
Multiple time points Adult male and —
up to 8 days female rats
Multiple time point Adult male and —
up to 60 h female rats
Multiple time point Male Wistar Rats Serum lead
analyses ranging concentrations
from 12 - 168 h peaking to
50-60 ug/dL between
12-24 hand
remaining up to
40 ug/dL up to 108h
30' - 3 h Adult male Wistar —
rats
Multiple analyses at Male Wistar rats —
3,6, 12, 24 and 36 h
Effects"
Lead nitrate induced liver hyperplasia exhibited sexual dimorphism where
mitogenic action was less effective and was delayed in females as
compared with males. Pre administration with choline partially filled
these sexual differences.
Lead nitrate induced liver hyperplasia exhibited sexual dimorphism. Pre
administration with choline partially filled these sexual differences.
Significant down regulation of PKC p and PKC a activities occurred
during lead induced proliferation
Effect of lead nitrate on protein kinase C (PKC) activity. A single dose of
lead nitrate resulted in enhanced activity of PKC in the purified particulate
fraction of the rat liver, reached its peak activity by 24 h which lasted for
48 h. This was accompanied by increased frequency of mitotic cells.
These results indicate, lead nitrate induced PKC activity may play a role in
liver cell proliferation.
Liver cell proliferation by enhanced DNA synthesis was observed with the
mitogens Cyproterone acetate, ethylene dibromide, and lead nitrate as
early as 30 mints after treatment and persisted even after 5 days of
treatment by lead nitrate administration.
hepatocytes isolated from pre neoplastic liver nodules have also exhibited
enhanced cell proliferation.
Stimulation of hepatocyte cell proliferation by lead nitrate was not
accompanied by changes in liver levels of Hepatocyte growth factor
(HGF), Transforming growth factor-a (TGF-a), or TGF-pl m-RNA.
Lead nitrate treatment resulted in the enhancement of Tumor necrosis
factor a at a time preceding the onset of hepatocyte DNA synthesis,
indicating its role in lead induced hepatic cell proliferation. The survival
of lead nitrate treated rats decreased significantly with an after treatment
of LPS (lipopolysaccharide).
Reference
Tessitore et al.
(1994)
Tessitore et al.
(1994)
Liuetal. (1997)
Coni(1991)
Shinozuka et al.
(1994)
-------
Table AX5-10.5 (cont'd). Lead-induced Liver Hyperplasia: Mediators and Molecular Mechanisms
to
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Concentration
Duration
Species
Blood Lead
Effects"
Reference
>
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O
15 mg/kg b. wt. lead
acetate
Pb+ LPS group
analyzed after 14 h
and the rest after
24 h after lead
administration
Male Sprague
Dawley rat
— Lead augments the lethality of endotoxin lipopolysaccharide (LPS) in rats
and enhances liver injury, which is further enhanced by TNF. Lead + LPS
treatment increased both serum TNF concentrations and TNF area as
compared to LPS alone, simultaneous administration of lead with either
LPS or TNF, serum aspartate transaminase, alanine transaminase, alkaline
phosphatase, glutamyl trans peptidase and plasma triglyceride levels were
markedly increased
Honchel et al.
(1990)
Lead nitrate
100 uM/kg
b. wt. i.v. single dose
100 umol/kgb. wt
lead nitrate, single,
i.v.
100 umol/kgb. wt.
single i.v.,
100 umol/kgb. wt.
i.v. single dose
Multiple time points
of analyses
extending up to 48 h
after treatment
Multiple time points
of analyses up to 48
h
Male Wistar rats
Multiple time points Male Sprague
of analyses up to 80 Dawley rats
h
Multiple time points Male Wistar rats
of analyses up to 24
h
100 umol/kg b. wt. Multiple time point Male Sprague
i.v., single dose analyses up to 96 h Dawley rats
Lead nitrate and ethylene bromide induce liver cell proliferation via Ledda-Columbano
induction of TNFa. Dexa methasone, a known TNF inhibitor, decreases et al. (1994)
TNF expression and liver cell proliferation by these mitogens. These
studies support the fact that TNF might mediate hepatic cell proliferation
by lead nitrate and ethylene bromide.
Lead nitrate (LN) treatment resulted in increased Brdu incorporation of Shinozuka et al.
hepatocytes and non parenchymal cells at 12 h after treatment and reached (1996)
the peak index at 36 h. Rats given a single iv of recombinant TNFa
enhanced proliferation in non parenchymal cells after 24 h, the labeling of
hepatocytes at 36 h. NAF, Nafenopin another mitogen which does not
induce liver TNFa, increased the number of labeled hepatocytes without
increasing the labeling of non parenchymal cells indicating that only lead
nitrate induced proliferation is mediated by TNFa and these mitogens
initiate proliferation in different cells based on their capacity to stimulate
TNFa production.
Lead nitrate induces liver cell proliferation in rats without accompanying Kubo et al. (1995)
liver cell necrosis. This proliferation involves enhanced TNF mRNA and
levels but not hepatocyte growth factor. The role of TNF in lead nitrate
induced liver cell proliferation is supported by the inhibition of TNF and
reduced hepatocyte proliferation by several TNF inhibitors.
Lead nitrate induced liver cell proliferation involves TNF a production, Menegazzi et al.
enhanced NF-KB activation increased hepatic levels of iNos mRNA as (1997)
opposed to other mitogens such as Cyproterone acetate or Nafenopine.
The role of neurotrophins, the nerve growth factor (NGF), the brain Nemoto et al.
derived neurotrophic factor (BDNF) and neurotrophin -3 (NT-3) in lead (2000 )
nitrate treated liver cells was studied. LN, treatment resulted in increased
in the levels of NGF, BDNF and NT-3. The increase in neurotrophin
receptors and the gene expression were correlate with liver weights. This
study demonstrates that lead nitrate induced hyperplasia may be mediated
by neurotrophins.
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Table AX5-10.5 (cont'd). Lead-induced Liver Hyperplasia: Mediators and Molecular Mechanisms
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Concentration
Duration
Species
Blood Lead
Effects"
Reference
>
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Multiple doses
0-50 uM, culture
medium
50 uM lead acetate,
culture medium
10 uM/110gb. wt,
single i.v.
lOumol/lOOgb. wt.
single i.v.
Multiple time points Hepatocytes from
up to 24 h Adult male Swiss-
mice, primary
24 h Rat hepatocyte
and Kupffer cell
and granulocyte
co-cultures
Multiple time point Adult male Wistar
analyses up to Rats
5 days
Multiple time points Male Wistar rats
up to 9 days
10 mmol/lOOg, lead
nitrate, i.v.
Multiple time points Male Wistar rats
Interaction between Pb and cytokines in hepatotoxicity- Pb potentiated Sieg and Billing
cytokine -induced oxidative stress by decreasing GSH and increased (1997)
efflux of Oxidized glutathione (GSSG). Combined treatment resulted in a
decline in intra cellular ATP concentration
Lead stimulates intercellular signaling between Kupffer cells and Milosevic and
hepatocytes which increased synergistically at low lipopolysaccharide Maier (2000)
levels. These signals promote proteolytic hepatocyte killing in
combination with a direct cellular interaction between the granulocytes
and hepatocytes.
Lead nitrate induced hepatocyte apoptosis was prevented by pre- Pagliara et al.
treatment with gadolinium chloride, a Kupffer cell toxicant - Role for (2003a)
Kuffer cell in hepatocyte apoptosis
Lead nitrate-induced liver hyperplasia in rats results in a significant Dini et al. (1993)
increase in the expression of aceyl glycoprotein receptor (ASGP-R) during
the involutive phase of lead nitrate induced hyperplasia in rat-liver, which
coincided with the massive death by apoptosis of the same cells.
A significant rise in the galactose-specific receptors was also observed
3 days after the treatment. These studies demonstrate that carbohydrate
receptors regulate lead nitrate induced liver cell apoptosis.
Demonstration of the expression of carbohydrate receptors on Kupffer Ruzittu et al. (1999)
cells. Lead nitrate induced apoptosis in Kupffer cells and internalization
of apoptotic cells (Phagocytes) is mediated by both Mannose and
Galactose receptors.
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Table AX5-10.5 (cont'd). Lead-induced Liver Hyperplasia: Mediators and Molecular Mechanisms
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O
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O
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Concentration
Pb (No3)2, , i.v.
100uM/110g.b. wt
GdCl3
0.75 mg/100 g. b. wt,
i.v.
In vitro, 10 mM lead
nitrate
Multiple
concentrations
varying from
300 nM-10 uM,
up to 100 uM in
certain in vitro expts
0-10 uM lead acetate
in the culture medium
Duration
1, 3, and 5 days.
2, 4, or 24 h before
lead nitrate injection.
Analyses at multiple
time points up to 24 h
in Hep G2 cells and at
24 and 48 h in
Kupffer cells
1,2,4 and 6 days
24 & 48 h
Species
In vivo Adult
male Wistar rats
Hep G2 cells
Hepatoma cell
line, H4- II-C3
H4-IIE - C3
hepatoma cell
culture model
Blood Lead Effects" Reference
— Hepatic apoptosis induced by lead nitrate in vivo is abolished by gadolium Pagliara (2003b)
chloride, a Kupffer cell toxicant that suppresses Kupffer cell activity and
reduces to half the apoptotic rate. Lead nitrate treatment also deprives the
hepatic cells from reduced glutathione and this process is reversed by
Gadolium chloride. Lead nitrate induces apoptosis in Kupffer cells, and
HepG2 cells in vitro.
— Acute effect of lead on glucocorticoid regulation of Tyrosine Heiman and Toner
aminotransferase (TAT) in hepatoma cells -Lead treatment does not (1995)
significantly alter initial glucocorticoid receptor number or ligand binding.
Lead may perturb PKC mediated phosphorylations in the glucocorticoid-
TAT signal transduction system. Lead also may be increasing the
turnover of TAT by actions at transcription, translation and /or post
translation.
— In HTC cells glucocarticoid signal transduction pathways involve calcium- Tonner and Heiman,
mediated events and PKC isoforms , lead exposure interferes with calcium (1997)
mediated events and aberrant modulation of PKC activities and may
contribute to the over all toxicity of lead.
b. wt. = body weight
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Table AX5-10.6. Effect of Lead Exposure on Liver Heme Synthesis
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Table AX5-10.6 (cont'd). Effect of Lead Exposure on Liver Heme Synthesis
to
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o
>
X
Concentration
Lead 500 ppm in
drinking water
0.5 or 2. 4 uM lead
acetate in culture
medium
500 ppm lead in
drinking water
A. Cu deficient diet- 1
mg/kg Cu in the
diet
Duration
14 days
Analyses at multiple
time points, 0-28 days
Rat exposure 62 days
Human occupational
exposure
0.3-38 yrs.
4 wks
Species
Male ddY mice
Hepatocyte
cultures on 3T3
cells
A. .Male Wistar
rats
B. Lead smelt
workers, males
Weanling
Sprague Dawley
rats
Blood Lead Effects"
— Urinary excretion of (3- Aminoisobutyric acid (ABA) and 8-aminolevulinic
acid (ALA) increased significantly in mice exposed to lead in drinking
water for 14 days. The degree of increasing excretion for ALA was higher
than urinary ABA. Liver and kidney ALA dehydratase was inhibited,
while ALA synthatase was not affected.
— Hepatocyte cultures on 3T3 cells produce and excrete porphyrins for 28
days. Lead exposure for 4 weeks alters cell morphology and produces
cytotoxic effects that could be monitored by altered porphyrin excretion.
— Lead exposure significantly increases the urinary ALA (Aminolevulinic
acid) and Coproporphyrins (CP-III>CP-I in rats and exposed workers.
Urinary 5-hydroxy indole acetic acid was not influenced by lead exposure.
— High Zn in the diet reduces plasma copper, but not plasma ceruloplasmin
activity or the recovery of plasma copper or ceruloplasmin activity after
oral copper sulphate of Cu-deficient rats. High dietary Zn also modifies
Reference
Tomokuni et al.
(1991)
Quintanilla-Vega
etal. (1995)
Ichiba and
Tomokuni (1987)
Panemangalore and
Bebe (1996)
B. Moderately
deficient- 2 mg/kg
C. High Zn diet
60 mg/kg b. wt.
the response of plasma SOD activity to dietary copper, but does not
influence RBC SOD activity
H
6
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O
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W
O
O
HH
H
W
1200 mg/kg b. wt. 4 wks
lead acetate in diet,
Sub acute toxic
studies 400 mg/lead
0- 1 00 uM lead acetate 1 9 h
in the culture medium
Broiler chickens
Primary Rat and
chick embryo
hepatocyte
cultures.
— Liver porphyrin levels increased during lead toxicosis. Concurrent
administration of selenium or monensin in the feed further enhances this
process.
— Formation of Zn protoporphyrins in cultured hepatocytes- Lead did not
specifically increase Zinc protoporphyrin accumulation or alter iron
availability in cultured hepatocytes.
Khan & Szarek
(1994)
Jacobs et al. (1998)
-------
Table AX5-10.6 (cont'd). Effect of Lead Exposure on Liver Heme Synthesis
to
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Concentration
Duration
Species
Blood Lead
Effects"
Reference
X
-------
>
X
oo
Table AX5-10.7. Lead and In Vitro Cytotoxicity in Intestinal Cells
to
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Compound and
Concentration
HgCl2, CdCl2, Ti2
S04, Pb(N03)2-
concentration not
given clearly,
Butathionine, up to
ImM
Glutathione 1 mM
N- Acetyl cysteine,
ImM
Duration Species
Cell proliferation 1-407, Intestinal
assays 48 h epithelial cell
line.
Glutathione depletion
assays 48 h
Sulphahydryl
repletion studies.
Blood Lead Effects"
— Rank order cytotoxicity of various metal salts in 1-407 intestinal epithelial
cells in terms of LC50 values- HgCl2 (32 uM) > CdCl2 (53 uM), CuCl2
(156nM)>Ti2SO4
(377 uM)>Pb (NO3)2(1.99 mM)
Role of Glutathione, in the cytotoxicity of these metals by the assessment
of GSH depletion by Butathionine sulfoxamine pretreatment at non
cytotoxic concentration increased the toxicity of HgCl2 (5. 7-fold), and
CuCl2(1.44-fold).
Administration of glutathione, with either HgCl2 or CdCl2 did not protect
the cells against the toxicity.
Reference
Keogh et al. (1993)
Af-acetyl cysteine reduced the cytotoxicity of mercury.
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Table AX5-10.8. Lead and Intestinal Uptake - Effect on Ultrastructure, Motility, Transport, and Miscellaneous
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Compound and
Concentration
Lead acetate, 0.1%, in
drinking water.
Duration
Multiple analyses at
2, 30, and 60 days
after lead exposure.
Species
Male Wistar rats
Blood Lead Effects"
— Small intestinal goblet cells are involved in lead detoxification.
Lead treatment for 30 days produces characteristic goblet cells in the
Authors
Tomczok et al.
(1988)
Prolonged exposure to lead more than 30 days caused silver sulphide
deposition (indicative of heavy metal deposition) in the mucus droplets of
cytoplasmic goblet cells.
>
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to
100 mg/lead acetate/kg.
b. wt.
Added lead concentration
in the milk - 0-80 ug/mL
Multiple analyses at
2, 30 and 60 days
Male Wistar rats
Adult & Infant
rats (16 days)
Fresh or frozen rat
or Avian milk
Lead poisoning changes the ultra structure of intestine.
30 day lead exposed rat intestinal enterocytes showed numerous, small
rough-membraned vesicles and prominent, dilated golgi complexes, in
their cytoplasm.
By 60th day, lead-exposed rats had a vacuolated cytoplasm and prominent
golgi filled with vacuoles.
90% of Pb in rat and bovine milk was found associated with caseine
micelles regardless of whether the milk is labeled in vitro or in vivo with
203 Pb. Similarly lead in infant milk formula was also predominantly
associated with casein, however, to a much lower extent than rat and
bovine milk formulae.
Tomczok et al.
(1991)
Beach and Henning
(1988)
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Pb as lead acetate, for
0.5-10.0 uM, ZnasZn
acetate 0, 5, 10, or 50 uM
Temperature variation
Expts, 5 uM Pb, and
incubated for 10 mints at
4, 22, or 37 °C
5, 10, 30 or 60 mints,
Simultaneously with
lead for 10 minutes
Incubation time
10 minutes
IEC-6 normal rat
intestinal
epithelial cells.
Lead tracer studies indicated that in infant rats, as the milk traversed
through the intestine, in the collected luminal fluid, Pb was primarily
associated with casein curd and remained as a non precipitable, non-
dialyzable fraction as it moved to the small intestine, indicating that Pb
remains with protein fraction as it traverses through the stomach and small
intestine fraction
Pb uptake by IEC-6 cells depends on the extracellular Pb concentration.
Pb transport in IEC-6 cells is time and temperature dependent, involves
sulphahydryl groups, and is decreased by the presence of Zn.
Dekaney et al.
(1997)
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Table AX5-10.8 (cont'd). Lead and Intestinal Uptake - Effect on Ultrastructure, Motility, Transport, and Miscellaneous
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Compound and
Concentration
OECD
(Organisation for
Economic Co-operation
and Development)
medium was artificially
contaminated at 1, 3, 5, or
10 times the Dutch
intervention value of
530 mg/Pb/kg dry wt.
Lead containing medium
was presented at the
apical surface of the cells
in 2 mL DMEM/chyme.
Duration
Cell viability
studies — 24 h
incubation.
Lead transport
studies, 1, 3, 5 and
24 h
Species Blood Lead Effects"
— Transport of bioaccessible lead across the intestinal epithelium — In Coco-
2 cells exposed to artificial chime, with in 24 hrs. App. 27% of the lead
was associated with the cells and 3% were transported across the cell
monolayer. Lead associated with cells showed a linear relationship with
the lead available in the system.
Results indicate that only a fraction of the bioavailable lead is transported
across the intestinal epithelium.
On the basis of lead speciation in chime, It could be attributed that
dissociation of labile lead species, such as lead phosphate, and lead bile
complexes and subsequent transport of the released free metal ions flow
toward the intestinal membrane.
Authors
Oomen et al. (2003)
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Neutral red uptake
studies had
DMEM/chime with low
5uM and high 50uM
lead content
44 mg/kg/day lead as
53 mmol/L lead acetate
2.5 mg/mL lead acetate in
drinking water
100 uM lead nitrate, in
vitro
4 weeks Rat —
55 days Colonic segments Exposed:
taken from 80 ug/dL
chronically
exposed guinea
pigs
Duration not specified Muscle - —
myenteric plexus
preparations of
distal ileum of
controlled animals
Lead exposure significantly decreases the amplitude of contraction in rat
duodenum.
Colonic propulsive activity as measured by the velocity of the
displacement of the balloon, from the oral to the aboral end, did not get
affected significantly by lead treatment. In longitudinal muscle-myenteric
plexus preparations of distal ileum, addition of lead nitrate (100 urn)
caused slight increase in cholinergic contractions.
Moderate decrease of electrically induced cholinergic contractions.
Karmakar and
Anand (1989)
Rizzietal(1989)
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Table AX5-10.9. Lead, Calcium, and Vitamin D Interactions, and Intestinal Enzymes
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Compound and
Concentration Duration
Ca- 0.5% in diet (low 10 days
Calcium)
1.2% in diet (high
calcium)
Species
White Leghorn
Cockerels
Blood Lead Effects11
— Dietary lead affects intestinal Ca absorption in two different ways
depending on the dietary Ca status.
A. In chicks fed low Ca diet (0.05%), ingested lead inhibited intestinal
47Ca absorption, intestinal Calbindin D, and alkaline phosphatase
synthesis in a dose dependent fashion.
Authors
Fullmer and Rosen
(1990)
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Pb chloride.
Ca-0.1%or 1.2% in the
diet with lead-0.1 -
0.8% as Lead chloride in
the diet
1 or 2 weeks
Leghorn
Cockerels
B. In normal calcium diets (1.2%) lead exposure had no bearing on the
intestinal Ca absorption, or Calbindin D, or Alkaline phosphatase
synthesis and in fact elevated their levels at higher lead
concentrations.
These results indicate that the primary effects of lead in both cases,
occur at or prior to intestinal protein synthesis involving
Cholecalciferol endocrine system.
- Dietary Ca deficiency, initially
(1* week) stimulates Ca absorption and Calbindin D levels regardless of
dietary Pb intake.
- At 2 weeks, this response is reversed by lead.
- Intestinal lead absorption was enhanced by Ca deficiency initially and
was inhibited by prolonged dietary lead intake.
- Intestinal Pb absorption was increased in adequate Ca situation, but only
after 2 weeks at the lower levels of dietary Pb.
Fullmer (1991)
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Ca-O.l-l.2%
Pb-0.8%
2 weeks
White Leghorn
Cockerels
Interactions between dietary lead and Ca-influence on serum vitamin D
levels.
- Lead ingestion and Ca deficiency alone or in combination generally
increased serum 1,25 (OH)2 D levels over the most of the range of
dietary lead and Ca.
- In severe Ca deficiency, Pb ingestion resulted in significant decreases in
hormone concentration.
- Similarities in response profiles for 1,25 ( OH)2 D, intestinal Ca
absorption and Calbindin- D suggested major interactions between lead
and calcium mediated changes via circulatingl,25(OH)2 D
concentration.
Fullmer (1997)
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Table AX5-10.9 (cont'd). Lead, Calcium, and Vitamin D Interactions, and Intestinal Enzymes
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Compound and
Concentration
Lead, Alkaline
phosphatase and Ca2+
ATPase
2.0- 10.0 mM
Duration Species
Incubation time- not Male Albino rats
specified
Blood Lead Effects"
Lead inhibited the activity of several intestinal brush border enzymes such
as Ca2+- ATPase, Sucrase, y-glutamyl -transpeptidase and acetyl choline
esterase with the exception of alkaline phosphatase. Inhibition of Ca2+-
ATPase was competitive and that of the other enzymes is by non-
competitive means.
Authors
Gupta etal. (1994)
Lead, Sucrase 0.5 mM -
6.0mM
Lead, y-glutamyl
transpeptidase 1.0-10 mM
Lead, Acetyl choline
esterase 10.00-35.00 mM
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Oral lead in Similac or
apple juice adjusted for
attainment of blood lead
levels 35-40 ug/dL.
Succimer 30 mg/kg/day
204 Pb 24.5 nM followed
by 206 Pb 352 nM, Single
dose
Administered from 8th
day post partum, until
age 26 weeks.
Two successive 19
days at age 53 weeks
and 65 weeks.
Administered
immediately before
chelation
Female infant Lead-exposed: Effect of oral succimer chelation on the Gastro intestinal absorption and
Rhesus Monkeys 35-40 ug/dL the whole body retention of lead—
Radio isotope Pb tracer technique—
Succimer significantly reduced Gastro intestinal absorption of lead and
increased urinary excretion of lead—
The initial decrease in whole body lead by 10% was over come when
majority of administered tracer was retained in the body after 5 days of
treatment
Creminetal. (2001)
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ANNEX TABLES AX5-11
May 2006 AX5-187 DO-NOT QUOTE OR CITE
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Table AX5-11.1. Lead-Binding Proteins
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Source
Goyer(1968a)
Goyer(1970a,b)
Choie and Richter (1972)
Moore etal. (1973)
Moore &Goyer( 1974)
Shelton and Egle (1982)
Egle and Shelton (1986)
Oskarsson et al. (1982)
Mistry etal. (1984)
Fowler and DuVal (1991)
Organ Species
Kidney Rat
Kidney Rat
Kidney Rat
Kidney Rat
Kidney Rat
Kidney Rat
Brain Rat, mouse,
dog, guinea
pig, and
chicken
Kidney Rat
cytosol &
brain
Kidney Rat
cytosol
Kidney Rat
cytosol
Molecular
Weight Protein Properties
Intranuclear lead inclusion
bodies
Lead is concentrated in the
intranuclear inclusion body
Initial inclusion bodies in
cytoplasm
Protein in inclusion bodies is
acidic, with high levels of
aspartic a, glutamic a, glycine
& cystine
Inclusion body
protein is 27.5
kDa
Inclusion body is Named p32/6 . 3
32 kDa with pi of
6.3
p32/6.3 found
11. 5 and 63 kDa
1 1 .5 kDa, 63 kDa, Respective Kd values: 13, 40
> 200 kDa 123 nM
Cleavage product of alpha-2
microglobulin
Inducible Separation Technique
Yes
Yes
Yes
Yes
Yes Acrylamide gel electrophoresis
Yes Two-dimensional gel electrophoresis
No(?)
No 203Pb binding followed by Sephadex G-
75 or G-200 chromatography, then
SDS-PAGE
No 203Pb binding followed by Sepharose-
6B column chromatography
No Chromatography followed by reverse
phase HPLC, then production of
antibodies. Kd 10-8 M
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Table AX5-11.1 (cont'd). Lead-Binding Proteins
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Table AX5-11.1 (cont'd). Lead-Binding Proteins
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g Source
Church etal. (1993a)
Church etal. (1993b)
Xie etal. (1998)
Organ Species
RBC Human lead
workers, one
asymptomatic
and one
symptomatic
RBC Human lead
workers
RBC Human lead
workers
Molecular
Weight
6-7 kDa
5, 7 and 12 kDa,
pi 4.7-4.9
240 -260 kDa,
< 30 kDa
Protein Properties
First pt had 67% of RBC Pb
bound to protein. Second pt
had 22% of RBC Pb in
protein
30 % cysteine Thought to be
MT on basis of greater UV
abs at 254 nm than 280 nm
High M.Wt. Peak identified
asALAD. Low M.Wt. peak
Inducible Separation Technique
Yes RBC hemolysate filtered through
Amicon YM 30 membrane. Superose
12 column. Lead quantitated by A.A.
Yes Superose 12, Amicon YM 30, Amicon
YM 2, HPLC
Yes Bio-gel A column. Pb determined by
AA.
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Goering & Fowler,
(1987a)
Goering & Fowler,
(1987b)
Kidney
Kidney and
liver
Rat
Rat
seen after adding lead in vitro
Pre-treatment with zinc
before injecting 203Pb leads to
zinc-thionein binding Pb
Pre-Rx with Zn or Cd induces
Zn or Zn, Cd-MT. The MT
decreases Pb inhibition of
ALAD
No 203Pb binding, Sephadex G-75
No 203Pb binding, Sephadex G-75
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Qu et al. (2002); Waalkes
et al. (2004)
Kidney
MT-null
phenotypic
Pb-exposed MT-null
developed no Pb inclusion
bodies, accumulated less
renal Pb than WT
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AX6. CHAPTER 6 ANNEX
May 2006 AX6-1 DRAFT-DO NOT QUOTE OR CITE
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ANNEX TABLES AX6-2
May 2006 AX6-2 DRAFT-DO NOT QUOTE OR CITE
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Table AX6-2.1. Prospective Longitudinal Cohort Studies of Neurocognitive Ability in Children
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
United States
Bellinger etal. (1992)
U.S.
148 subjects from the Boston Prospective Study
were re-evaluated at 10 years of age. The WISCR
was used to index intellectual status. Extensive
assessment of medical and sociodemographic
covariates.
Cord and serial postnatal
blood lead assessments.
Cord blood lead grouping
<3, 6-7, >10 ug/dL.
Blood lead at 2 years 6.5
(SD 4.9) ug/dL
Increase of 10 ug/dL in blood lead level at age two was
associated with a decrement of approximately 6 IQ points.
Relationship was stronger for verbal compared to
performance IQ. Prenatal exposure to lead as indexed by
cord blood lead levels was unrelated to psychometric
intelligence.
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Dietrich et al.
(1991, 1992, 1993a);
Ris et al. (2004)
U.S.
Canfield et al. (2003a)
U.S.
253-260 children followed since birth in the
Cincinnati Lead Study were re-evaluated at 4, 5,
and 6.5 years of age. At 4 and 5 years the KABC,
was used to index intellectual status. At 6.5 years,
the WISCR was administered. At 15-17 years of
age, 195 Cincinnati Lead Study subjects were re-
evaluated with a comprehensive
neuropsychological battery that yielded a
"Learning/IQ" factor in a principal components
analysis. Extensive assessment of medical and
sociodemographic covariates.
172 predominantly African-American, lower
socioeconomic status children in Rochester, NY
followed since they were 5 to 7 months were
evaluated at 3 and 5 years. An abbreviated form of
the Stanford-Binet Intelligence Scale-4 (SBIS-4)
was used to index intellectual status. Extensive
assessment of medical and sociodemographic
covariates.
Prenatal (maternal) and serial
postnatal blood lead
assessments.
Prenatal blood lead 8.3
(SD 3.7) ug/dL
Blood lead at 2 years 17.4
(SD 8.8) ug/dL
Serial postnatal blood lead
Blood lead at 2 years
9.7 ug/dL
Few statistically significant relationships between blood
lead indices and covariate-adjusted KABC scales at 4 and
5 years of age. One KABC subscale that assesses visual-
spatial skills was associated with late postnatal blood lead
levels following covariate adjustment. After covariate
adjustment, average postnatal blood lead level was
significantly associated with WISCR performance IQ at
6.5 years. Blood lead concentrations in excess of 20 ug/dL
were associated with deficits in performance IQ on the
order of 7 points compared with children with mean blood
lead concentrations of less than 10 ug/dL. At 15-17 years,
late childhood blood lead levels were significantly
associated with lower covariate-adjusted Learning/IQ
factor scores.
Following covariate adjustment, there was a significant
inverse relationship between blood lead indices and IQ at
all ages. Overall estimate indicated that an increase in
average lifetime blood lead concentration of 1 ug/dL was
associated with a loss of 1A IQ point. Effects were stronger
for subjects whose blood lead levels never exceeded
10 ug/dL. Semiparametric analysis indicated a decline in
IQ of 7.4 points for a lifetime average blood lead
concentration up to 10 ug/dL while for levels between
10 and 30 ug/dL a more gradual decrease in IQ was
estimated. Authors concluded that the most important
aspect of their findings was that effects below 10 ug/dL
that have been observed in previous cross-sectional studies
have been confirmed in a rigorous prospective
investigation.
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Table AX6-2.1 (cont'd). Prospective Longitudinal Cohort Studies of Neurocognitive Ability in Children
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Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
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United States (cont'd)
Bellinger and
Needleman (2003)
U.S.
Chen et al. (2005)
U.S.
Reanalysis of data from the Boston Prospective
Study focusing on 48 subjects at 10 years of age
whose blood lead levels never exceeded 10 ug/dL.
WISCR was used to index intellectual status, (see
Bellinger, etal. (1992)
Repeat measure psychometric data on 780 children
enrolled in the Treatment of Lead-Exposed
Children (TLC) clinical trial for were analyzed to
determine if blood lead concentrations at 2 years of
age constitute a critical period of exposure for the
expression of later neurodevelopmental deficits.
Data for placebo and active drug groups were
combined in these analyses, which spanned the
ages of approximately 2 to 7 years of age.
Measures of intellectual status included the Bayley
Mental Development Index (MDI), and full scale
IQ derived from age-appropriate Wechsler scales.
Serial postnatal blood lead
Blood lead at 2 years 6.5
(SD 4.9) ug/dL
Blood lead
Range 20-44 ug/dL
Baseline blood lead 26
(SD 26.5) ug/dL in both drug
and placebo groups.
Blood lead at 7 years 8.0
(SD 4.0) ug/dL
IQ was inversely related to two-year blood lead levels
following covariate adjustment. Blood lead coefficient
(-1.56) was greater than that derived from analyses
including children with concentrations above 10 ug/dL
(-0.58). Authors conclude that children's IQ scores are
reduced at lead levels still prevalent in U.S.
Association between blood lead and psychometric
intelligence increased in strength as children became older,
whereas the relation between baseline (2 year) blood lead
and IQ attenuated. Peak blood lead concentration thus
does not fully account for the observed association in older
children between their lower blood lead concentrations and
IQ. The effect of concurrent blood lead on IQ may
therefore be greater than currently believed. Authors
conclude that these data support the idea that lead exposure
continues to be toxic to children as they reach school age,
and does not lend support to the interpretation that
majority of the damage is done by the time the child
reaches 2 to 3 years of age.
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Table AX6-2.1 (cont'd). Prospective Longitudinal Cohort Studies of Neurocognitive Ability in Children
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
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Europe
Wasserman et al. (1992,
1994, 2003); Factor-
Litvaketal. (1999)
Yugoslavia
Birth cohort of approximately 300-400 infants
followed since birth residing in two towns in
Kosovo, Yugoslavia, one group near a
longstanding lead smelter and battery
manufacturing facility and another in a relatively
unexposed location 25 miles away. Intellectual
status was monitored from 2 to 10-12 years of age
with the Bay ley Scales of Infant Development,
McCarthy Scales of Children's Abilities, and
WISCIII. Extensive assessment of medical and
sociodemographic covariates.
Maternal prenatal, umbilical
cord and serial postnatal blood
lead
Maternal blood lead in:
exposed area 19.9 (SD 7.7)
ug/dL, unexposed area 5.6
(SD 2.0) ug/dL
Umbilical cord blood lead in:
exposed area 22.2 (SD 8.1)
ug/dL, unexposed area 5.5
(SD 3.3) ug/dL
Blood lead at 2 years in:
exposed area 35.4 ug/dL,
unexposed area 8.5 ug/dL
Rise in postnatal blood lead from 10 to 30 ug/dL at two
years of age associated with a covariate-adjusted decline of
2.5 points in Bay ley MDI. Maternal and cord blood lead
not consistently associated with Bayley outcomes. Higher
prenatal and cord blood lead concentrations associated
with lower McCarthy General Cognitive Index (GCI)
scores at 4 years. Scores on the Perceptual-Performance
subscale particularly affected. After covariate-adjustment,
children of mothers with prenatal blood lead levels
>20 ug/dL scored a full standard deviation below children
in the lowest exposure group (<5 ug/dL prenatal blood
lead). Postnatal blood lead also associated with poorer
performance. Increase in blood lead level from 10-25
ug/dL was associated with a reduction of 3.8 points in GCI
after covariate-adjustment. Effects even more pronounced
on the Perceptual-Performance subscale. At 7 years,
significant inverse associations between lifetime average
blood lead and WISCIII IQ were observed, with
consistently stronger associations with Performance IQ and
later blood lead measures. Adjusted intellectual loss
associated with an increase in lifetime average blood lead
from 10-30 ug/dL was over 4 points in WISCIII Full-Scale
and Performance IQ. At 10-12 years, subjects were again
assessed with the WISCIII. Following covariate-
adjustment, average lifetime blood lead was associated
with all components of the WISCIII with effect sizes
similar to those observed at 7 years. In most instances,
bone lead-IQ relationships were stronger than those for
blood lead among subjects residing near the lead smelter.
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Table AX6-2.1 (cont'd). Prospective Longitudinal Cohort Studies of Neurocognitive Ability in Children
to
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Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
X
ON
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Latin America
Schnaas et al. (2000)
Mexico
Gomaa et al. (2002)
Mexico
112 children followed since birth with complete
psychometric data from the Mexico City
Prospective Study were examined. Intellectual
status was indexed with the General Cognitive
Index (GCI) from the McCarthy Scales of
Children's Abilities (MSCA). Purpose of the study
was to determine if the magnitude of the effect of
postnatal blood lead levels on cognition varies with
the time between blood lead and cognitive
assessments.
197 two year-olds residing in Mexico City
followed since birth. The Bay ley Scales of Infant
Development Mental Development Index (MDI)
was used to index intellectual status. Extensive
assessment of medical and sociodemographic
covariates.
Serial postnatal blood lead
Average blood lead
24-36 months 9.7
(range 3-48) ug/dL.
Umbilical cord and serial
postnatal blood lead
Umbilical cord blood
lead 6.7
(SD 3.4) ug/dL
Blood lead at 2 years 8.4
(SD 4.6) ug/dL.
Maternal tibial and patellar
bone lead
Patellar (trabecular)
bone lead
17.9 (SD 15.2) ug/g
A number of significant interactions observed between
blood lead levels and age of assessment. Greatest effect
observed at 48 months where a 5.8 deficit in adjusted GCI
scores was observed for each natural log increment in
blood lead. Authors concluded that four to five years of
age appears to be a critical period for the manifestation of
earlier postnatal blood lead level effects on cognition.
Umbilical cord blood lead and patellar (trabecular) bone
lead were significantly associated with lower scores on the
Bayley MDI. Maternal trabecular bone lead levels
predicted poorer sensorimotor functioning at two years
independent of the concentration of lead measured in cord
blood. Increase in cord blood lead level from 5-10 ug/dL
was associated with a 3.1 point decrement in adjusted MDI
scores. In relation to lowest quartile of trabecular bone
lead, the second, third, and fourth quartiles were associated
with 5.4, 7.2, and 6.5 decrement in MDI following
covariate adjustment. Authors concluded that higher
maternal trabecular bone lead concentrations constitute an
independent risk factor for impaired mental development
in infancy, likely due to the mobilization of maternal bone
lead stores over gestation.
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Table AX6-2.1 (cont'd). Prospective Longitudinal Cohort Studies of Neurocognitive Ability in Children
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Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
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Latin America (cont'd)
Tellez-Rojo et al.
(in press)
294 one and two year-olds residing in Mexico City
followed since birth. The Bay ley Scales of Infant
Development-II (MDI and PDI) were used to index
developmental status. There was extensive
assessment of medical and sociodemographic
covariates.
Umbilical cord blood lead and
blood lead at 12 and
24 months.
Umbilical cord blood lead
4.8 (SD 3.0) ug/dL.
Blood lead at 1 year
4.27(SD2.1)ng/dL
Blood lead at 2 years
4.3 (SD 2.2) ug/dL
Blood lead at 12 months was not associated with MDI at
either age. Blood lead at 24 months was significantly
associated with 24 month MDI. An increase of one
logarithmic unit in 24 month blood lead level was
associated with a reduction of approximately 5 points in
MDI. Findings for PDI were similar. In comparison to a
subsample of subjects with blood lead levels greater than
10 ug/dL, the coefficient for blood lead was significantly
larger for infants never exceeding that level of internal
dose. A steeper inverse slope was observed over the blood
lead range up to 5 ug/dL (-1.71 points per 1 ug/dL
increase in blood lead, p = 0.01) compared to the range
between 5 and 10 ug/dL (-0.94 points, p = 0.12); however,
these slopes were not significantly different (p = 0.34).
In conclusion, a major finding of this prospective study
was that a significant inverse relationship between blood
lead concentration and neurodevelopment was observed
among children whose blood lead levels did not exceed
10 ug/dL at any age.
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Australia
Baghurst et al. (1992);
McMichael et al.
(1994); long etal.
(1996)
Australia
400-500 subjects residing in and near Port Pirie,
Australia and followed since birth were re-evaluated
at 7 to 8 and 11-13 years of age. WISCR was used
to index intellectual status at both ages. Extensive
assessment of medical and sociodemographic
covariates.
Maternal prenatal, umbilical
cord and serial postnatal blood
lead
Antenatal average blood lead
10.1(SD3.9)ug/dL
Umbilical cord blood lead
9.4 (SD 3.9) ug/dL
Blood lead at 2 years
geometric mean 21.3
(SD 1.2) ug/dL
Deciduous central incisor
whole tooth lead
Tooth lead geometric 8.8
(SD1.9)ug/g
Significant decrements in covariate-adjusted full scale IQ
were observed in relationship to postnatal blood lead levels
at both ages. At seven to eight years a loss of 5.3 points
was associated with an increase in blood lead from 10 to
30 ug/dL. At 11-13 years mean full scale IQ declined by
3.0 points for an increase in lifetime average blood lead
concentrations from 10 to 20 ug/dL. Lead levels in central
upper incisors were also associated with lower 7-8 year IQ
following covariate adjustment. Adjusted estimated
decline in IQ across the range of tooth lead from 3 to
22 ppm was 5.1 points.
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Table AX6-2.1 (cont'd). Prospective Longitudinal Cohort Studies of Neurocognitive Ability in Children
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Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
Australia (cont'd)
Cooneyetal. (1991)
Australia
175 subjects from the Sydney, Australia Prospective
Study were assessed at 7 years of age. The WISCR
was used to index intellectual status. Extensive
assessment of medical and sociodemographic
characteristics.
Maternal and cord blood lead
Cord blood lead 8.4 ug/dL
(SD not given)
Blood lead at 2 year 15.8
ug/dL (SD not given)
Blood indices of lead exposure were not associated with
any measure of psychometric intelligence. Authors
conclude that the evidence from their study indicates that if
developmental deficits do occur at blood lead levels
<25 ug/dL, the effect size is likely to be small (<5%).
Sydney results are difficult to interpret from the statistical
presentation in their report. It is not clear which covariates
were entered into regression analyses nor is the empirical
or substantive basis for their conclusion.
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Asia
Shenetal. (1998)
China
Pregnant women and newborns in Shanghai, China
recruited from health care facilities in the community
on the basis of cord blood lead concentration
percentiles (30th and 70th) yielding a total N of 173
subjects. TheBayley Scales of Infant Development
Mental Development Index (MDI) and Psychomotor
Development Index (PDI) were used to index
sensorimotor/intellectual status at 3, 6, and 12
months. Extensive assessment of medical and
sociodemographic characteristics.
Cord blood lead
Cord blood lead "high group"
13.4 (SD 2.0) ug/dL
"low group" 5.3 (SD 1.4)
ug/dL
Blood lead at 1 year
"high group" 14.9
(SD 8.7) ug/dL
"low group" 14.4
(SD 7.7) ug/dL
At all ages the Bayley MDI was associated with cord
blood lead groupings following adjustment for covariates.
Postnatal blood lead unrelated to any Bayley measures.
Differences in MDI between prenatal blood lead exposure
groupings generally in accord with similar investigations
in Boston, Cincinnati, and Cleveland. Authors conclude
that the adverse effects of prenatal lead exposure are
readily discernible and stable over the first year of life.
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Table AX6-2.2. Meta- and Pooled-Analyses of Neurocognitive Ability in Children
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Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
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United States
Lanphear et al. (2005)
International
Needleman and
Gatsonis(1990)
International
Schwartz (1994)
International
Pooled analysis of seven international prospective
studies involving 1,333 school-age children.
Primary outcome measure was full-scale IQ as
assessed by age-appropriate Wechsler scale.
Measures of exposure were concurrent, peak,
average lifetime and "early" blood lead (i.e. mean
blood lead from 6-24 months). Cord blood lead was
also investigated for those studies that collected
these samples at birth. Multivariate regression
models were developed adjusting for site as well as
10 common covariates. Blood lead measure with the
largest adjusted R2 was nominated a priori as the
preferred index related lead exposure to IQ in
subsequent analyses. Results evaluated by applying
a random-effects model.
Meta analysis of 12 studies chosen on the basis of
quality—covariate assessment and application of
multiple regression techniques. Studies weighted on
basis of sample size. Studies divided according to
tissue analyzed (blood or teeth). Joint p-values and
average effect sizes calculated using two different
methods.
Meta analysis of 7 recent studies relating blood lead
to IQ were reviewed, three longitudinal and four
cross-sectional. Measure of effect was estimated
decrease in IQ for an increase in blood lead from
10-20 ug/dL. Studies were weighted by the inverse
of the variances using random
Umbilical cord blood lead
Serial postnatal blood lead
Lifetime average blood lead
12.4 (range 4.1-34.8) ug/dL
Blood lead
Tooth lead
Blood lead
Concurrent blood lead level exhibited the strongest
relationship with IQ, although results of regression analyses
for all blood lead variables were similar. Steepest declines
in IQ were at blood lead concentrations below 10 ug/dL.
For the entire pooled data set, a decline of 6.2 IQ points
(95% CI: 3.8-8.6) was observed for an increase in blood
lead from 1-10 ug/dL.
Joint p-values for blood lead studies were <0.0001 for both
methods while for teeth joint p-values of <0.0006 and
<0.004 were obtained. Partial correlations ranged from
-0.27 to -0.0003. No single study was responsible for the
significance of the final findings. Authors concluded that
the hypothesis that lead lowers children's IQ at relatively
low dose is strongly supported by results of this quantitative
review.
Estimated decrease in IQ for increase in blood lead from
10-20 ug/dL was -2.6 points (SE 0.41). Results were not
determined by any individual study. Effect estimates
similar for longitudinal and cross-sectional studies.
For studies with mean blood lead concentrations below
15 ug/dL estimated effect sizes were larger. When the
study with the lowest exposures was examined alone using
nonparametric smoothing (Boston), no evidence of a
threshold was observed down to a blood lead level of
1 ug/dL. Author concludes that these data provide further
evidence of lead effects on cognition at levels below
10 ug/dL.
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Table AX6-2.2 (cont'd). Meta- and Pooled-Analyses of Neurocognitive Ability in Children
to Reference, Study
§ Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
United States (cont'd)
Pococketal. (1994)
International
Meta-analysis of five prospective and fourteen cross-
sectional studies (including tooth and blood tissues)
were included. The fixed effect method of
Thompson and Pocock (1992) was employed.
Only blood lead at or near two years of age was
considered for the prospective studies.
Blood lead
Tooth lead
Overall conclusion was that a doubling of blood lead levels
from 10-20 ug/dL, or tooth lead from 5-10 ug/g was
associated with an average estimated deficit in IQ of
around 1-2 points. Authors caution interpretation of these
results and lead literature in general citing questions
surrounding the representativeness of the samples, residual
confounding, selection bias, and reverse causality.
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Table AX6-2.3. Cross-sectional Studies of Neurocognitive Ability in Children
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Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
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United States
Lanphear et al. (2000)
U.S.
Emory et al. (2003)
U.S.
Chiodo et al. (2004)
U.S.
4,853 U.S. children ages six to 16 years enrolled in
NHANES-III. Two subtests of the WISC-R (Block
Design and Digit Span) used to assess intellectual
status. Medical and sociodemographic covariates were
assessed
77 healthy, lower-risk African-American infants age 7
months. The Fagan Test of Infant Intelligence (FTII)
was administered to assess intellectual status. Birth
weight and gestational age examined as potential
covariates/confounders.
237 African-American inner-city children assessed at
7.5 years of age. Cohort was derived from a larger
study of the effects of prenatal ETOH exposure on
child development. 83% of children in lead study had
little or no gestational exposure to ETOH. WISC-III
was administered to assess intellectual status. Medical
and sociodemographic covariates were assessed.
Blood lead at time of testing
Geometric blood Lead 1.9
(SE 0.1) ug/dL 2.1%
with blood lead > 10 ug/dL
Maternal blood lead
Blood lead 0.72
(SD 0.86) ug/dL
Blood lead at time of testing
Blood lead 5.4
(SD 3.3) ug/dL
Multivariate analyses revealed a significant association
between blood lead levels and both WISC-R subtests.
Associations remained statistically significant when
analyses were restricted to children with blood lead levels
below 10 ug/dL. Authors caution that lack of control for
parental intelligence and variables like the HOME scale
should temper any conclusions regarding observed effects.
Infants scoring in the upper 5th to 15th percentiles for the
FTII had mother with significantly lower maternal blood
lead levels when compared to those scoring in the lower
5th or 15th percentile. Findings of this study should be
considered preliminary due to small sample size and lack
of covariate assessment or control.
Following covariate adjustment statistically significant
relationships between blood lead and full-scale, verbal and
performance IQ were observed. Significant effects of lead
on full-scale and performance IQ was evident at blood lead
concentrations below 7.5 ug/dL.
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Europe
Walkowiak et al.
(1998)
Germany
Prpic-Majic et al.
(2000)
Croatia
384 six-year-old children in three German cities. Two
subtests of the WISC (Vocabulary and Block Design)
used to estimate IQ. Both subscales were combined to
form a "WISC Index." Medical and sociodemographic
covariate covariates were assessed.
275 third and fourth grade students in Zagreb, Croatia.
WISC-R was administered to assess intellectual status.
Covariate factors limited to parents' educational status
and gender of child.
Blood lead at time of testing
Blood lead 4.2 ug/dL
95th percentile 8.9 ug/dL
Blood lead at time of testing
Blood lead 7.1
(SD 1.8) ug/dL
Following covariate-adjustment, WISC Vocabulary was
significantly associated with blood lead but combined
WISC index was borderline. Authors conclude that
findings roughly correspond with those of other studies
that find effects below 10 ug/dL but caution that
potentially important covariates such as HOME scores
were not controlled.
Following covariate adjustment, no statistically significant
associations were observed for lead or other indicators of
toxicity (ALAD, EP) on WISC-R. Authors argue that
study had sufficient power and that the "no-effect"
threshold for lead must be in the upper part or above the
study's range of exposures.
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Table AX6-2.3 (cont'd). Cross-sectional Studies of Neurocognitive Ability in Children
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Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
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Latin America
Kordas et al. (2004,
2006)
Mexico
602 first grade children in public schools in a highly
industrialized area of northern Mexico. Premise of
study was that effects of lead could be explained by
correlated nutritional factors such as iron status,
anemia, and growth. Peabody Picture Vocabulary
Test-Revised (PPVT-R), Cognitive Abilities Test
(CAT), and an abbreviated form of the WISC-R were
administered to assess intellectual status. Medical and
sociodemographic covariates were assessed.
Blood lead at time of testing
Blood lead 11.5
(SD 6.1) ug/dL
Counter etal. (1998)
Ecuador
77 chronically lead-exposed children living in
Ecuadorian villages where lead is used extensively in
commercial ceramics production. Ravens Colored
Progressive Matrices (RCPM) used to index
intellectual status. Only half of the sample was
assessed. No assessment of medical or
sociodemographic covariates.
Blood lead at time of testing
Blood lead 47.4
(SD 22) ug/dL
Following covariate adjustment blood lead levels were
significantly associated with poorer performance on the
PPVT-R, WISC-R Coding, and Number and Letter
Sequencing, a Math Achievement Test, and the Sternberg
Memory Test. Authors concluded that lead's association
with iron deficiency anemia or growth retardation could
not explain relationship between lead and cognitive
performance. Non-linear analyses of selected
neurocognitive outcomes revealed that dose-response
curves were steeper at lower than at higher blood lead
levels. Moreover, the slopes appeared negative at blood
lead levels below 10 ug/dL, above which they tend to
plateau. Effects of lead on neurocognitive attainment
appeared to be greatest among the least advantaged
members of the cohort.
Simple regression analysis revealed a correlation between
blood lead and RCPM of only borderline significance.
Results difficult to interpret because there was no attempt
to age-adjust. When analysis restricted to children 9 to
11 years of age, a highly significant negative correlation
was obtained. Study has little relevance to the question of
lead hazards in the U.S. because of unusually high levels
of exposure.
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Asia
Rabinowitz et al.
(1991)
Taiwan
Bellinger et al. (2005)
India
443 children in grades one to three in Taipei City and
three schools near lead smelters. Ravens Colored
Progressive Matrices (RCPM) used to index
intellectual status. Medical and sociodemographic
covariate factors were assessed.
74 four to fourteen year-old children residing in
Chennai, India were enrolled in the study, 31 of which
were assessed with the Binet-Kamath Intelligence test.
Data were collected on sociodemographic features of
subjects' families.
Dentin tooth lead
Taipei City 4.3
(SD 3.7) ug/g
Smelter areas 6.3
(SD 3.3) ug/g
Blood lead at time of testing
Blood lead 11.1
(SD 5.6) ug/dL
Scores on the RCPM were negatively correlated with tooth
lead concentrations. In multivariate analyses, parental
education was the most important predictor of RCPM
scores, but tooth lead concentrations still significantly
predicted lower scores in females residing in low-income
families.
Covariate-adjusted blood lead coefficient was negative but
nonsignificant, perhaps due to small sample size and
highly variable performance of subjects with the least
elevated blood lead concentrations.
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S Table AX6-2.3 (cont'd). Cross-sectional Studies of Neurocognitive Ability in Children
^ Reference, Study
O Location, and Period Study Description Lead Measurement Findings, Interpretation
Middle East
Al-Saleh et al. (2001) 533 Riyadh, Saudi Arabia girls (6-12 years of age) Blood lead at time of testing Blood lead levels had no impact on TONI scores but this
were administered a variety of standardized tests Blood lead 8.1 (SD 3.5) test has limited evidence of validity in this population.
including the TONI, and the Beery VMI. Extensive ug/dL Significant negative associations were noted between
data were collected on potentially confounding blood lead levels and the Beery VMI suggesting an
variables including sociodemographic variables, early association between impairment in visual-spatial skills in
developmental milestones and child health status. Saudi children with blood lead levels in the range of 2.3 to
27.4 ug/dL.
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Table AX6-2.4. Effects of Lead on Academic Achievement in Children
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Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
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United States
Lanphear et al. (2000)
U.S.
Needlemanetal. (1990)
U.S.
Design: Cross-sectional. 4,853 U.S. children ages
six to 16 years enrolled in NHANES-III. Subjects
were administered the Arithmetic and Reading
subtests of the Wide Range Achievement Test-
Revised (WRATR). A number of medical and
sociodemographic covariates were assessed and
entered into multivariable models.
Blood lead at time of testing
Geometric blood lead 1.9
(SE0.1)ng/dL. 2.1%
with blood lead > 10 ng/dL
Design: Prospective cohort. Re-examination of the
Chelsea and Somerville cohort recruited in the
1970's (Needleman et al., 1979). 132 adolescents
were recalled. Large battery of tests was
administered to examine neurobehavioral deficits
and academic achievement in high school and shortly
following graduation. Extensive assessment of
medical and sociodemographic covariates.
Tooth (dentin) lead
Tooth lead median
8.2 iig/g
Following covariate adjustment, a statistically significant
relationship between blood lead and WRATR performance
was found. A 0.70 point decrement in Arithmetic scores
and a 1 point decrement in Reading scores for each
1 ng/dL increase in blood lead concentration was observed.
Statistically significant inverse relationships between blood
lead levels and performance for both Reading and
Arithmetic subtests were found for children with blood
lead concentrations <5 |ig/dL. Authors concluded that
results of these analyses suggest that deficits in academic
skills are associated with blood lead concentrations lower
than 5 |ig/dL. They cautioned, however, that two
covariates that have been shown to be important in other
lead studies (i.e., parental IQ and HOME scores) were not
available. This may have over or under estimated deficits
in academic skills related to lead. They further caution
that, as with all cross-sectional studies utilizing blood lead
as the index of dose it is not clear whether deficits in
academic skills were due to lead exposure that occurred
sometime during early childhood or due to concurrent
exposure. Nevertheless, concurrent blood lead levels
reflect both ongoing exposure and preexisting body
burden.
Subjects with dentin lead levels >20 ppm were at higher
risk of dropping out of high school (adjusted OR = 5.8,
95% CI: 1.4-40.7) and of having a reading disability
(adjusted OR: 5.8, 95% CI: 1.7-19.7). Higher dentin lead
levels were also significantly associated with lower class
standing, increased absenteeism, and lower vocabulary and
grammatical reasoning scores on the Neurobehavioral
Evaluation System (NES). Authors conclude that undue
exposure to lead has enduring and important effects on
objective parameters of success in life.
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Table AX6-2.4 (cont'd). Effects of Lead on Academic Achievement in Children
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Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
United States (cont'd)
Bellinger et al. (1992)
U.S.
Design: Prospective longitudinal. 148 children in
the Boston Lead Study cohort were examined at
10 years of age. The short-form of the Kaufman Test
of Educational Achievement (KTEA) was used to
assess academic achievement. Primary outcome was
the Battery Composite Score. Extensive assessment
of medical and sociodemographic covariates.
Cord and serial postnatal
blood lead assessments.
Cord blood lead grouping
<3, 6-7, >10 ug/dL.
Blood lead at 2 years 6.5
(SD 4.9) ug/dL
After covariate-adjustment, blood lead levels at 24 months
were significantly predictive of lower academic achievement
(P = -0.51, SE 0.20). Battery Composite Scores declined by
8.9 points for each 10 ug/dL increase in blood lead. This
association was significant after adjustment for IQ. Authors
conclude that lead-sensitive neuropsychological processing and
learning factors not reflected in measures of global intelligence
may contribute to deficits in academic achievement.
Levitonetal. (1993)
U.S.
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Design: Prospective cohort. Teachers of
approximately 2000 eight year-old children bom in
1 hospital in Boston between 1979 and 1980 filled
out the Boston Teachers Questionnaire (BTQ) to
assess academic performance and behavior. Limited
information is provided on the assessment of
covariate factors but a number were considered and
controlled for in multivariable analyses.
Cord blood lead
Cord blood lead 6.8 ug/dL
Tooth (dentin) lead
Tooth lead 2.8 ug/g
Following adjustment for potential confounding variables,
elevated dentin lead concentrations were associated with
statistically significant reading and spelling difficulties as
assessed by the BTQ among girls in the sample. Authors
conclude that their findings support the case for lead-associated
learning problems at levels that were prevalent at that time in
the general population. However, authors add that the inability
to assess child-rearing quality in this study conducted by mail
limits the inferences that can be drawn.
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Australia
Fergusson et al. (1993,
1997); Fergusson and
Horwood(1993)
New Zealand
Design: Prospective cohort. Academic performance
was examined in a birth cohort of 1200 New Zealand
children enrolled in the Christchurch Health and
Development Study. Measures of academic
performance at 12-13 years included the Brut
Reading Test, Progressive Achievement Test, Test of
Scholastic Abilities, and teacher ratings of classroom
performance in the areas of reading, writing, and
mathematics. The growth of word recognition skills
from 8 to 12 years was also examined using growth
curve modeling methods. Academic achievement in
relationship to lead was re-examined in this cohort at
18 years. Measures of academic achievement
included the Burt Reading Test, number of years of
secondary education, number of certificates passed
(based on national examinations), and leaving school
without formal qualifications (failing to graduate).
Extensive assessment of medical and social
covariates.
Tooth (dentin) lead
Tooth lead 6.2
(SD 6.2) ug/g
Following covariate adjustment, dentin lead levels were
significantly associated with virtually every formal index of
academic skills and teacher ratings of classroom performance
in 12-13 year-olds. After adjustment for covariates, tooth lead
levels greater than 8 ug/g were associated with significantly
slow growth in word recognition abilities with no evidence of
catch up. At 18 years, tooth lead levels were significantly
associated with lower reading test scores, having a reading
level of less than 12 years, failing to complete three years of
high school, leaving school without qualifications, and mean
number of School Certificates passed. Authors conclude that
early exposure to lead is independently associated with
detectable and enduring deficits in children's academic
abilities. They further conclude that their findings are
particularly significant in that they confirm the findings of
Needleman (1990), albeit in a cohort with lower levels of
exposure to environmental lead.
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Table AX6-2.4 (cont'd). Effects of Lead on Academic Achievement in Children
to
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Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
X
ON
Asia
Wang et al. (2002a)
Taiwan
Rabinowitz et al. (1992)
Taiwan
Design: Cross-sectional. 934 third graders living in
an urban industrial area of Taiwan. Outcome
variables were grades for Chinese (reading, writing),
mathematics, history, and natural science. Grades
were converted into individual class rankings to
avoid teacher bias. Limited data on medical and
sociodemographic covariates.
Design: Cross-sectional. Teachers of 493 children
in grades 1-3 filled out the Boston Teachers
Questionnaire (BTQ) to assess academic
performance and behavior. Sociodemographic and
medical covariate factors were assessed.
Blood lead at time of
evaluation
Blood lead 5.5
(SD1.9)ng/dL
Tooth (dentin) lead
Tooth lead 4.6
(SD 3.5) ug/g
Following covariate adjustment, blood lead was
significantly associated with lower class ranking in all
academic subjects. Major shortcoming of this study is lack
of control for potentially important covariates such as
parental IQ. However, the relatively low levels of
exposure in this sample and strength and consistency of the
reported relationships suggest that lead may be playing
some role in lowering academic performance.
Prior to adjustment for covariates, girls with higher
exposures to lead evinced a borderline significant trend for
reading difficulties while byes displayed significantly
increased difficulties with respect to activity levels and
task attentiveness. In logistic regression models that
include significant covariate factors, the tooth lead terms
failed to achieve statistical significance. Authors conclude
that lead levels found in the teeth of children in this
Taiwanese sample are not associated with learning
problems as assessed by the BTQ.
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Table AX6-2.5. Effects of Lead on Specific Cognitive Abilities in Children — Attention/Executive Functions, Learning, and
Visual-Spatial Skills
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
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United States
Bellinger etal. (1994)
U.S.
Stiles and Bellinger
(1993) U.S.
Canfield et al. (2003b,
2004)
U.S.
Design: Prospective cohort. 79 subjects from the
original Chelsea and Somerville, MA lead study
were re-evaluated at 19-20 years of age with the
Mirsky battery of attentional measures. Extensive
measures of medical and sociodemographic
covariates.
Design: Prospective longitudinal. 148 subjects from
the Boston Lead Study were re-evaluated at 10 years
of age with an extensive neuropsychological battery.
Tests included the California Verbal Learning Test,
Wisconsin Card Sorting Test, Test of Visual-Motor
Integration, Rey-Osterieth Complex Figure, Story
Recall, Finger Tapping, and Grooved Pegboard.
Extensive measures of medical and
sociodemographic covariates.
Design: Prospective longitudinal. 170-174 children
from the Rochester Lead Study were administered a
number of learning and neuropsychological
functioning at 48, 54, and 66 months of age. At 48
and 54 months the Espy Shape School Task was
administered while at 66 months the Working
Memory and Planning assessment protocols of the
Cambridge Neuropsychological Test Automated
Battery (CANTAB) was given. Extensive measures
on medical and sociodemographic covariates.
Tooth (dentin) lead
Tooth lead 13.7
(SD11.2 ug/g)
KXRF Bone lead
Tibial bone lead
(range <1 ->10 ug/g)
Patellar bone lead
(range <1 ->15 ug/g)
Cord and serial postnatal
blood lead assessments.
Cord blood lead grouping
Blood lead at 2 years 6.5
(SD 4.9) ug/dL
Serial postnatal blood lead
Blood lead at 2 years
9.7 ug/dL
Lifetime average blood
lead 7.2 ug/dL
(range 0-20 ug/dL)
Higher tooth lead concentrations were significantly
associated with poorer scores on the Focus-Execute and
Shift factors of the Mirsky battery. Few significant
associations were observed between bone lead levels and
performance. Authors conclude that early lead exposure
may be associated with poorer performance on
executive/regulatory functions, which are thought to
depend on the frontal or prefrontal regions of the brain.
Authors point out that the number of significant
associations was about equal to those that would be
expected by chance. However, tasks that assess attentional
behaviors and executive functions tended to among those
for which lead was a significant predictor of performance.
Following covariate adjustment, higher blood lead
concentrations at two year were significantly associated
with lower scores on Freedom from Distractibility factor of
the Wechsler scales, increase in percentage of
perseverative errors on the Wisconsin Card Sorting Test
and the California Verbal Learning Test.
Following covariate adjustment, blood lead level at
48 months was negatively associated with children's
focused attention while performing the Shape School
Tasks, efficiency at naming colors, and inhibition of
automatic responding. Children with higher blood lead
concentrations also completed fewer phases of the Espy
tasks and knew fewer color and shape names. On the
CANTAB battery, children with higher lifetime average
blood lead levels showed impaired performance on spatial
working memory, spatial memory span, and cognitive
flexibility and planning. Authors conclude that the effects
of pediatric lead exposure are not restricted to global
measures of intellectual functioning and executive
processes may be at particular risk.
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Table AX6-2.5 (cont'd). Effects of Lead on Specific Cognitive Abilities in Children — Attention/Executive Functions,
Learning, and Visual-Spatial Skills
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
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United States (cont'd)
Ris et al. (2004)
U.S.
Design: Prospective longitudinal. 195 subjects from
the Cincinnati Lead Study were administered an
extensive and comprehensive neuropsychological
battery at 16-17 years of age. Domains assessed
included Executive Functions, Attention, Memory,
Achievement, Verbal Skills, Visuoconstructional,
and Fine Motor. Factor scores transformed to ranks
derived from a principal components factor analysis
of the neuropsychological test scores were the
primary outcome variables. Extensive measures on
medical and sociodemographic covariates.
Prenatal (maternal) and
serial postnatal blood lead
assessments.
Prenatal blood lead 8.3
(SD 3.7) ug/dL
Blood lead at 2 years 17.4
(SD 8.8) ug/dL
Following covariate adjustment, strongest associations
between lead exposure and performance were observed for
factor scores derived from the Attention component, which
included high loadings on variables from the Conners
Continuous Performance Test. This relationship was
strongest in males. Authors speculate that since the
incidence of Attention Deficit/Hyperactivity Disorder is
greater in males in general, early exposure to lead may
exacerbate a latent potential for such problems.
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Table AX6-2.6. Effects of Lead on Disturbances in Behavior, Mood, and Social Conduct in Children
to
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Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
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United States
Sciarillo et al. (1992)
U.S.
Bellinger etal. (1994)
U.S.
Demo (1990)
U.S.
Design: Cross-sectional. 150 2-5 year-old children
in Baltimore separated into "high" (2 consecutive
blood lead levels >15 ug/dL) and "low" groups.
Mothers filled out the Achenbach Child Behavior
Checklist (CBCL). The Center for Epidemiologic
Studies Depression Scale (CESD) was administered
to mothers as a control measure.
Design: Prospective cohort: 1782 children bom
within a 1-year period at a single Boston hospital
were examined at 8 years of age. Teachers filled out
the Achenbach Child Behavior Profile (ACBP).
Medical and sociodemographic characteristics
assessed by questionnaire and chart review.
Design: Prospective cohort. Survey of 987
Philadelphia African-American youths enrolled in
the Collaborative Perinatal Project. Data available
from birth through 22 years of age. Analysis
considered 100 predictors of violent and chronic
delinquent behavior.
Screening Blood leads at
various times before
assessment.
Blood lead high group 28.6
(SD 9.3) ug/dL, blood
lead low group 11.3
(SD 4.3) ug/dL
Umbilical cord blood lead
Cord blood lead 6.8
(SD 3.1) ug/dL
Tooth (dentin) lead 3.4
(SD2.4)ug/g
Blood lead
Values not provided
When compared to lower exposed group, children in the
high group had a significantly higher CBCL Total
Behavior Problems Score (TBPS) and Internalizing and
Externalizing scores. After adjustment for maternal
depression, blood lead concentrations were still
significantly associated with an increase in the TBPS.
Children in high group were nearly 3 times more likely to
have a TBPS in the clinical range. A significantly higher
percentage of children in the high group scored in the
clinical range for CBCL subscales measuring aggressive
and destructive behavioral tendencies.
Cord blood lead levels were not associated with the
prevalence or nature of behavioral problems reported by
teachers. Tooth lead levels were significantly associated
with ACBP Total Problem Behavior Scores (TPBS).
Statistically significant tooth lead-associated increases in
both Externalizing and Internalizing scores were observed.
Each log unit increase in tooth lead was associated with a
1.5-point increase in T scores for these scales. Authors
caution that residual confounding cannot be ruled out
because of the lack of information on parental
psychopathology or observations of the family
environment. However, these results are in accord with
other studies that social and emotional dysfunction may be
an important expression of elevated lead levels during
early childhood.
Repeat offenders presented consistent features such as low
maternal education, prolonged male-provider
unemployment, frequent moves, and higher lead
intoxication. In male subjects, a history of lead poisoning
was among the most significant predictors of delinquency
and adult criminality.
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Table AX6-2.6 (cont'd). Effects of Lead on Disturbances in Behavior, Mood, and Social Conduct in Children
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Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
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United States (cont'd)
Needleman et al.
(1996)
U.S.
Dietrich et al. (2001)
U.S.
Needleman et al.
(2002)
U.S.
Design: Prospective cohort. 850 boys enrolled in the
Pittsburgh Youth Study were prescreened to assess
delinquent behavioral tendencies. Subjects who
scored in the 30th percentile on the risk score and an
equal number randomly selected from the remainder
form the sample of 530 subjects. Measures of
antisocial behavior were administered at 7 and 11
years of age including the Self Reported Antisocial
Behavior scale (SRA), Self Report of Delinquent
Behavior (SRD), and parents' and teachers' versions
of the Achenbach Child Behavior Profile (CBCL).
Extensive assessment of medical and
sociodemographic covariates.
Design: Prospective longitudinal. 195 subjects from
the Cincinnati Lead Study were examined at 16-17
years of age. Parents were administered a
questionnaire developed specifically for the study
while CLS subjects were given the Self Report of
Delinquent Behavior. Extensive assessment of
medical and sociodemographic covariates.
Design: Case-control. 194 adjudicated delinquents
and 146 non-delinquent controls recruited from high
schools in the City of Pittsburgh and Allegheny
County, PA. Covariate assessments were not
extensive but did include race, parental
sociodemographic factors, and neighborhood crime
rates.
Bone lead by K-XRF
Bone lead (exact
concentrations not reported)
Negative values treated
categorically as 1 and
positive values grouped into
quintiles.
Prenatal (maternal) and
serial postnatal blood lead
assessments.
Prenatal blood lead 8.3
(SD 3.7) ug/dL
Blood lead at 2 years 17.4
(SD 8.8) ug/dL
Bone lead by KXRF
Bone lead Cases 11.0
(SD 32.7 ug/g),
Controls 1.5
(SD 32.1 ug/g)
Following covariate-adjustment, parents of subjects with
higher lead levels in bone reported significantly more
somatic complaints, more delinquent and aggressive
behavior, and higher Internalizing and Externalizing
scores. Teachers reported significant increase in scores on
somatic complaints, anxious/depressed, social problems,
attention problems, delinquent behavior, aggressive
behavior, internalizing and externalizing problems in the
higher bone lead subjects. At 11 years, subject's SRD
scores were also significantly related to bone lead levels.
More high lead subjects had CBCL T scores in the clinical
range for attention, aggression, and delinquency. Authors
conclude that lead exposure is associated with increased
risk for antisocial and delinquent behavior.
Prenatal (maternal) blood lead was significantly associated
with a covariate-adjusted increase in the frequency of
parent-reported delinquent and antisocial acts. Prenatal
and measures of postnatal lead exposure were significantly
associated with self-reported delinquent and antisocial
behaviors. Authors concluded that lead might play a
measurable role in the development of behavioral problems
in inner-city children independent of other important social
and biomedical cofactors.
Cases had significantly higher average concentrations of
lead in tibia than controls. Following covariate adjustment,
adjudicated delinquents were 4 times more likely to have
bone lead concentration >25 ug/g then controls. Bone lead
level was the second strongest factor in the logistic
regression models, exceeded only by race. In models
stratified by race, bone lead was exceeded as a risk factor
only by single parent status. Authors conclude that
elevated body lead burdens are associated with increased
risk for adjudicated delinquency.
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Table AX6-2.6 (cont'd). Effects of Lead on Disturbances in Behavior, Mood, and Social Conduct in Children
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Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
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Europe
Wasserman et al.
(1994)
Yugoslavia
Design: Prospective longitudinal. Birth cohort of
approximately 300-400 infants followed since birth
residing in two towns in Kosovo, Yugoslavia, one
group near a longstanding lead smelter and battery
manufacturing facility and another in a relatively
unexposed location 25 miles away. 379 children at
3 years of age were examined. Parents were
interviewed with the Achenbach Child Behavior
Checklist (CBCL). Extensive assessment of medical
and sociodemographic covariates.
Maternal prenatal, umbilical
cord and serial postnatal
blood lead
Maternal blood lead in:
exposed area 19.9 (SD 7.7)
ug/dL, unexposed area 5.6
(SD 2.0) ug/dL
Umbilical cord blood lead
in: exposed area 22.2 (SD
8.1) ug/dL, unexposed area
5.5 (SD 3.3) ug/dL.
Blood lead at 2 years in:
exposed area 35.4 ug/dL,
unexposed area 8.5 ug/dL.
Following covariate adjustment, concurrent blood lead levels
were associated with increased Destructive Behaviors on the
CBCL subscale, although the variance accounted for by lead
was small compared to sociodemographic factors. As blood
lead increased from 10 to 20 ug/dL, subscale scores
increased by 0.5 points. The authors conclude that while
statistically significant, the contribution of lead to social
behavioral problems in this cohort was small compared to the
effects of correlated social factors.
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Australia
Burns etal. (1999)
Australia
Fergusson et al. (1993)
New Zealand
Design: Prospective longitudinal. 322 subjects
residing in and near Port Pirie, Australia and
followed since birth were re-evaluated at 11-13 years
of age. Parents completed the Achenbach Child
Behavior Checklist. Extensive assessment of
medical and sociodemographic characteristics.
Design: Prospective cohort. 690-891 children ages
12 and 13 years from the Christchurch Child and
Health Study, New Zealand were examined.
Mothers and teachers were asked to respond to a
series of items derived from the Rutter and Conners
parental and teacher questionnaires. Extensive
assessment of sociodemographic and medical
covariates.
Maternal prenatal, umbilical
cord and serial postnatal
blood lead
Antenatal average blood
lead 10.1(SD 3.9) ug/dL
Umbilical cord blood lead
9.4 (SD 3.9) ug/dL
Blood lead at 2 years
geometric mean 21.3
(SD 1.2) ug/dL
Tooth (dentine) lead
Tooth lead
(range 3-12 ug/g)
After adjustment for covariates, regression models revealed
that for an increase in average lifetime blood lead
concentrations from 10 to 30 ug/dL, the Externalizing
behavior problem T score increased by 3.5 points in boys
(95% CI: 1.6, 5.4), but only 1.8 points (95% CI: -0.1, 11.1)
in girls. Internalizing behavior problems were predicted to
rise by 2.1 points (95% CI: 0.0, 4.2) in girls by only 0.8
(95% CI: - 0.9, 2.4) in boys. Authors concluded that lead
exposure is associated with an increase in externalizing
(undercontrolled) behaviors in boys.
Statistically significant dose-effect relationships were
observed between tooth lead levels and the
inattention/restlessness variable at each age. Authors
conclude that this evidence is consistent with the view that
mildly elevated lead levels are associated with small but long
term deficits in attentional behaviors.
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Table AX6-2.7. Effects of Lead on Sensory Acuities in Children
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Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
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United States
Schwartz and Otto
(1991)
U.S.
Design: Cross-sectional. 3545 subjects 6-19 years old who
participated in the Hispanic Health and Nutrition Examination
Survey. Pure tone audiometric evaluations were performed at
500 Hz, 2000 Hz, and 4000 Hz. Extensive measures on
medical and sociodemographic covariates.
Blood lead at the time of
testing.
Blood lead 50th
percentile
8 ng/dL
Dietrich etal. (1992)
U.S.
Design: Prospective/longitudinal. 215 subjects drawn from
the Cincinnati Lead Study at the age of 5 years. Children were
administered the SCAN-a standardized test of central auditory
processing. Extensive measurement of medical and
sociodemographic covariates
Prenatal (maternal) and
serial postnatal blood lead
assessments.
Prenatal blood lead 8.3
(SD 3.7) ng/dL
Blood lead at 2 years
17.4 (SD 8.8) ng/dL
Following covariate adjustment, higher blood lead
concentrations were associated with an increased risk
of hearing thresholds that were elevated above the
standard reference level at all four frequencies.
Blood lead was also associated higher hearing
threshold when treated as a continuous outcome.
These relationships extended to blood lead levels
below 10 ng/dL. An increase in blood lead from 6 to
18 ng/dL was associated with a 2-dB loss at all
frequencies. Authors conclude that HHANES results
those reported earlier for NHANES-II.
Higher prenatal (maternal), neonatal and postnatal
blood lead concentrations were associated with more
incorrect identification of common monosyllabic
words presented under conditions of muffling.
Following covariate adjustment, average childhood
blood lead level remained significantly associated
with impaired performance on the SCAN subtest.
Authors conclude that lead-related deficits in hearing
and auditory processing may be one plausible
mechanism by which an increased lead burden might
impede a child's learning.
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Europe
Osmanetal. (1999)
Poland
Design: Cross-sectional. 155 children 4-14 year-old living in Blood lead at the time
an industrial region of Poland. Pure tone audiometric
evaluations were performed at 500 Hz, 1000 Hz, 2000 Hz,
4000 Hz, 6000Hz, and 8000 Hz. Basic data on medical
history, limited information on sociodemographic covariates
such as family structure and income.
of testing
Blood lead median
7.2 ng/dL
(range 1.9-28 ng/dL)
Higher blood lead concentrations were significantly
associated with increased hearing thresholds at all
frequencies studied. This relationship remained
significant when analyses were limited to subjects
with blood lead levels below 10 ng/dL. Authors
conclude that auditory function in children is
impaired at blood lead concentrations below
10
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Table AX6-2.8. Effects of Lead on Neuromotor Function in Children
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Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
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Dietrich et al. (1993b);
Bhattacharya et al.
(1995);Risetal.
(2004)
U.S.
Design: Prospective longitudinal. Relationship
between lead exposure and neuromotor function has
been examined in several studies on the Cincinnati
Lead Study Cohort from 6 to 17 years of age.
At 6 years of age 245 subjects were administered the
Bruininks-Oseretsky Test of Motor Proficiency
(BOTMP); at 6-10 years of age subjects were
assessed for postural instability using a
microprocessor-based strain gauge platform system
and at 16-17 years of age the fine-motor skills of
study subjects were assessed with the grooved
pegboard and linger tapping tasks (part of a
comprehensive neuropsychological battery).
Extensive measurement of medical and
sociodemographic factors.
Prenatal (maternal) and
serial postnatal blood lead
assessments.
Prenatal blood lead 8.3
(SD 3.7) ug/dL
Blood lead at 2 years 17.4
(SD 8.8) ug/dL
Following covariate adjustment, postnatal lead exposure was
significantly associated with poorer scores on BOTMP
measures of bilateral coordination, visual-motor control,
upper-limb speed and dexterity and the Fine Motor
Composite score. Low-level neonatal blood lead
concentrations were also significantly associated with poorer
scores on the aforementioned subtests, as well as measures
of visual-motor control. Postnatal lead exposure was
significantly associated with greater postural instability in 6-
10 year-old subjects and poorer fine-motor coordination
when examined at 16-17 years.
Authors conclude that effects of early lead exposure extend
into a number of dimensions of neuromotor development.
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Europe
Wasserman et al.
(2000a)
Yugoslavia
Design: Prospective longitudinal. Birth cohort of
approximately 300-400 infants followed since birth
residing in two towns in Kosovo, Yugoslavia, one
group near a longstanding lead smelter and battery
manufacturing facility and another in a relatively
unexposed location 25 miles away. 283 children at
age 54 months were administered the Beery
Developmental Test of Visual-Motor Integration
(VMI) and the Bruininks-Oseretsky Test of Motor
Proficiency (BOTMP). Extensive measurement of
medical and sociodemographic factors.
Maternal prenatal, umbilical
cord and serial postnatal
blood lead
Maternal blood lead in:
exposed area 19.9 (SD 7.7)
ug/dL, unexposed area
5.6 (SD 2.0) ug/dL
Umbilical cord blood lead
in: exposed area 22.2
(SD 8.1) ug/dL, unexposed
area 5.5
(SD 3.3) ug/dL.
Blood lead at 2 years in:
exposed area 35.4 ug/dL,
unexposed area 8.5 ug/dL.
Following covariate-adjustment, the log average of serial
blood lead assessments to 54 months was associated with
lower Fine Motor Composite and VMI scores. Lead
exposure was unrelated to gross motor performance. With
covariate adjustment, an increase in average blood lead from
10 to 20 ug/dL was associated with a loss of 0.62 and 0.42
points respectively, in Fine Motor Composite and VMI.
Authors noted that other factors such as indicators of greater
stimulation in the home make a larger contribution to motor
development than lead.
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Table AX6-2.9. Effects of Lead on Direct Measures of Brain Anatomical Development and Activity in Children
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Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
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United States
Trope etal. (1998)
U.S.
Trope etal. (2001) U.S.
Cecil et al. (2005)
U.S.
Design: Case-control. One 10 year-old subject
with history of lead poisoning and unexposed 9
year-old cousin. Magnetic Resonance Imaging
(MRI) and Magnetic Resonance Spectroscopy
(MRS) were used to assess differences in cortical
structures and evidence of neuronal loss. This was
the first study to attempt to determine in vivo
structural and/or metabolic differences in the brain
of a child exposed to lead compared with a healthy
control.
Design: Case-control. 16 subjects with a history of
elevated blood lead levels before 5 years of age and
5 age-matched siblings or cousins were evaluated.
Average age at time of evaluation was 8 years.
Magnetic Resonance Imaging (MRI) and Magnetic
Resonance Spectroscopy (MRS) were used to
assess differences in cortical structures and
evidence of neuronal loss.
Design: Prospective/longitudinal. 48 young adults
ages 20 to 23 years were re-examined. Functional
MRI (fMRI) was used to examine the influence of
childhood lead exposure on language function.
Subjects performed a verb generation/finger-
tapping paradigm. Extensive measurement of
medical and sociodemographic covariates
Blood lead
lead poisoned case
51 ug/dL
at 38 mos.
Unexposed control
not reported.
Blood lead range in lead-
exposed 23 to 65 ug/dL
Controls <10 ug/dL
Blood lead
Average childhood blood
lead 13.9 (SD 6.6 ug/dL
(range 4.8-31.1 ug/dL)
Both children presented with normal volumetric MRI. MRS
revealed a significant alteration in brain metabolites, with a
reduction in N-acetylaspartate:creatine ratio for both gray
and white matter compared to the subject's cousin. Authors
conclude that results suggest neuronal loss related to earlier
lead exposure.
All children had normal MRI examinations, but lead-
exposed subjects exhibited a significant reduction in
N-acetylaspartate:creatine and pohosphocreatine ratios in
frontal gray matter compared to controls. Authors conclude
that lead has an effect on brain metabolites in cortical gray
matter suggestive of neuronal loss.
Higher average childhood blood lead levels was significantly
associated with reduced activation in Broca's area in the left
hemisphere and increased activation in the right temporal
lobe, the homologue of Wemicke'a area in the left
hemisphere. Authors conclude that elevated childhood lead
exposure strongly influences neural substrates of semantic
language function on normal language areas with
concomitant recruitment of contra-lateral regions resulting in
a striking dose-dependent atypical organization of language
function.
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Table AX6-2.9 (cont'd). Effects of Lead on Direct Measures of Brain Anatomical Development and Activity in Children
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Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
Latin America
Rothenberg et al. (2000)
Mexico
Design: Prospective/longitudinal. 113 5-7 year-
old children from the Mexico City Prospective
Study were re-examined. Brain stem auditory
evoked potentials were recorded to assess the
impact of prenatal and postnatal lead exposure on
development of auditory pathways. Results
adjusted for gender and head circumference.
Blood lead
Prenatal (20 wks) 8.1
(SD 4.1) ug/dL
Cord 8.7 (SD 4.3) ug/dL
Postnatal 18 mos. 10.8
(SD 5.2) ug/dL
Prenatal blood lead at 20 weeks was associated with
decreased interpeak intervals. After fitting a nonlinear
model to these data, I-V and III-V interpeak intervals
decreased as blood lead rose from 1 to 8 ug/dL and
increased as blood lead rose from 8 to 30 ug/dL. Increased
blood lead at 12 and 48 months was related to decreased
conduction intervals for I-V and II-V across the entire blood
lead range suggesting pathway length effects.
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Meng et al. (2005)
China
Design: Case-control. 6 subjects with blood lead
concentrations >27 ug/dL and 6 controls with
blood lead concentrations <10 ug/dL were
evaluated with Magnetic Resonance Imaging
(MRI) and Magnetic Resonance Spectroscopy to
evaluate structural abnormalities and differences in
N-acetylaspartate, creatine, and choline in frontal
lobes and hippocampus of cases and controls.
Blood lead
Blood lead cases 37.7
(SD 5.7) ug/dL
Blood lead controls 5.4
(SD 1.5) ug/dL
All children presented with normal MRI. Peak values of N-
acetylaspartate, choline, and creatine in all four brain regions
were reduced in lead exposed children relative to controls.
Authors conclude that reduced brain N-acetylaspartate in
cases may be related to decreased neuronal density or loss.
Reduced choline signal may indicate decreased cell
membrane turnover or myelin alterations while lower
creatine may indicate reduced neuronal cell viability.
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Table AX6-2.10. Effects of Lead on Reversibility of Lead-Related Deficits in Children
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Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
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United States
Ruff etal. (1993)
U.S.
Roganetal. (2001);
Dietrich et al. (2004)
U.S.
Design: Intervention study, non-randomized.
126 children with complete data age 13 to 87
months and with blood lead levels between 25 and
55 ug/dL were given chelation with ETDA and/or
therapeutic iron where indicated. At baseline and
follow-up, patients were evaluated with the Bayley
Scales of Infant Development, Mental
Development Index, or Stanford Binet Scales of
Intelligence depending upon age.
Design: Double blind, placebo-controlled
randomized clinical trial. The Treatment of Lead-
Exposed Children (TLC) clinical trial of 780
children in 4 centers was designed to determine if
children with moderately elevated blood lead
concentrations given succimer would have better
neuropsychological outcomes than children given
placebo. Children between 12 and 33 months of
age were evaluated 3 years following treatments
and again at 7 and 7.5 years of age. A wide range
of neurological, neuropsychological, and
behavioral tests was administered. Assessment of
potentially confounding factors included
sociodemographics and parental IQ.
Blood lead at time of
treatment 31.2 (SD 6.5)
ug/dL.
Blood lead
Range 20-44 ug/dL
Baseline blood lead 26
(SD 26.5) ug/dL in both
drug and placebo groups.
Without respect to treatment regimen, changes in
performance on cognitive measures after 6 months were
significantly related to changes in blood lead levels after
control for confounding factors. Standardized scores on tests
increased 1 point for every 3 ug/dL decrement in blood lead.
Succimer was effective in lowering blood lead levels in
subjects on active drug during the first 6 months of the trial.
However, after 1 year differences in the blood lead levels of
succimer and placebo groups had virtually disappears.
3 years following treatment, no statistically significant
differences between active drug and placebo groups were
observed for IQ or other more focused neuropsychological
and behavioral measures. When evaluated at 7 and 7.5 years
of age, TLC could demonstrate no benefits of earlier
treatment on an extensive battery of cognitive, neurological,
behavioral and neuromotor endpoints. Authors conclude that
the TLC regimen of chelation therapy is not associated with
neurodevelopmental benefits in children with blood lead
levels between 20 and 44 ug/dL and that these results
emphasize the importance of taking environmental measures
to prevent exposure to lead in light of the apparent
irreversibility of lead-associated neurodevelopmental
deficits.
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Table AX6-2.10 (cont'd). Effects of Lead on Reversibility of Lead-Related Deficits in Children
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Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
United States (cont'd)
Liu et al. (2002)
U.S.
T.atin Amprira
Design: Prospective longitudinal clinical trial.
Data from the Treatment of Lead-Exposed Children
(TLC) used to examine prospective relationships
between falling blood lead levels and changes in
cognitive functioning. 741 children recruited
between 13 and 33 months of age were assessed at
baseline and 6 months later with the Bayley Mental
Development Index (MDI) and 36 months post-
randomization with the Wechsler Preschool and
Primary Scales of Intelligence-Revised to
obtain IQ.
Blood lead
Baseline blood lead 26.2
(SD 5.1) ug/dL
36 months post-
randomization blood lead
12.2 (SD 5.2) ug/dL
TLC found no overall effect of changing blood lead level on
change in cognitive test scores from baseline to 6 months.
Slope estimated to be 0.0 points per 10 ug/dL change in
blood lead. From baseline to 36 months and 6 months to
36 months, falling blood lead levels were significantly
associated with increased cognitive test scores, but only
because of an association in the placebo group. Authors
conclude that because improvements were not observed in
all children, the data do not provide support that lead-
induced cognitive impairments are reversible. Although the
possible neurotoxicity of succimer cannot be ruled out.
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Kordasetal. (2005);
Rico et al. (2006)
Torreon, Mexico
Australia
Design: Double-blind, placebo-controlled
nutritional supplementation clinical trial conducted
among 602 first grade children ages 6-8 years in
Torreon, Mexico. Subjects received iron, zinc,
both or placebo for 6 months. Parents and teachers
filled out the Conners Rating Scales at baseline and
follow-up six months following the end of
supplementation to index behavioral changes
following therapy. In addition, 11 cognitive tests
of memory, attention, visual-spatial abilities, and
learning were administered, including WISCR-M at
baseline and follow-up 6 months later.
Blood lead
Baseline blood lead 11.5
(SD 6.1) ug/dL
No significant effects of treatment on behavior or cognition
could be detected with any consistency. Authors conclude
that this regimen of supplementation does not result in
improvements in ratings of behavior or cognitive
performance.
Tongetal. (1998)
Australia
Design: Prospective longitudinal. 375 children
from the Port Pirie Prospective Study were
followed from birth to the age of 11-13 years.
Bayley Mental Development Index (MDI) at
2 years, the McCarthy Scales General Cognitive
Index (GCI) and IQs from the Wechsler
Intelligence Scale served as the primary indicators
of intellectual status. The purpose of the study was
to assess the reversibility of lead effects on
cognition in relationship to declines in blood lead
over time.
Postnatal Blood lead
Average Blood lead at
2 years 21.2 ug/dL declining
to 7.9 ug/dL at 11-13 years.
Although blood lead levels declined substantially, covariate
adjusted scores on standardized measures of intellectual
attainment administered at 2, 4, 7, and 11-13 years of age
were unrelated to declining body burden. Authors conclude
that effects of early exposure to lead during childhood are
not reversed by a subsequent decline in blood lead
concentration.
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ANNEX TABLES AX6-3
May 2006 AX6-28 DRAFT-DO NOT QUOTE OR CITE
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Table AX6-3.1. Neurobehavioral Effects Associated with Environmental Lead Exposure in Adults
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Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
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United States
Krieg et al. (2005)
1988-1994
Muldoonetal. (1996)
4,937 adults aged 20-59years from
NHANES III completed three
neurobehavioral tests. Regression analyses
of neurobehaqvioral test and log of blood
lead concentration adjusted for sex, age
education, family income, race/ethnicity,
computer or video game familiarity, alcohol
use, test language, and survey phase.
325 women from rural location (mean age
71) and 205 women from a city location
(mean age 69) participants in the Study for
Osteoporotic Fractures had the association
of nonoccupational lead exposure and
cognitive function examined. Logistic
regression determined effect of blood lead
on neuropsychological performance.
Mean blood lead 3.3 ug/dL
Range 0.7 to 41.7 ug/dL
Rural group
Blood lead 5 ug/dL
Urban group
Blood lead 5 ug/dL
No statistically significant relationship between blood lead
concentration and mean simple reaction time, symbol-digit
substitution latency and errors and serial digit learning trials to
criterion and total score after adjustments for covariates.
Groups were significantly different with the urban group more educated
and smoked and drank more. Performance in each group stratified by
exposure into three groups (low <4 ug/dL, medium 4-7 ug/dL, high
>7 ug/dL rural and >8 ug/dL) - no significant associations were present
in the urban group but the rural group had significantly poorer
performance with increasing blood lead for Trails B (OR = 2.6, 95%
CI: 1.04, 6.49), Digit Symbol (OR 3.73, 95% CI: 1.57, 8.84),and
Reaction Time in the lower (OR 2.84, 95% CI: 1.19, 6.74) and upper
extremities (OR 2.43, 95% CI: 1.01,5.83). The fact that marked
differences exist between the low lead groups for rural and urban (the
lowest 15th percentile) suggests the differences between the two groups
are unrelated to lead. Response time for reaction time across lead
groups increased for the rural group and decreased or remained the
same for the urban group. As response time is sensitive to lead effect,
this raises question whether factors not measured accounted for
difference. Namely MMSE for the whole population was 25 (15-26)
with poorer performance in the rural group. The clinical cutoff score
for MMSE is 24 suggesting the presence of clinical cognitive disorders.
Even though this is a simple neuropsychological battery up to 9 were
unable to perform some of the tests including 3 on the MMSE.
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Table AX6-3.1 (cont'd). Neurobehavioral Effects Associated with Environmental Lead Exposure in Adults
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
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United States (cont'd)
Paytonetal. (1998)
Rhodes et al. (2003)
Wright et al. (2003)
141healthy men in VA normal aging study
evaluated every 3 to 5 years with cognitive
battery and blood lead and once a
measurement of patella and tibia bone lead.
Statistics are confusing as it is not clear
when ANCOVA is used and how the
groups are created.
526 participants with mean age 67 years,
47% had education level of high school or
less. Mood symptoms evaluated with Brief
Symptom Inventory (BSI). Use of logistic
regression adjusting for covariates
examined association of BSI scales and
blood lead and bone lead levels.
736 healthy men (mean age 68) in
Normative Aging Study examined every 3
to 5 years were administered the Mini-
Mental State Exam (MMSE). Linear
regression examined relationship of
MMSE and blood lead, Patella and Tibia
bone lead measurements after adjusting for
covariates.
Mean blood lead 6 ug/dL,
patella bone lead 32 and
tibia bone lead 23 ug/g
bone mineral
Mean blood lead 6 ug/dL
Mean tibia Pb 22 ug/g
Mean patella Pb 32 ug/g
Mean blood lead 5 ug/dL,
patella bone lead 30 and
tibia bone lead 22 ug/g
bone mineral
Regressions adjusted for age and education found significant relationship of
blood lead with Pattern Comparison (perceptual speed), Vocabulary, Word
List Memory, Constructional Praxis, Boston Naming Test, and Verbal
Fluency Test. Only for Constructional Praxis were bone lead and blood lead
significantly associated. Mechanism most sensitive to low levels lead
exposure believed to be response speed. It is unusual that Vocabulary, a test
resistant to neurotoxic insult is significantly associated with blood lead.
This may be related to the significant negative correlation of bone lead with
education, a similar trend is present for blood lead. It is not clear how
multiple comparisons were handled.
BSI found mood symptoms for anxiety and depression were potentially
associated with bone lead levels. However education was inversely related
to bone lead and high school graduates had significantly higher odds of
Global Severity Index and Positive Symptom Total. BSI appears to be
detecting general stress related to socioeconomic status.
Mean MMSE score 27. Relation of MMSE scores <24 (n = 41) and blood
lead by logistic regression found OR 1.21 (95% CI: 1.07, 1.36) and for
patella lead OR 1.21 (95% CI: 1.00, 1.03) and tibia lead OR 1.02 (95% CI:
1.00, 1.04). Risk of MMSE <24 when comparing the lowest and highest
quartiles of patella lead was 2.1 (95% CI: 1.1, 4.1), for tibia lead was 2.2
(95% CI: l.l,3.8)andbloodleadwas3.4(95%CI: 1.6,7.2). Interaction
between patella lead and age, and blood lead and age in predicting MMSE
found steeper decrease in MMSE score relative to age in the higher quartiles
of patella lead and blood lead.
MMSE very sensitive to years of education below 8 years. In this study 213
subjects had less than high school education. If the community dwelling
population had older individuals with less education living in areas with
higher past pollution the confounding may be impossible to sort out.
Initially at beginning of NAS subjects were eliminated with chronic medical
problems or blood pressure > 140/90. It is not addressed how the
development of medical conditions during the duration of the study are
handled.
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Table AX6-3.1 (cont'd). Neurobehavioral Effects Associated with Environmental Lead Exposure in Adults
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
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United States (cont'd)
Weisskopf et al.
(2004b)
466 men, mean age 70 years, in the VA
Normative Aging Study had 2 MMSE tests
3.5 years apart.
Mean blood lead 4 ug/dL,
patella bone lead 23 and
tibia bone lead 19 ug/g
bone mineral
Baseline mean MMSE score was 27 and mean change in MMSE
score over 3.5 years was 0.3. Change in MMSE associated with one
interquartile range increment for bone lead and blood lead found
relationship between patella lead and change in MMSE was unstable
when patella lead is >90 ug/g bone mineral. Examination of patella
lead below this level found a greater inverse association with MMSE
at lower lead concentrations ((3 = -0.25, 95% CI: -0.45, -0.05).
A similar but weaker association existed for tibia lead when values
>67 ug/g bone mineral were removed ((3 = -0.19, 95% CI: -0.39,
0.02). There was no association of MMSE change and blood lead
(P = -0.01, 95% CI: -0.13,0.11). There was no interaction of age
and bone lead. These are very high bone lead levels for
environmental exposure. The biological plausibility of change in the
MMSE over 3.5 years would have been reinforced if the change by
functional domain in the MMSE was provided.
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Europe
Nordberg et al. (2000)
Sweden
762 participants, mean age 88 years, in a
study of aging and dementia examined
MMSE. Used blood lead as dependent and
examined contribution of covariates and
MMSE.
Mean blood lead 3.7 ug/dL
Mean MMSE 25 found no relation of blood lead and MMSE. In this
population was fairly homogenous, all elderly Swedes, and the
likelihood of prior exposure to elevated lead levels was low.
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Table AX6-3.2. Symptoms Associated with Occupational Lead Exposure in Adults
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
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Canada
Lindgren et al. (1999)
Holnessetal. (1988)
Smelter workers (n = 467) with a mean age
of 40 years completed the Profile of Mood
Scale. Factor structure of POMS validated in
this occupational population. Regression
analysis determined association with lead
exposure.
47 demolition workers with acute lead
intoxication - Phase 1-were followed with
blood lead and symptoms during engineering
modifications to control exposure -Phases
2-4. Workers stratified by blood lead and
symptom frequency was analyzed.
Mean blood lead 28
(8.5,4-62)ng/dL
Mean IBL 711
(415.5, 1-1537) ng-yr/dL.
Phase I- Mean blood lead
59 ug/dL SDN/A
Phase 2-Mean blood lead
30 ug/dL SD N/A
Phase 3-Mean blood lead
19 ug/dL SDN/A
Phase 4-Mean blood lead
17 ug/dL SDN/A
Factor analysis found one factor labeled "general distress" composed
of POMS subscales anger, confusion, depression, fatigue and tension
and a second factor labeled 'psychological adjustment'. IBL was
significantly associated with 'general distress' after adjustment for the
covariates (P = 0.28 [SE 1.51 x 10~4] p = 0.01) while there was no
relation with blood lead. The factor structure of POMS originally
validated in a clinical population had six mood subscales however the
factor structure in this occupational population was found to have
only two subscales.
Below blood lead <50 ug/dL percentage of workers reporting
symptoms was fatigue-25, headache-14 dizzy-9, sleep-8, abdominal
cramps-8, muscle ache-8, paresthesiae-8, appetite-7, constipation-6,
and weakness-6. All symptoms were significantly lower except for
paresthesiae when compared to group with blood lead >70 ug/dL.
Of interest, at beginning of Phase 4 when mean blood lead was
13 ug/dL, no symptoms were reported. At the end of Phase 4, mean
blood lead was 17 ug/dL and one worker complained of fatigue.
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Europe
Lucchini et al. (2000)
Italy
66 workers in lead manufacturing, mean age
40 (8.7) years and 86 controls mean age 43
(8.8) years were administered a questionnaire
with neuropsychological (14 items), sensory-
motor (3 items), memory (4 items) and
extrapyramidal (8 items), 10 Parkinson
symptoms and the Mood Scale. Group
comparisons and linear regression examined
relationship of symptoms and lead exposure.
Mean blood lead 27 (11.0,
6-61) ug/dL
Mean TWA 32 (14.1, 6-61)
ug/dL
Mean IBL 410 (360.8,
8-1315)ug-yr/dL.
Controls-mean blood lead 8
(4.5, 2-21) ug/dL
Lead exposed worker reported confusion, somnolence, abnormal
fatigue, irritability, and muscular pain more frequently (p < 0.04).
There were no group differences for the parkinsonism symptoms or
Mood Scale. Linear regression combing exposed and control group
found neurological symptoms significantly associated with blood lead
r = 0.22, p = 0.006). Neuropsychological symptoms were
significantly higher in the High-IBL compared to the Low-IBL group.
The estimated threshold for a significant increase (prevalence of 5%)
of a high score for neurological symptoms was at a blood lead of
12 ug/dL.
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Table AX6-3.2 (cont'd). Symptoms Associated with Occupational Lead Exposure in Adults
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Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
Latin America
Maizlish et al. (1995)
Venezuela
43 workers from a lead smelter, mean age 34
(9) years and 47 nonexposed workers, mean
age35(ll) years completed the profile of
mood states (POMS) questionnaire and a
questionnaire of symptoms of the central and
peripheral nervous system, and
gastrointestinal. Prevalence ratios used to
examine symptoms and lead. ANCOVA and
linear regression adjusting for potential
confounders examined relationship of lead
exposure and POMS.
Mean blood lead 43 (12.1)
ug/dL
Mean peak blood lead 60
(20.3) ug/dL
Mean TWA 48 (12.1)
ug/dL
Controls
mean blood lead 15 (6)
ug/dL
mean peak blood lead 15
(6) ug/dL
mean TWA 15 (6) ug/dL
Significantly increased relative risks found for difficulty
concentrating (RR 1.8 [95% CI: 1.0-3.1]), often being angry or upset
without reason (RR 2.2 [95% CI: 1.2,4.1]), feeling abnormally tired
(RR 2.2 [95% CI: 0.9, 5.3]) and joint pain (RR 1.8, [95% CI: 1.0,
3.3]). The six subscales of the POMS were not significantly different
between the exposed and control groups. However dose-related
analysis found significantly poorer scores for tension-anxiety and
blood lead (p = 0.009), hostility and blood lead (p = 0.01) and TWA
(p = 0.04), and depression and blood lead (p = 0.003) and peak lead
(p = 0.003) and TWA (p = 0.004).
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Asia
Schwartz et al.
(200 la)
Korea
Lee et al. (2000)
Korea
803 lead-exposed Korean workers, mean age
40 years completed the Center for
Epidemiologic Studies Depression Scale.
Linear regression examined for association
of CES-D and lead biomarkers after
adjusting for the covariates.
95 Korean lead exposed workers, mean age
43 years, completed questionnaire of lead-
related symptoms present over last three
months. Relationship between symptom
score and measures of lead exposure
assessed by linear regression. Logistic
regression use to model presence or absence
of symptoms for gastrointestinal,
neuromuscular, and general.
Mean blood lead 32 (15.0)
ug/dL
Mean tibia lead 37 (40.3)
ug/g bone mineral
DMSA-chelatable lead
Mean 289 (167.7) ug
ZPP 108 (60.6) ug/dL
MeanALAU3(2.8)mg/l
Mean blood lead 45 (9.3)
ug/dL
After adjustment for age, gender and education significant
associations found for CES-D and tibia lead ((3 = 0.0021 [SE 0.0008];
p < 0.01) but not with blood lead. This occupational lead-exposed
populations had higher past lead exposure compared to the current
mean blood lead of 32 ug/dL.
Workers with DMSA -chelatable lead above the median of 261 ug
were 6.2 (95% CI: 2.4, 17.8) times more likely to have tingling or
numbness in their extremities, 3.3 (95% CI: 1.2, 10.5) times more
likely to experience muscle pain and 3.2 (95% CI: 1.3, 7.9) times
more likely to feel irritable. The workers with higher chelatable lead
were 7.8 (95% CI: 2.8,24.5) times more likely to experience
neuromuscular symptoms compared to workers with lower chelatable
lead. In this study ZPP predicted weakness of ankle and wrist (OR
2.9[95%CI: 1.1, 8.1]) and fatigue (OR 2.9 [95% CI: 1.1, 8.7]) while
ALAU predicted inability to sleep (OR 5.4 [95% CI: 1.2, 33.2]) and
blood lead was not significantly associated with any symptoms.
A measure of lead in bioavailable storage pools was the strongest
predictor of symptoms particularly neuromuscular.
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Table AX6-3.2 (cont'd). Symptoms Associated with Occupational Lead Exposure in Adults
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Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
Asia (cont'd)
Niu et al. (2000)
China
44 lead-exposed workers (17 men, 27
women) from lead printing houses, mean age
35 (4.9) and education 9.3 (no SD) years and
34 controls (19 men and 15 women), mean
age 33 (7.4) years and education 9.5 (no SD)
years completed the profile of mood state as
part of the NCTB. ANCOVA controlling for
age, sex and education examined group
differences and linear regression for dose-
response relationship.
Mean blood lead 29 (26.5)
(8 workers blood lead
exceeded 50 ng/dL)
Controls
Mean blood lead 13 (9.9)
POMS subscales for confusion (F = 3.02, p < 0.01), fatigue (F = 3.61,
p < 0.01), and tension (F = 2.82, p < 0.01) were significantly elevated
in the lead exposed group. Regression analyses found a dose response
(data not shown).
(1 control blood lead
exceeded 50 |ig/dL)
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Table AX6-3.3. Neurobehavioral Effects Associated with Occupational Lead Exposure in Adults
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Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
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United States
Fiedler et al. (2003)
New Jersey
Balbus-Kornfeld et al.
(1995)
40 workers with lead exposure, mean age
48 (9.5) years completed a neurobehavioral
battery and was compared to 45
lead/solvent workers, mean age 47 (10.2),
39 solvent exposed workers, mean age 43
(9.4), and 33 controls, mean age 44 (10.2).
Group differences and dose-effect
relationships were assessed after adjusting
for potential confounding.
Reviewed 21 studies from 28 publications;
number of subjects ranged from 9-708.
Mean blood lead ug/dL
Mean bone lead ppm ug/g
(dw) Lead workers
14 (11.7)/2.7(0.7)
Lead/Solvent workers
12(11.6)72.8(0.6)
Solvent workers
5(4.1)7-1.8(1.8)
Controls
4(1.4)7-1.1(1.6)
Mean blood lead in most
exposed group 28-68
ug/dL. Only 5 studies used
a measure of cumulative
exposure or absorption of
Pb, 2 studies used duration
of exposure.
Of nineteen outcomes, significant differences found on the California
verbal learning test (CVLT) (p = 0.05) and positive symptom distress
index on the Symptom checklist-90-R. On the CVLT the controls
performed significantly better on trials 2 and 3 demonstrating
efficiency of verbal learning. Symbol digit substitution (SDS)
approached significance (p = 0.09) with lead and lead/solvent group
slower on latency of response but not accuracy. Bone lead was a
significant predictor of latency of response on SDS, total errors on
paced auditory serial addition task and simple reaction time non-
preferred hand. Bone lead and SRT, preferred hand approached
significance. This is a confusing study design as bone lead is used as
a predictor in workers both with and without occupational lead
exposure.
Dexterity (17/21 studies) and executive or psychomotor 11/21 studies
were the functional domains most commonly associated with lead.
Age not adequately controlled in most studies, usually matching
means or medians. Intellectual abilities prior to exposure usually
adjusted for with education however Vocabulary, a measure of overall
intellectual ability still different between the groups. The conclusion
reached that evidence of effects from cumulative exposure or
absorption of lead was inadequate.
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Canada
Lindgren et al. (1996)
467 Canadian former and current, French
and English speaking lead smelter workers,
mean age 43 (11.0) years and education
10 (3.2) years were administered a
neuropsychological battery in English or
French. Data analyses used MANCOVA
adjusting for age, education, measure of
depressive symptoms and self reported
alcohol use.
Mean years employment 18
(7.4)
Mean blood lead 28 (8.4)
ug/dL
Mean TWA 40 (4-66)
ug/dL
Mean IBL 765 (1-1626)
ug-yr/dL
Fourteen neuropsychological variables examined by MANCOVA
with the grouping variable exposure (high, medium and low) and the
covariates, age, education, CES-D, and alcohol use found no exposure
term significant until years of employment, a suppressor term, was
added as a covariate. IBL exposure groups differed significantly (df
2,417) on digit symbol (F = 3.03, p = 0.05), logical memory (F = 3.29,
P = 0.04), Purdue dominant hand (F = 4.89, p = 0.01), and trails A
(F = 3.89, p = 0.02) and B (F = 3.2, p = 0.04). This study showed a
dose-effect relationship between cumulative lead exposure (IBL) and
neuropsychological performance at a time when there was no
association with current blood lead.
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Table AX6-3.3 (cont'd). Neurobehavioral Effects Associated with Occupational Lead Exposure in Adults
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Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
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ON
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Canada (cont'd)
Bleecker et al. (2002)
Bleecker et al. (2005a)
256 smelter workers from the above
population were currently employed and
took the test battery in English. Their mean
age was 41 (7.9) years, and education 10
(2.8) years. The goal was to determine if
educational achievement as measured by
WRAT-R Reading modified performance on
MMSE. Linear regression assessed the
contribution of age, WRAT-R, education,
alcohol intake, cigarette use, IBL and
IBLxWRAT-R on MMSE performance.
256 smelter workers currently employed and
took the test battery in English. Their mean
age was 41 (7.9) years, and education
10 (2.8) years. The purpose was to
determine whether components of verbal
memory as measured on the Rey Auditory
Verbal Learning Test (RAVLT) were
differentially affected by lead exposure.
Linear regression and ANCOVA assessed
the relationship of lead and components of
verbal learning and memory.
Mean blood lead 28 (8. 8)
Mean IBL 725 (434) ng-
yr/dL
Mean blood lead 28 (8. 8)
lig/dL
Mean TWA 39 (12.3)
Mean IBL 725 (434)
Hg-yr/dL
MMSE had a median (range) score of 29 (19-30). The most common
errors were recall of 3 items (38%), spell world backwards (31%),
repetition of "no ifs ands or buts" (21%) and copy a design to two
intersecting pentagons (16%). WRAT-R reading used as an additional
measure of educational achievement because it was a stronger predictor
of MMSE performance than years of education. The significant
interaction (AR2 = 2%, p = 0.01) explained by a dose-effect between
IBL and MMSE only in the 78 workers with a WRAT-R reading grade
level less than 6. The workers with higher reading grade levels and the
same cumulative lead exposure were able to compensate for the effects
of lead on the MMSE because of increased cognitive reserve.
Outcome variables RAVLT a word list test included measures of
immediate memory span and attention (Trial 1), best learning (Trial V),
incremental learning across the five trials (Total Score), and storage
(Recognition) and retrieval (Delayed Recall) of verbal material. TWA
significantly contributed to the explanation of variance for Trial V
(AR2 = 1.4%, p < 0.03) and Delayed Recall (AR2 = 1.4%, p = 0.03) after
adjusting for age and WRAT-R while IBL did the same with
Recognition (AR2 = 2.0%, p = <0.02) and Delayed Recall (AR2 = 1.1%,
p = 0.06). Workers stratified into 3 group by increasing clinical
memory difficulties-Group 1 had normal encoding, storage and
retrieval; Group2 could encode and store verbal information but had
difficulty with retrieval and Group 3 had abnormal encoding, storage
and retrieval but was still able to learn new verbal information.
ANCOVA adjusting for age and WRAT-R compared lead exposure
across the memory groups. Blood lead showed no difference but TWA
and IBL were significantly higher in Group 3 compared to Group 1
(p < 0.05 for both). Internal strategies used on the RAVLT over the
five trials found that Groups 1 and 2 remembered more words from the
beginning of the list while group 3 remembered more from the end.
At a time when blood lead was not associated with performance,
cumulative lead exposure resulted in poorer storage and retrieval of
previously learned material. Alterations in the ability to organize
materials in long term memory interferes with retrieval efficiency.
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Table AX6-3.3 (cont'd). Neurobehavioral Effects Associated with Occupational Lead Exposure in Adults
Reference, Study
Location, and Period Study Description Lead Measurement Findings, Interpretation
Canada (cont'd)
Bleeckeretal. (1997a)
New Brunswick
1992-1993
Bleeckeretal. (1997b)
New Brunswick
1992-1993
The performance of the 467 current and
retired smelter workers as described in
Lindgren et al. (1996) administered a
screening neuropsychological battery by
testers blinded to the degree of lead exposure
of the worker had their performance
compared to age matched norms.
If performance on two or more tests in any
functional domain was below 1.5 standard
deviations the worker was invited for a
complete clinical evaluation. Eighty current
workers were identified by this criterion.
Mean years- age 44 (8.4), education 8 (2.8)
and duration employed 20 (5.3). Five
neuropsychological tests commonly
associated with lead exposure were
examined for a differential association with
blood lead, IBL,TWA and bone lead.
Of the 80 current smelter workers described
above 78 completed a simple visual reaction
time (SRT) and had mean years age 44 (8.2)
years, education 8 (7.2) and duration
employed20 (5.6).
Mean blood lead 26 (7.07)
ug/dL
Mean TWA 42 (8.4) ug/dL
Mean IBL 903 (305.9)
ug-yr/dL,
Mean tibial bone lead
41 ug/g bone mineral
Mean blood lead, 26 (7.2)
ug/dL
Mean blood lead from bone
7 (4.2) ug/dL
Mean blood lead from
environment 19 (7.0) ug/dL
Mean bone lead 40 (25.2)
ug/g bone mineral
Relationship of 5 neuropsychological tests with 4 measures of lead dose
after adjusting for age age2 and education, education2 found RAVLT trial
V and Verbal Paired Associates were associated with blood lead
(AR2 = 6.2%, p = 0.02; AR2 = 5.5%, p = 0.07) and TWA (AR2 = 3.2%,
p = 0.09; AR2 = 13.9%; P = 0.00) while Digit Symbol and Grooved
Pegboard were associated with TWA (AR2 = 6.1%, p = 0.00; AR2 = 5.5%,
p = 0.02) and IBL (AR2 = 4.8%, p = 0.01; AR2 = 5.7%, p = 0.02). Only
grooved pegboard was associated with bone lead (AR2 = 4.2%, p = 0.05).
Block design was not associated with any measures of lead dose. Age was
an effect modifier with grooved pegboard. There was enhanced slowing in
older workers when compared to younger workers with the identical IBL.
SRT consisted of 44 responses to a visual stimulus at interstimulus
intervals (ISI) varying between 1 through 10 seconds with a mean SRT
(median) of 262 (179 to 387) ms. Blood lead and median SRT had a
curvilinear relationship R = Pb+ Pb , 13.7%, p < 0.01) after adjusting for
age and education with slowing of SRT beginning at a blood lead of
approximately 30 ug/dL. No relationship existed between bone lead and
SRT. There was a stronger association between Pb and Pb2 and SRT for
the longer ISI = s, 6 to 10 seconds (R2 = 13.9%, p < 0.01), as age was
significantly related to the shorter ISI = s but not the longer ones. In this
population the contribution of bone lead to blood lead had been previously
where estimated where for a bone lead level of 100 ug Pb/g bone mineral,
17 ug Pb/dL of the blood lead was derived from internal bone stores with
the remainder from the environment. Blood lead was fractionated to that
from bone (blood lead-bn) versus blood lead from the environment (blood
lead-en). Regression analysis to examine the relationship of blood lead-bn
and blood lead-en and SRT after adjusting for the covariates found
significant contribution to the variance of SRT only for blood lead-en (R
for blood lead-en + blood lead-en2 = 14.4%, p < 0.01). The absence of a
contribution by age and more stable responses with ISIs of 6 to 10 sec
supports using this component of SRT.
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Table AX6-3.3 (cont'd). Neurobehavioral Effects Associated with Occupational Lead Exposure in Adults
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Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
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Canada (cont'd)
Lindgren et al. (2003)
New Brunswick
1992-1993
Braun and Daigneault
(1991)
Quebec
In an attempt to separate the effects of past
high lead exposure from a lower proximate
exposure, examination of the pattern of lead
levels of the 467 Canadian lead smelter
workers found 40 workers who had high past
exposure followed by years where 90% of
blood lead were above 40 ug/dL (High-
high = H-H) while another group of 40
workers had similar past high lead exposure
followed by years where 90% of blood lead
were below 40 ug/dL (High-low = H-L).
The groups did not differ on age, education,
years of employment or CES-D. Five
outcomes examined-Purdue Pegboard
assembly, Block Design, Digit Symbol, Rey
Auditory Verbal Learning Test-total score,
delayed Logical Memory.
41 workers from a secondary lead smelter,
mean age 35 (9.6) years and years of
education 10 (2.1) were compared to a
control group mean age 37 (10.1) years and
years of education 11 (1.3) on tests of
cognitive and motor function. MANCOVA
and dose-effect relationships after adjusting
for potential confounders were performed.
Mean IBL for past
exposure
H-H 633 (202.2) ug-yr/dL
H-L 557 (144.8) ug-yr/dL
Mean IBL for the
proximate exposure
H-H 647 (58.7) ug-yr/dL
H-L 409 (46.4) ug-yr/dL
Mean blood lead
H-H 37 (5.1) ug/dL
H-L 24 (5.2) ug/dL
Mean TWA 53 (7.5) ug/dL
Mean maximum blood lead
87 (22.4) ug/dL
Of the five neuropsychological measures examined only RAVLT
(total score) and Logical Memory (delayed) were significantly
different after adjusting for the covariates in the two pattern groups.
Use of regression analyses found pattern group contributed
significantly (R2 = 4%, p < 0.05) to the explanation of variance in
RAVLT after accounting for current blood lead (R2 = 3%, p < 0.10)
and IBL measures (R2 = 7%, p < 0.01). For past IBL, H-H pattern
correlated more strongly with RAVLT (r = -0.21) while H-L pattern
had no relationship with past exposure (r = 0.08). For proximate IBL
the difference was maintained between H-H (r = -0.11) and H-L
pattern (r = 0.00). The authors suggested that the absence of an
association between past high lead exposure and verbal memory in the
H-L pattern group may reflect reversibility of function when blood
lead is maintained below 40 ug/dL.
None of the measures of cognitive executive function showed group
differences. Partial correlation adjusting for age and education with
dose related variables found no statistical significance. On motor
function the exposed workers had significantly slower simple reaction
time (p = 0.05). However partial correlations with measures of dose
found dose-effect correlation in both negative and positive directions.
Group of exposed workers was mixed for lead exposure with 11
currently working and the remainder with no exposure up to 84
months. Also two of the exposed workers had been treated with
chelation.
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Table AX6-3.3 (cont'd). Neurobehavioral Effects Associated with Occupational Lead Exposure in Adults
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Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
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6
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Europe
Hanninen et al. (1998)
Finland
Lucchini et al. (2000)
Italy
Fifty-four lead battery workers were
stratified by those whose blood lead never
exceeded 50 ng/dL (n = 26) (group 1)
and those who had higher exposure in the
past (n = 28) (group 2) to examine the
neuropsychological effects of current low
level blood lead from higher blood lead in
the past. Mean age group Iwas 42 (9.3)
years, education 8 (1.7) years and years of
exposure 12 (6.7). Mean age group 2 was 47
(6.2) years, education 8(1.0) years and years
of exposure 21 (6.9). Analysis included
partial correlations within the groups and
ANCOVA within group 1 divided at the
median TWA3 of 29 ng/dL.
66 workers in lead manufacturing, mean age
40 (8.6) years, mean education 8 ( 2.4) years
and mean exposure time 11 (9) years and a
control group of 86 with mean age 43 (8.8)
years, mean years of education 9 (2.7) years.
Group differences examined and dose-effect
relationship with correlation and ANOVA.
Markers of lead exposure
for the group 1 were mean
IBL 330 ng-yr/dL,
Maximum blood lead
40 ng/dL, TWA 29 ng/dL
Tibial lead 20 ng/g
Calcaneal lead 79 |ig/g
Past high exposure, group 2
Mean IBL 823 ng-yr/dL,
Maximum blood lead
69 (ig/dL, TWA 40 ng/dL,
Tibial lead 35 ng/g
Calcaneal lead 100 jig/g
IBL, TWA and maximum
blood lead were also
calculated for the previous
3 years with a median
TWA3 of 29 ng/dL
Mean blood lead 28 (11)
Control-mean blood lead 8
(4.5) ng/dL
Mean IBL 410 (360.8)
Hg-yr/dL,
Mean TWA 32 (14.1)
Mean years exposed
11(8.1)
Partial correlations controlling for age, sex and education in group 1
found block design, digit symbol, digit span, similarities, Santa Ana 1
and memory for design significantly associated with recent measures
of exposure and embedded figures with maximum blood lead. In
group 2 embedded figures, digit symbol, block design, and associative
learning were associated with IBL and /or maximum blood lead.
Calcaneal lead was weakly associated with digit symbol, digit symbol
retention, and synonyms. There was no association with tibial lead in
either group. Group 1 divided at the median TWA3 of 29 ng/dL
found the high group had lower scores for visuospatial and
visuoperceptive tasks (digit symbol, embedded figures and memory
for design). Overall past high exposure, blood lead >50 ng/dL, had
the greatest effect on tests requiring the encoding of complex visually
presented stimuli. The authors conclude that the effect of lead on
brain function is better reflected by history of blood lead than content
of lead in bone.
No association with neuropsychological tests (addition, digit span,
linger tapping symbol digit and motor test from Luria) and blood lead,
TWA or IBL were found. Blood lead and visual contrast sensitivities
at the high frequencies were significantly associated for the entire
group. Blood lead and serum prolactin in the whole group was
significantly associated. Increased prolactin secretion occurs with a
variety of neurotoxins and reflects impaired dopamine function in the
pituitary. The estimated threshold for a significant increase of high
prolactin levels was at a blood lead of 10 ng/dL.
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Table AX6-3.3 (cont'd). Neurobehavioral Effects Associated with Occupational Lead Exposure in Adults
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Study Description
Lead Measurement
Findings, Interpretation
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Europe (cont'd)
Osterbergetal. (1997)
Sweden
38 workers, median age 42 (no range) years
at a secondary smelter stratified by finger
bone lead concentration and along with
19 controls matched triplets for age,
education and job level. Median years
employed 10 (2-35).
High bone lead
Median bone 32
(17-101) ug/g
Median blood lead 38
(19-50) ng/dL
Median peak blood lead 63
(46-90) ng/dL
Median IBL 408
(129-1659) ng-yr/dL
Low bone lead
Median bone 16
(-7-49)ug/g
Median blood lead 34 (17-
55) ng/dL
Median peak blood lead 57
(34-78) ng/dL
Median IBL 250
(47-835) ng-yr/dL
Controls
Median bone 4
(-19-18)ug/g
Median blood lead 4(1-7)
A cognitive test battery (36 tests) covering learning and memory,
visuomotor function, visuospatial function, concentration and
sustained attention found no impairment or dose-response
relationships with any of the markers of lead exposure. Deviating test
scores (belong to 10% lowest reference norms) were less in high bone
lead (1 vs. 4 vs. 4). None of the deviating parameters were
significantly correlated with any of the lead indices. Even when age
was taken into account the significant associations between outcome
and lead exposure metrics did not exceed chance in light of the
numerous analyses performed. These were the most heavily lead-
exposed workers in Sweden. It was unusual that the 2 visuomotor
tasks significantly different had better performance in the lead-
exposed workers compared to the controls.
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Table AX6-3.3 (cont'd). Neurobehavioral Effects Associated with Occupational Lead Exposure in Adults
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Study Description
Lead Measurement
Findings, Interpretation
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Europe (cont'd)
Stolleiyetal. (1991)
England
Stollery (1996)
England
Earth et al. (2002)
Austria
Seventy lead-exposed workers, mean age
41 (no SD) years, grouped by blood lead
(<20 |ig/dL, 21-40 ng/dL and 41-80 ng/dL)
examined on three occasions each separated
by four months. Tested on a computer for
syntactic reasoning, delayed five choice
reaction time, visual spatial memory, and
category search task.
Same as above except this was a further
analysis of the five choice reaction time.
47 lead storage-battery workers, mean age
40 (9.7) years and 53 nonexposed controls,
mean age 39 (8.4) years were matched for
age and verbal intelligence. Group
differences and dose-response relationship
were explored.
Low blood lead (no SD
provided)
Mean blood lead 14 |ig/dL
MeanZPP13mg/dL
Mean urinary ALA 2 mg/L
Mean years exposed 7
Medium blood lead
Mean blood lead 31 ng/dL
MeanZPP33mg/dL
Mean urinary ALA 3 mg/L
Mean years exposed 10
High blood lead
Mean blood lead 52 ng/dL
Mean ZPP 77 mg/dL
Mean urinary ALA 6 mg/L
Mean years exposed 11
Same as above
Mean blood lead 31 (11.2)
IBL 384 (349.0) ng-yr/dL
Years employed 12 (9.0)
Controls
Mean blood lead 4 (2.0)
Lead exposure was stable over the 8 months of testing. The low lead
group drank significantly less alcohol and rated their work as less
demanding. Performance and exposure stable except in the high lead
group where decision time was slowed more than movement time
along with concentration difficulties that remained stable across
testing sessions. Movement and decision times were significantly
correlated for each duration of waiting. On the memory test of
recalled nouns, the memory deficit associated with lead (r = -0.35,
p = 0.003) was restricted to recall of nouns unrelated to task
(distracters) (p = 0.04) that did not improve with repetition suggestive
of difficulties with incidental learning. Workers with blood lead
>40 ng/dL had impairments that correlated best with average blood
lead over the preceding 8 months. Workers with blood lead between
21 to 40 ng/dL had essentially no impairment.
Movement and decision slowing was correlated with blood lead.
Slowed movement time was constant across response-stimulus
intervals in contrast to decision time that was increasingly affected by
lead especially at the shortest response-stimulus intervals.
This supported the finding that decision gaps, central in origin, as
opposed to movement gaps are selectively affected by lead exposure
in this population.
Significant differences were found for block design (p < 0.01), visual
recognition (p < 0.01) and Wisconsin card sorting (categories
p = 0.0005, total errors p = 0.0025, perseverations p = 0.001, loss of
sorting principle p = 0.003) but not SRT or digit symbol. In the
exposed group partial correlation adjusting for age found no
significant associations with IBL (n = 53). In the entire group the full
correlation was significant for blood lead and Wisconsin card sorting,
block design and visual recognition (n = 100). Visuospatial abilities
and executive function were better predicted by blood lead than
cumulative lead exposure. It is unusual that a frontal lobe task is
associated with blood lead when SRT and digit symbol sensitive to
the affects of lead are not.
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Table AX6-3.3 (cont'd). Neurobehavioral Effects Associated with Occupational Lead Exposure in Adults
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Study Description
Lead Measurement
Findings, Interpretation
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Europe (cont'd)
Winker etal. (2005)
Austria
Winker et al. (2006)
Austria
48 workers formerly lead-exposed, mean
duration since last exposure 5 (3.5) years,
and mean age 40 (8.8) years were matched
with 48 controls for age, verbal intelligence,
years of education and number of alcoholic
drinks. Group differences and dose-response
relationship were explored.
The same 48 workers formerly lead-exposed
described above were compared to the
47 exposed workers described by Barth et al.
(2002). Both groups were comparable for
age and verbal intelligence. Group
differences and differences by duration of
exposure and exposure absence were
evaluated.
Formerly lead-exposed
Mean blood lead 5.4 (2.7)
ug/dL
Range 1.6 to 14..5 ug/dL
IBL4153.3 (36930.3)
ug-yr/dL
Controls
Mean blood lead 4.7 (2.5)
ug/dL
Range 1.6 to 12.6 ug/dL
Exposed workers
Mean blood lead 31 (11.2)
ug/dL
Range 10.6-62.1 ug/dL
IBL 4613 (4187.6)
ug-yr/dL
Formerly lead-exposed
Mean blood lead 5.4 (2.7)
ug/dL
Range 1.6 to 14.5 ug/dL
IBL 4153 (36930.3)
ug-yr/dL
No significant differences on neurobehavioral battery were present
when groups compared by t-tests for paired samples. When the
groups were combined, partial correlation adjusting for age found
significant negative correlation between blood lead and Block Design,
(r = "0.28, p < 0.01) Visual Recognition (r = "0.21, p < 0.05) and
Digit Symbol Substitution (r = "0.26, p < 0.01). The authors
conclude that the cognitive deficits associated with low-level lead
exposure are reversible. However there appears to be a residual effect
primarily from those with the highest past lead exposure.
Mann-Whitney test found significantly better performance in the
formerly lead-exposed workers for Block Design (p = 0.005) and
Wisconsin Card Sorting Test (categories p = 0.0005, total errors
p = 0.005, perseverations p = 0.0095 and loss of sorting principle
p = 0.02). To further examine the reduction of cognitive impairment
with absence of exposure, workers were stratified by duration of
exposure and exposure absence - short exposure and long absence;
long exposure and long absence; short exposure and short/no absence
and long exposure and short/no absence. Linear contrasts for Block
Design (p = 0.003) and Wisconsin Card Sorting Test (categories-
p < 0.001, total errors p = 0.001, perseverations p = 0.019 and loss of
sorting principle p = 0.030) were highly significant in the
hypothesized direction. Results were believed to support reversibility
of cognitive deficits related to occupational lead exposure.
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Table AX6-3.3 (cont'd). Neurobehavioral Effects Associated with Occupational Lead Exposure in Adults
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Study Description
Lead Measurement
Findings, Interpretation
Latin America
Maizlishetal. (1995)
Venezuela
43 workers from a lead smelter, mean age 34
(9) years and 47 nonexposed workers, mean
age 35 (11) years completed the WHO
neurobehavioral core test battery. ANCOVA
and linear regression adjusting for potential
confounders examined relationship of lead
exposure and NCTB.
Mean blood lead 43 (12.1)
Mean peak blood lead 60
(20.3) ng/dL
Mean TWA 48 (12.1)
Group comparison was significant for SRT (p = 0.06) but the lead
exposed workers performed faster. Linear regression found SRT
poorer performance with blood lead and TWA but not significant.
With peak blood lead SRT improved with increasing lead exposure.
In this study only symptoms were significantly different between
the groups. (See above).
Controls
Mean blood lead 15 (6)
Mean peak blood lead 1 5
(6) ng/dL
MeanTWA15(6)|ig/dL
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Asia
Schwartz et al.
(200 la)
South Korea
803 Korean lead exposed workers, 80% men
and 20% women, mean age 40 (10.1) years
from a variety of industries, and 135
controls, 92% men and 8% women, mean
age 35 (9.1) years. Educational levels lead-
exposed workers/controls
<6 years = 23% / 7%, 7-9 years 23% / 11%,
10-12 years = 46% / 70%, and >12 years
8% / 12%. Group differences on
neurobehavioral testing after controlling for
covariates and linear regression controlling
for covariates examined the presence of a
dose-effect relationship.
Lead-exposed workers
Mean blood lead 32 (15)
Tibia bone lead 37 (40. 3)
(Wg
DMSA-chelatable lead
level 186 (208. l)ng
Controls
Mean blood lead 5 (1.8)
Tibia bone lead 6 (7) |ig/g
Nineteen outcomes examined. Compared to controls lead exposed
workers performed significantly worse on SRT, Digit Span, Benton
Visual Retention, Colored Progressive Matrices, Digit Symbol, and
Purdue Pegboard after controlling for age, gender and education. The
association of DMSA with test performance was lost by the addition
of blood lead. Bone lead was not associated with neurobehavioral
performance, blood lead was the best predictor for significant
decrements in neurobehavioral performance on trails B (P = -0.0025
[SE 0.0009], p < 0.01), Purdue Pegboard (dom p = -0.0159 [SE
0.0042], p < 0.01; non-dom p = 0.0169 [SE 0.0042], p < 0.01; both
P = -0.0142 [SE 0.0038], p < 0.01; assem p = -0.0493 [SE 0.0151],
p<0.01 )and Pursuit Aiming (#corr p = -0.1629 [SE0.0473],
p < 0.01; #incorr p = -0.0046 [SE 0.0023], p < 0.05). The magnitude
of the effect for these eight tests significantly associated with blood
lead was an increase in blood lead of 5 ng/dL was equivalent to an
increase of 1.05 years in age. Use of Lowess lines for Purdue
Pegboard (assembly) and Trails B suggested a threshold at blood lead
18 ng/dL after which there is a decline of performance.
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Table AX6-3.3 (cont'd). Neurobehavioral Effects Associated with Occupational Lead Exposure in Adults
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Study Description
Lead Measurement
Findings, Interpretation
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Asia (cont'd)
Schwartz etal. (2005)
South Korea
1997-2001
Longitudinal decline in neurobehavioral
performance examined in 576 of the above
group of lead exposed workers who
completed 3 visits at one year intervals.
Mean age at baseline was 41 (9.5) years and
job duration 9 (6.3) years and 76% were
men.
Compared to non-completers lead workers
who completed 3 visits were 3.3 years older,
baseline mean blood lead was 2.0ug/dL
lower, on the job 1.6 years longer, 24%
women vs. 10% of noncompleters, and
usually had less than high school education.
Models examined short-term versus long-
term effects. Final model had current blood
lead, tibia bone lead and longitudinal blood
lead and covariates.
Baseline mean blood lead
31(14.2)ug/dL
Tibia lead 38 (43) ug/g
Blood lead from baseline correlated with those from visit 2 and 3 and
baseline tibial lead correlated with that measured at visit 2. Cross-
sectional associations of blood lead or short-term change occurred
with Trails A (P = -0.0020 [95% CI: -0.0040, -0.0001]) and B
(P = -0.0037 [95% CI: -0.0057, -0.0017]), Digit Symbol
(P = -0.0697 [95% CI: -0.1375, -0.0019]), Purdue Pegboard
(dom P = -0.0131 [95% CI: -0.0231, -0.0031]; non-dom
(P = -0.0161 [95% CI: -0.0267, -0.0055]); both (P = -0.0163, [95%
CI: -0.0259, -0.0067]); assem (P = -0.0536 [95% CI: -0.0897,
-0.0175]), and Pursuit Aiming #corr (P = 0.1526, [95% CI: -0.2631,
-0.0421]) after covariates. However longitudinal blood lead was only
associated with poorer performance on Purdue Pegboard non-dom
(P = -0.0086 [95% CI: -0.0157, -0.0015](; both (P = -0.0063 [95%
CI: -0.0122, 0.0004]); assem (P = -0.0289 [95% CI: -0.0532,
-0.0046]). Historical tibial bone lead was associated with digit
symbol (P = -0.0067 [95% CI: -0.0120, 0.0014]) and Purdue
Pegboard dom (P = -0.0012, [95% CI: -0.0024, -0.0001]).
Magnitude of lead associations was expressed as the number of years
of increased age at baseline that was equivalent to an increase of lead
from the 25th to 75th percentile. At baseline, these lead associations
were equivalent to 3.8 years of age for cross-sectional blood lead, 0.9
years of age for historical tibial lead and 4.8 years of age for
longitudinal blood lead. Analyses showed decline in performance
over time related to tibia lead.
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Table AX6-3.3 (cont'd). Neurobehavioral Effects Associated with Occupational Lead Exposure in Adults
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Study Description
Lead Measurement
Findings, Interpretation
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Asia (cont'd)
Hwang et al. (2002)
South Korea
From the above cohort of 803 Korean lead
workers, 212 consecutively enrolled workers,
were examined for protein kinase C (PKC)
activity and the relations between blood lead
and neurobehavioral performance. PKC
activity assessed by measuring levels of
phosphorylation of three erythrocyte
membrane proteins. Seventy-four percent of
workers were men, mean age 36(0.8)years,
duration of exposure 9 (0.6) and education
93% had high school or less. For the female
workers, mean age 47 (0.9) years, duration of
exposure 6 (0.5), and education 95% had
high school or less.
Male workers
Mean blood lead 32 (13.0)
Mean tibia lead 38 (39.6)
Hg/g
Mean ZPP 69(47. 8) ng/dL
Female workers
Mean blood lead 20 (9.2)
Mean tibia lead 26 (14.7)
Hg/g
Mean ZPP 72 (29.7) ng/dL
Blood lead was associated significantly with decrements in Trails B
(P = -0.003 [SE 0.002], p < 0.10), SRT (P = -0.0005 [SE 0.0003],
p < 0.10) and Purdue Pegboard (dom p = -0.21 [SE 0.010], p < 0.05);
non-dom (P = -0.021 [SE 0.010], p < 0.05); both (P = -0.021 [SE
0.009], p < 0.05). PKC activity as measured by back-phosphorylation
of erythrocyte membrane proteins was not associated with
neurobehavioral test scores. Addition of the interaction term of blood
lead by back-phosphorylation dichotomized at the median found
significant effect modification with the association of higher blood
lead and poorer neurobehavioral performance occurring only among
workers with lower back-phosphorylation levels that corresponds to
higher in vivo PKC activity. Association of blood lead and SRT for
the 52 kDa subunit with high in vivo PKC activity (adjusted
P = -0.001, p < 0.01) and for low in vivo PKC (adjusted p = -0.0001,
p = 0.92). The authors suggest that PKC activity may identify a
subpopulation at increase risk of neurobehavioral effects of lead.
Chuangetal. (2005)
Taiwan
H
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27 workers from a glazing factory were
administered a computerized
neurobehavioral battery 3 times over 4 years.
At year 1, the mean age was 40 (9.6) years.
In the first year workers were compared to a
referent group matched for age and
education. Neurobehavioral performance
compared in first year to referent group with
adjustment for age and Vocabulary.
Generalized mixed linear mixed models
analyzed relationship between blood lead
level and neurobehavioral test performance
after adjusting for age and Vocabulary.
Yearl
Mean blood lead 26 (12)
Year3
Mean blood lead 11 (6.4)
Year 4
Mean blood lead 8 (6.9)
Referent
Mean blood lead 7 (4.2)
Referents scored significantly lower on questionnaire for chronic
symptoms in year 1. In the mixed model analyses linger tapping
dominant (p = 0.008) and non-=dominant (p = 0.025) were
significantly inversely associated with blood lead. Pattern
comparison (p < 0.001) and Pattern memory (p = 0.06) improved
significantly as blood lead levels improved. Chronic symptoms and
neurobehavioral performance appear to reverse when lead exposure is
decreased. However since the referent group was not tested in year 3
and year 4 it was not possible to control for practice effect known to
occur with repeat neurobehavioral testing even at two year intervals.
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Table AX6-3.3 (cont'd). Neurobehavioral Effects Associated with Occupational Lead Exposure in Adults
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Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
X
ON
H
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O
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O
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H
W
Asia (cont'd)
Tsai et al. (2000)
Taiwan
Chia et al. (2004)
Singapore
Chia etal. (1997)
Singapore
19 lead workers and 19 referents included in
the above publication, mean age 39 years in
both groups and mean education 10 (2.9) and
9 (3.2) years, respectively, were tested with a
computerized neurobehavioral battery.
Alcohol use was similar. Mean duration of
lead exposure 6 (2.5) years. Student's t
compared neurobehavioral performance
between the two groups.
120 workers from lead stabilizer factories,
mean age 40 (10.7) years, duration of
exposure 10.2 (7.9) were given a
neurobehavioral battery. Genotyping of
ALAD polymorphisms was performed.
ANCOVA used to test for differences in
neurobehavioral performance among ALAD
polymorphism types adjusting for age,
exposure duration and blood lead.
50 lead battery manufacturing workers, mean
age 36 (10.6) years, education 8.6 (2.1) years
duration of employment 9 (7.4) years and
97 controls, mean age 34 (3.7) years, and
education 12 (1.8) years were administered a
neurobehavioral battery. ANCOVA and
linear regression used to assess relationship
of lead dose and performance.
Mean blood lead 32 (12.2)
ug/dL
Referent
Mean blood lead 7 (2.7)
ug/dL
Mean blood lead 22 (9.4)
ug/dL
ALAD0.6 (0.25) urn of
porphobilinogen/h/ml of
RBC
ALAU0.9 (0.56) mg/g cr
Median blood lead of 38
(13.2-64.6) ug/dL
Median IBL 264
(10.0-1146.2) ug-yr/dL
Controls
Median blood lead 6
(2.4 - 12.4) ug/dL
Poorer performance in lead workers for finger tapping, dominant and
non-dominant, and continuous performance task but only finger
tapping was significant. Lead workers performed better than referents
on Associate Learning, Pattern Comparison Test, Pattern Memory
Test, Visual Delay and Associate Learning Delayed that was
attributed to higher mean education.
Frequency of ALAD1 1, 87%, ALAD1 2, 12%, and ALAD2 2, 1%.
Mean blood lead adjusting for age and exposure duration was 20
ug/dL for ALAD1 1 (n = 107) and 20.4 ug/dL for ALAD1 2 and 2 2
(n = 13). However ALAU was significantly higher in ALAD1 1
(p = 0.023). After adjusting for the covariates significant differences
for grooved pegboard dominant hand (p = 0.01), non-dominant hand
(p = 0.04), and grooved pegboard mean time (p = 0.006) were found
between ALAD 1 1 and ALAD 1 2 and 2 2. Considering cognitive
tests were part of battery it is surprising education was ignored.
As noted by the authors the study only had 13 in the group with better
performance and the ALAD1 2 or 2 2 genotypes limiting the power.
Significant group differences for Santa Ana, grooved pegboard, digit
symbol, pursuit aiming and Trails A and B after adjusting for age,
education, smoking, ethnic group and alcohol use. When the exposed
group was stratified by age, in the group >35 years the poorer
performance on digit symbol and Trails A was significantly
associated with cumulative lead and not blood lead after adjusting
for age and education.
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Table AX6-3.3 (cont'd). Neurobehavioral Effects Associated with Occupational Lead Exposure in Adults
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Study Description
Lead Measurement
Findings, Interpretation
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Asia (cont'd)
Niu et al. (2000)
China
Boey and Jeyaratnam
(1988)
Singapore
44 lead-exposed workers (17 men,
27 women) from lead printing houses, mean
age 35 (4.9) and education 9.3 (no SD) years
and 34 controls (19 men and 15 women),
mean age 33 (7.4) years and education
9.5 (no SD) years completed the NCTB.
ANCOVA controlling for age, sex and
education examined group differences and
linear regression for dose-response
relationship.
49 lead -exposed workers, mean age 26 (7.6)
years and 36 controls, mean age 30 (6.4)
years completed SRT and 8 psychological
tests covering attention, vigilance, visual-
motor speed, short-term memory, visuomotor
tracking, visual scanning, and manual
dexterity. Control group was matched for
education level. Discriminate analysis of
neurobehavioral tests performed to determine
which best discriminate the groups.
Mean blood lead 29 (26.5)
(8 workers blood lead
exceeded 50 |ig/dL)
Controls
Mean blood lead 13 (9.9)
(1 control blood lead
exceeded 50 ng/dL)
Mean blood lead 49 (15)
Controls
Mean blood lead 15 (3)
SRT (F = 2.30, p < 0.05), digit symbol (F = 4.81, p < 0.01) pursuit
aiming # correct (F = 7.186, p < 0.01) and pursuit aiming total
(F = 6.576, p < 0.01) had significantly poorer performance compared
to controls. No repression analyses provided.
Six tests were significantly different between the two groups-Digit
Symbol, Bourdon-Wiersma, Trails A, Santa Ana dominant, Flicker
Fusion and SRT. The group of tests that best differentiates lead-
exposed workers from nonexposed workers were Simple Reaction
Time, Digit Symbol (WAIS) and Trail Making Test (Part A) with
long latency in reaction time contributing three times more to the
derived function than Digit Symbol (WAIS) or Trails A.
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Table AX6-3.4. Meta-analyses of Neurobehavioral Effects with Occupational Lead Exposure in Adults
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Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
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United States
Davis etal. (1990)
Meyer-Baron and
Seeber (2000)
Goodman et al. (2002)
Schwartz et al. (2002)
Meta-analysis of 32 studies of nerve
conduction studies and lead exposure.
Meta-analysis of studies with blood lead
<70 ug/dL found 12 studies with comparable
test procedure and sufficient documentation
of results. Thirteen tests from the 12 studies
examined.
Meta-analysis of 22 studies with median
blood lead <70 ug/dL, numbers of exposed
and unexposed workers given with scores
and dispersion on neurobehavioral tests.
Letter to the Editor commenting on
shortcomings in the Goodman et al. (2002)
meta-analysis on studies of neurobehavioral
testing in workers occupationally exposed to
lead.
Exposed group
Range of mean blood lead
31 to 49 ug/dL
Controls
Range of mean blood lead 6
to 18 ug/dL
Exposed group
Range blood lead 24 to
63 ug/dL
Unexposed group
Range blood lead 0 to
28 ug/dL
Presented 41 effect sizes with the overall effect size for all studies
D = -0.369 (p < 0.001). All median nerves combined was
D = -0.481 (p < 0.001) and for all ulnar nerves D = -0.211
(p < 0.001). The median motor was most sensitive with an effect
size of D = -0.553 (p < 0.001). Overall blood lead was a weak
measure of exposure for the peripheral nervous system. Paradoxical
association found effect size smaller with increasing blood lead but
increased with duration of exposure.
Block Design, Logical Memory, and Santa Ana had performance
deficits with small effect size. For Block Design the effect size was
comparable to changes observed with 20 years of aging. Aiming,
SRT, Trials A and B, Digit Span and Digit Symbol also had poorer
performance but the large variance for effect sizes suggest other
factors besides lead exposure influenced performance. The authors
conclude, "that the evidence of neurobehavioral deficits at a blood
lead of approximately 40 ug/dL is obvious."
Digit symbol and D-2 errors significant effect for fixed effects,
weighted random effects and unweighted random effects. Simple
reaction time, grooved pegboard, Trails A and B, picture completion
visual reproduction, eye-hand coordination and vocabulary had
significant effects for the fixed effects model only. The authors
conclude none of the individual studies were adequate or conclusive
of subclinical neurobehavioral effects of exposure to lead as the
biological effects of blood lead <70 ug/dL are inconsistent. (See
Schwartz et al. (2002) for comments).
The six points regarding problems with the methodology included:
(1) no evaluation of quality of study design or statistical methods,
(2) data from poorly done and well done studies are combined,
(3) included 6 studies with no age adjustment and 3 with no
adjustment for education, (4) confounding of age and education
when addressed the variation across studies not discussed, (5) main
effect only examined exposed versus nonexposed comparisons that
are known to have the lowest power, cannot evaluated dose-effect
relationships and have a tendency for selection bias, and (6) few of
the 22 studies included contributed to effect size.
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Table AX6-3.4 (cont'd). Meta-analyses of Neurobehavioral Effects with Occupational Lead Exposure in Adults
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Study Description
Lead Measurement
Findings, Interpretation
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United States (cont'd)
Seeber et al. (2002)
A comparison of the two meta-analyses
Meyer-Baron and Goodman) performed to
evaluate recommendations of a German BEI
of 40 ug/dL.
Graves etal. (1991)
A meta-analysis on 11 case-control studies of
Alzheimer's disease for occupational
exposure to solvents and lead.
Effect size calculated for 12 tests in two meta-analyses and 10 tests
from one meta-analyses found subtle impairments associated with
blood lead between37 ug/dL and 52 ug/dL for Logical Memory,
Visual Reproduction, Simple Reaction Time, Attention Test d2,
Block Design, and Picture Completion, Santa Ana, Grooved
Pegboard and Eye-hand Coordination. Effect sizes related to age
norms between approximately 40 to 50 years. For example, -3 score
on Block Design = 10 to 15 years; -3.5 score on Digit Symbol =
10 years; -21 score on Cancellation d2 = 10 years; and +5 to +6 on
Trails A = 10 to 20 years. This analyses concluded that both meta-
analyses supported recommendation for German BEI of 40 ug/dL.
Four studies had data for lead exposure with a pooled analysis of
relative risks for occupational lead of 0.71 (95% CI: 0.36, 1.41).
The exposure frequencies was 16/261 for the cases and 28/337 for
the controls.
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Table AX6-3.5. Neurophysiological Function and Occupational Lead Exposure in Adults
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Study Description
Lead Measurement
Findings, Interpretation
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Canada
Bleecker et al. (2005b)
New Brunswick
1992-1993
74 current smelter workers, mean age
44 (8.4) years, education 8 (2.8) years and
employment duration 20 (5.3) year had
current perception threshold (CPT) measured
for large and small myelinated and
unmyelinated nerve fibers in the finger.
Linear regression modeled CPT on metrics
of lead dose after adjusting for covariates.
Interaction of lead dose and ergonomic
stressor on peripheral nerve function was
assessed.
Mean blood lead 26 (7.1)
ug/dL
Mean IBL 891 (298.8)
ug-yr/dL
Mean TWA 42 (8.4)ug/dL
Mean tibia bone 40 (23. 8)
5 metrics relating to IBL
cumulated only exposure
above increasing blood lead
ranging from 20 to
60 ug/dL
Blood lead and tibial bone lead were not associated with any of the
three nerve fiber populations. IBL and TWA accounted for a
significant percentage of the variance only for the large myelinated
nerve libers (AR2 = 3.9%, AR2 = 8.7% respectively). The relationship
of CPT and TWA was curvilinear with a minimum at a TWA of 28
ug/dL. Unique variance of CPT for large myelinated fibers explained
by different thresholds of IBL were IBL - 3.9%, p = 0.08; IBL20 -
5.8%, p < 0.03, IBL30 - 7.8%, p < 0.02; IBL40, p < 0.005; IBL50,
p < 0.005; and IBL60, p < 0.005. IBL60 also explained significant
variance of CPT for small myelinated nerve fibers demonstrating an
increased impairment in peripheral nerve function. This effect on
myelinated sensory nerve fibers was enhanced when a measure of
ergonomic stress was added to the model for IBL60.
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Europe
Kovalaetal. (1997)
Finland
60 workers in a lead battery factory with a
mean age of 43 (9) years and mean exposure
duration of 16 (8) years. Nerve conduction
studies, vibration thresholds, and quantitative
EEG were performed. Relationship of lead
exposure with peripheral nerve function and
quantitative EEG were examined by partial
correlation and regression analyses adjusting
for age.
Mean Tibial lead 26 (17)
mg/kg
Mean Calcaneal lead
88 (54) mg/kg
Mean IBL 546 (399)
ug-yr/dL
Mean TWA 34 (8.4) ug/dL,
Mean Max blood lead 53
(19) ug/dL,
Mean blood lead 27 (8.4)
ug/dL
The sensory amplitude of the median and sural nerves had a negative
correlation with IBL and duration of exposure that was not related to
age. Vibration threshold at the ankle related significantly to IBL and
duration of exposure after adjusting for age. Vibration threshold in
the finger was associated with blood lead and blood lead averages
over the past three years. The alpha and beta frequencies were more
present in workers with higher long term lead exposure such as tibial
and calcaneal, IBL and TWA. Overall historical blood lead measures
were more closely associated with peripheral nerve function than
bone lead concentrations. The study had no comparison group and
did not account for the effect of smoking and alcohol use or give their
usage in this population.
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Table AX6-3.5 (cont'd). Neurophysiological Function and Occupational Lead Exposure in Adults
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Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
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Asia
Schwartz et al.
(200 la)
South Korea
1997-1999
Schwartz etal. (2005)
South Korea
1997-2001
804 workers from 26 different lead using
facilities and 135 controls with a mean age of
40 (10.1) and 35 (9.1) years respectively,
job duration of 8 (6.5) and 9 (5.3) years
respectively, and education level 42 % and
69% completed high school respectively had
comparable alcohol and smoking use. Linear
regression used to compare vibration
threshold in lead exposed and controls
controlling for potential confounders.
Longitudinal decline in neurobehavioral
performance examined in 576 of the above
group of lead exposed workers who
completed 3 visits at one year intervals.
Mean age at baseline was 41 (9.5) years and
job duration 9 (6.3) years and 76% were
men.
Compared to non-completers lead workers
who completed 3 visits were 3.3 years older,
baseline mean blood lead was 2.0 ug/dL
lower, on the job 1.6 years longer, 24%
women vs. 10% of noncompleters, and
usually had less than high school education.
Models examined short-term versus long-
term effects. Final model had current blood
lead, tibia bone lead and longitudinal blood
lead and covariates.
Lead-exposed workers
Mean blood lead 32 (15)
ug/dL
Tibia bone lead 37 (40.3)
ug/g
DMSA-chelatable lead
level 186(208. l)ug
(4 hour collection)
Baseline mean blood lead
31 (14.2) ug/dL
Tibia lead 38 (43) ug/g
After adjustment for age, gender, education and height, tibia lead but
not blood lead was significantly associated with poorer vibration
threshold in the dominant great toe but not the finger (P = -0.0020
[SE 0.0007], p < 0.01). These results contrast with those for
neurobehavioral measures (see above) performed in the same study
where tibial lead was not a predictor of performance.
After adjustment for age, visit number, education, gender, height
(for vibration) and BMI (for grip strength and pinch) vibration
threshold in the dominant great toe and not the finger was associated
with tibia lead (P = -0.0006 [95% CI: -0.0010, -0.0002]) and
longitudinal blood lead (P = -0.0051 [95% CI: -0.0078, -0.0024])
in one Model and blood lead (P = -0.0019 [95% CI: -0.0039,
0.0001]) in another model.
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Table AX6-3.5 (cont'd). Neurophysiological Function and Occupational Lead Exposure in Adults
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Study Description
Lead Measurement
Findings, Interpretation
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Asia (cont'd)
Chuang et al. (2000)
Taiwan
Chiaetal. (1996a)
Singapore
206 lead battery workers, mean age 41 years,
with annual blood lead for the previous five
years had vibration perception measured in
hand and foot. Relationship of lead exposure
term and vibration perception threshold
assessed with multiple regressions, hockey
stick regression analysis after adjusting for
potential confounders.
72 workers in a lead battery manufacturing
factory with a mean age of 30 years and
reference group of 82 workers had nerve
conduction studies and blood lead performed
every 6 months over the course of three
years. Only 28 lead battery workers
completed the program. At the end of the
first year of the 82 workers in the comparison
group only 26 remained and by year 3 this
had decreased to 4. Mean nerve conduction
values examined by ANCOVA between the
exposed and reference after adjustment for
age, ethnic group, smoking and drinking
habits. Analysis of serial nerve conduction
values and blood lead treated as a clustered
sample had the within-cluster regression
coefficient examined. The 28 exposed
workers were stratified by blood lead level
and the relationship between nerve
conduction values and blood tested within
the cluster.
Mean blood lead 28 ug/dL,
Mean blood lead over past
5 years 32 ug/dL
Mean maximum blood lead
39 ug/dL
Mean index of cumulative
exposure 425 ug-yr/dL,
Mean TWA 32 ug/dL
Mean working duration
13 years and life span in
work 31%.
The geometric mean blood
lead concentrations for the
6 testing periods were 37,
41, 42,40, 41, and
37 ug/dL.
The overall range for blood
lead was 16-73 ug/dL.
After adjustment for age, sex, body height, smoking, alcohol
consumption, and use of vibrating hand tools, significant association
between mean blood lead and mean TWA and vibration perception in
the foot were found. After adjustment for the covariates, a hockey
stick regression analysis of foot vibration threshold versus mean
blood lead concentration for 5 years found an inflection point around
30 ug/dL with a positive linear relation above this point suggesting
a potential threshold.
The relationship between blood lead levels and nerve conduction
values for the 28 exposed workers was significant for all outcomes
except median motor conduction velocity and ulnar sensory nerve
conduction velocity and ulnar sensory amplitude. The regression
correlation coefficients for blood lead >40 ug/dL was significant for
all parameters except the median sensory conduction velocity and
for blood lead <40 ug/dL there was no association with nerve
conduction values. Therefore the blood lead level associated with
no change in nerve conduction studies was <40 ug/dL.
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Table AX6-3.5 (cont'd). Neurophysiological Function and Occupational Lead Exposure in Adults
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Study Description
Lead Measurement
Findings, Interpretation
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Asia (cont'd)
Chiaetal. (1996b)
Singapore
Chuang et al. (2004)
Taiwan
Yokoyama et al.
(1998)
Japan
Extension of above study - 72 workers in
lead battery manufacturing and 82 controls.
Mean duration of exposure 5.3 years.
181 lead battery manufacture workers were
stratified by milk drinkers, n = 158 and non-
or rare mild drinkers n = 23. Mean age in the
two groups was 40 and 36 years and working
duration 10/8 years respectively. Peripheral
nerve evaluation was with current perception
threshold at 3 frequencies 5Hz = C fibers,
250 Hz = A-delta fibers and 2000 Hz = A-
beta fibers. Linear regression estimated the
association of CPT and lead exposure
variable and adjustment of milk intake and
potential confounders.
17 gun-metal workers, mean age 48 years
and a 20 controls with a mean age of 45 years
had distribution of conduction velocities
(DCV) measured and the maximum median
sensory conduction velocity (SVC)
performed twice at a year interval. Group
differences controlling for confounders and
dose-effect relationships were examined.
Mean blood lead 37 ug/dL
Mean blood lead
cumulative
137 ug-yr/dL
Blood lead 25 ug/dL
milk drinkers
30 ug/dL non or rare milk
drinkers
TWA 28 ug/dL milk
drinkers
32 ug/dL non or rare milk
drinkers
IBL 316 ug-yr/dL milk
drinkers
245 ug-yr/dL non or rare
mild drinkers
Mean blood lead 40 ug/dL
Mean mobilized Pb
(CaEDTA) in urine
1 mg/24 h
ANCOVA found significant differences for all nerve conduction
parameters except three for the ulnar nerve, after adjusting for age,
ethnic groups, smoking and drinking habits. There was no
significant correlation between blood lead and blood leadCum with
nerve conduction values after linear regression with adjustment for
confounders. When blood leadCum was stratified- 12 workers
<40 ug-yr/dL, 28 workers 40-300 ug-yr/dL, 21 workers >300 ug-
yr/dL ANCOVA found significant differences for 5 nerve conduction
parameters. The strongest dose effect relationship was for sensory
nerve conduction velocity.
Age was significantly different but distributions of gender, smoking,
alcohol use, use of hand vibration tool, working history and height
were not different. Linear regressions found association of 5 Hz CPT
and 250 Hz CPT in hand and foot with blood lead and TWA but not
IBL. However the protective effects of drinking milk was present for
all fiber populations only in the hands. This paper presents an
unusual finding of subclinical lead neuropathy involving the
unmyelinated and small myelinated fibers. Toxic axonopathies
classically involve the large nerve libers. The main group difference
may be related to other nutritional deficiencies associated with the
malabsorption syndrome that lead to the non-milk drinking status.
ANCOVA controlling for age and alcohol found mobilized lead was
associated with significant slowing in the large nerve fibers while
blood lead was not. Workers with increased change in mobilized
lead over 1 year interval (mean 0.44 mg/24hr) had significant
reduction in large fiber (V95) conduction velocity while those
workers with less change in mobilized lead (0.08 mg/24hr) did not
have significant change in DCV or SVC. It appears that larger faster
conducting nerve fibers are susceptible to lead and a measure of body
burden (readily mobilized lead from soft tissue) is a stronger
predictor of this change than blood lead.
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Table AX6-3.5 (cont'd). Neurophysiological Function and Occupational Lead Exposure in Adults
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Study Description
Lead Measurement
Findings, Interpretation
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Asia (cont'd)
Heetal. (1988)
China
Niu et al. (2000)
China
40 workers in a lead smelter with age range
20 to 45 years (no mean provided) and
duration of exposure 5.4 years. Fifty controls
age 20 to 55 years. Nerve conduction studies
examined 11 parameters. Student = s t-test
examined for differences between exposed
and controls.
44 lead-exposed workers (17 men,
27 women) from lead printing houses, mean
age 35 (4.9) and education 9.3 (no SD)years
and 34 controls (19 men and 15 women),
mean age 33 (7.4) years and education 9.5
(no SD) years had nerve conduction studies
for maximal motor nerve conduction
velocity. ANCOVA controlling for age, sex
and education examined group differences
and linear regression for dose-response
relationship.
Mean blood lead 40 ug/dL
Mean urinary lead 71 ug/dL
Mean ALAU5 ug/dL
Mean blood lead 29 (26.5)
ug/dL
(8 workers blood lead
exceeded 50 ug/dL)
Controls
Mean blood lead 13 (9.9)
ug/dL
(1 control blood lead
exceeded 50 ug/dL)
There were no symptoms or signs of peripheral nerve disorder.
Both motor and sensory conduction velocities were slowed in the
lead exposed groups. 10 nerve conduction parameters were
significant in the group with blood lead >40 ug/dL and 6 parameters
were significant in the group with blood lead <40 ug/dL. An unusual
finding in this study was the lack of age association with nerve
conduction values and therefore it was not controlled for in the
analyses.
Only 12 lead exposed workers and 24 controls examined for NCV.
Left ulnar nerve was significantly slower but the left median and
right ulnar were faster in the lead exposed and the right median was
slightly slower. This appears to be a finding of chance due to the
small n. For the lead exposed group mean left ulnar C V was
52 while the mean right ulnar CV was 59 while for the controls left
ulnar CV was 58 while the mean right ulnar CV was 55.
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Table AX6-3.6. Evoked Potentials and Occupational Lead Exposure in Adults
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Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
Canada
Bleecker et al. (2003)
New Brunswick
1992-1993
359 currently employed smelter workers, mean age 41
years, had brainstem auditory evoked potentials
(BAEP) measured. Relationship between absolute
latencies and interpeak latencies assessed using linear
regression after adjusting for potential confounders.
Exposure was assessed in cases with clinical
abnormalities in Wave I and I-V interpeak latency
compared to those workers with normal BAEP using
post-hoc analysis.
Mean blood lead 28 ug/dL
Mean TWA 39 ug/dL
MeanIBL719 ug-yr/dL
Linear regression after the contribution of age found blood lead and
TWA were significantly associated with Wave I while IBL was
significantly associated with Wave III and I-III interpeak interval.
Four groups created with increasing abnormalities based upon
clinical cut-off scores for Wave I and I-V interpeak interval had
similar age. blood lead, TWA and IBL were all significantly higher
in the group with prolonged Wave I and I-V interpeak interval
compared to the group with normal BAEP = s. These findings
support involvement of the brainstem and auditory nerve with lead
exposure.
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Europe
Abbateetal. (1995)
Italy
Discalzi et al. (1992)
Italy
Discalzi et al. (1993)
Italy
300 lead exposed men ages 30 to 40 years in good
health with no other neurotoxic exposure had PI00
latency measured for visual evoked potentials (VEP)
for 15 and 30 minute of arc. Groups created based
upon blood lead had VEPS examined followed by
linear regression for each group.
49 lead exposed workers and 49 age and sex matched
controls had BAEPs measured. Relationship of 6
BAEP outcome variables and lead exposure examined
with analysis of variance and linear regression.
22 battery storage workers, mean age 35 years and 22
control group, age and sex matched, with normal
hearing had BAEPs recorded. Latencies I and V and
lead exposure examined by ANOVA after stratifying
blood lead.
Blood lead 17 to 60 ug/dL
range
Mean blood lead for
4 groups
n=39
n= 113
n=89
n=59
23 ug/dL
30 ug/dL
47 ug/dL
56 ug/dL
Mean blood lead 55 ug/dL
and TWA for previous
3 years 54 ug/dL
Mean blood lead 48ug/dL
ANOVA of the blood lead and PI00 latencies were significantly
prolonged for 15 and 30 minutes of arc. Linear regression found the
association of blood lead and PI00 were significant in each group
but the relationship was not proportional (angular coefficient).
Effect of blood lead on VEP began at 17-20 ug/dL. With age
limited to one decade, contribution from age was not a concern.
Even though no comparison group, careful screening ruled out other
medical and eye conditions and other potential exposures.
Latencies for waves I, III, V and interpeak latencies, I-V, I-III, and
III-V were all significantly prolonged in the lead-exposed workers
(p < 0.04). No significant association found with linear regression
between BAEP outcomes and exposure variables. In those workers
with TWA >50 ug/dL, I-V latency was significantly prolonged
compared to workers with TWA <50 ug/dL.
Interpeak latency I-V was significantly prolonged in lead exposed
workers (p = 0.001). No significant associations by linear
regression between I-V and lead exposure. Stratifying lead exposed
workers by blood lead 50 ug/dL found I-V interpeak latency
significantly prolonged (p = 0.03) in subgroup with higher blood
lead.
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Table AX6-3.6 (cont'd). Evoked Potentials and Occupational Lead Exposure in Adults
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Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
X
ON
Latin America
Counter and Buchanan
(2002)
Ecuador
30 lead-glazing workers, median age 35 years, had pure-tone
thresholds and BAEPs performed. Regression analyses examined
relations between auditory outcomes and blood lead.
Mean blood lead 45 ug/dL
(range 11 to 80 ug/dL)
Sixty percent of the men and 20 percent of the
women had abnormal high-frequency thresholds,
however there was no significant relationship with
blood lead and pure tone threshold at all frequencies.
Analysis of BAEPs found agreement between
latencies for Waves I, III and V and peripheral
hearing status. Interpeak latencies were within
normal limits but no analysis provided with lead
exposure. Workers lived in a lead contaminated
environment from discarded lead-acid storage
batteries. Therefore a measure of chronic lead
exposure may have been more appropriate.
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Asia
Holdstein et al. (1986)
Israel
20 adults and 8 children (mean age 27 years, range 8-56 years)
accidentally exposed to lead through food until one year prior to
measurement of BAEP.
Hirata and Kosaka
(1993)
Japan
41 lead-exposed men from lead-glass-based colors manufacturing
(n = 20), production of lead electrode plates (n = 8), casting of
lead-bronze (n = 4) and casting of lead pipes and plates (n = 9) had
mean age 41 years, mean duration of exposure 13 years. A battery
of tests administered including radial nerve conduction study,
electroretinogram (ERG), visual evoked potential (VEP),
brainstem auditory evoked potential (BAER), and short-latency
somatosensory evoked potential (SSEP). Comparison group of 39
unexposed used only for BAER analysis by Student's t test.
Correlation and linear regression controlling for age examined the
relationship of lead and the other variables.
Adult mean blood lead
31 ug/dL
Children mean blood lead
22 ug/dL
In the adults 10 month
average blood lead in adults
43 ug/dL and in children
36 ug/dL
Mean blood lead 43 ug/dL
(13-70)
Mean TWA based upon
previous 5 years 43 ug/dL
(13-70)
Mean duration of exposure
13 (0.6-29) years.
In adults, latencies I, III and I-III and I-V interpeak
intervals were significantly longer than the control
group (p < 0.05). When group stratified by 10
month average blood lead I-III interpeak interval was
longer in the high group. Age and blood lead were
not studied due to few subjects. The I-III interpeak
interval reflects transmission in the lower brainstem
and Vlllth nerve.
Significant partial correlation after adjusting for age
included TWA and radial motor conduction velocity,
blood lead and sensory conduction velocity,
exposure duration and VEP, blood lead and SSEP-
N20. Comparison of BAERs of 15 lead exposed and
39 controls found interpeak interval III-V was
prolonged significantly. It is not clear why
comparison group only used for BAERs.
Considering the large number of variables examined
with three exposure terms some of the findings could
be by chance alone.
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Table AX6-3.6 (cont'd). Evoked Potentials and Occupational Lead Exposure in Adults
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Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
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Asia (cont'd)
Murataetal. (1993)
Japan
22 gunmetal foundry workers with age range of 32 to 59 years and
work duration of 1 to 19 years and control group matched for age,
no chronic disease and no lead exposure participated. No
significant difference between groups for age, height, skin
temperature, alcohol consumption, and years of schooling. The
test battery consisted of visual evoked potential (VEP), brainstem
auditory evoked potential (BAEP), short latency somatosensory-
evoked potential (SSEP), event related potential (P300) and EKG
R-R interval variability. Paired-sample t test examined for
differences between the matched groups. Dose-effect
relationships examined with partial correlation adjusting for age
and stepwise linear regression.
Blood lead 12 to 64 ug/dL
(no mean provided)
For VEPs, N75 and N145 were significantly
prolonged in the lead exposed workers. N9-N13
interpeak latency of the SSEP was significantly
prolonged. BAEP latencies showed no significant
differences. P300 believed to reflect cognitive
function was prolonged in the lead workers and
correlated with blood lead, and PbU. Autonomic
nervous system effects were significantly diminished
for CVR.Rand for a measure of parasympathetic
activity C-CVRSA. Fifty percent of the outcome
variables showed significant group differences but
there is limited dose effect for any outcome within
the exposed group. Small sample size limited
conclusions with 20 outcome variables and 8
biomarkers of lead exposure.
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Occupational Lead Exposure in Adults
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
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Dick etal. (1999)
145 workers from a secondary lead smelter, mean
age 33 (8.7) and duration of employment 5 (4.8)
years and 84 comparison workers mean age 30
(9.3) and duration of employment 4 (4.3) years had
postural sway testing performed. The analysis of
exposure with test conditions and covariates used
mixed models.
Lead workers
Mean blood lead 39 (8.5)
ug/dL
Mean ZPP 55 (42.2) ug/dL
Mean CBL 230 (217.9) ug-
yr/dL
Mean TWA 35 9 (8.5) ug/dL
Comparison workers
Mean blood lead 2(1.7) ug/dL
The postural sway test had 6 conditions that varied the
challenge to the vestibular and proprioceptive affenernts and
visual system. Only blood lead had a significant effect
primarily on the one leg condition after the effects of the
covariates age, height, mass, and race. For the left leg,
exposure slope estimate for area (b = 0.0067, t = 3.88,
p = 0.0001) and length (b = 0.0046, t = 4.11, p = 0.0001)
were significant. For the right leg only the exposure slope
estimate for length (b = 0.0033, t = 3.02, p = 0.0029) was
significant. Dose effect was only significant when lead
workers were combined with comparison workers. If
comparison workers with blood lead level below 12 ug/dL
removed no significant exposure effect was found.
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Europe
Kovalaetal. (1997)
Finland
60 workers in a lead battery factory with a mean
age of 43 (9) years and mean exposure duration of
16 (8) years. Quantitative EEG were performed.
Relationship of lead exposure with quantitative
EEG were examined by partial correlation and
regression analyses adjusting for age.
Mean tibial lead 26 (17) mg/kg
Mean calcaneal lead
88 (54) mg/kg
Mean IBL 546 (399)
ug-yr/dL,
Mean TWA 34 (8.4) ug/dL,
Mean max blood lead 53(19)
ug/dL,
Mean blood lead 27 (8.4)
ug/dL
The alpha and/or beta frequencies were more present in
workers with higher long term lead exposure such as tibial
(p < 0.05) and calcaneal (p < 0.05), IBL (p < 0.01) and TWA
(p < 0.05). Slow alpha in workers was believed to correlate
with increased episodes of 'microdrowsiness'. The study had
no comparison group and did not account for the effect of
smoking and alcohol use or give their usage in this
population.
Asia
Iwataetal. (2005)
Japan
121 workers from a battery recycling plant and 60
age matched comparison group, mean age 46 (11)
years. Height, body weight, body mass index, and
alcohol use was similar in both groups. Lead
group had significantly more smokers. ANCOVA
used to evaluate postural sway after controlling for
age, height, and smoking and drinking status.
Benchmark dose level was calculated as the 95%
lower confidence limit of the benchmark dose.
Mean blood lead 40 (15) ug/dL
Referent
Not done
Except for sagittal sway, all postural sway parameters with
eyes open were significantly larger in lead workers. Blood
lead level in workers was significantly associated with to
sagittal sway at 1-2 Hz and 2-4 Hz with eyes open, and
sagittal and transversal sways at 1-2 Hz and 2-4 Hz with
eyes closed. The mean benchmark dose level of current
blood lead level for postural sway was 14.3 ug/dL for the
linear model and 14.6 ug/dL for the K power model.
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Table AX6-3.7 (cont'd). Postural Stability, Autonomic Testing, Electroencephalogram, Hearing Thresholds, and
Occupational Lead Exposure in Adults
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
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Asia (cont'd)
Yokoyama et al.
(1997)
Japan
Chiaetal. (1994a)
Singapore
Chiaetal. (1997)
Singapore
49 chemical workers exposed to lead
stearate, mean age 48 (1.3) years and
23 controls, mean age 47 (2.5) had postural
sway evaluated. ANCOVA examined group
differences after adjusting for covariates.
60 lead storage workers, mean age 32 (7.7)
years and 60 controls, mean age 35 (7.4) had
postural sway parameters measured.
ANCOVA used to examine group differences
after adjusting for covariates. Linear
regression examined relationship between
lead exposure and postural sway.
The same 60 lead storage workers as above
and 60 control had postural sway data
examined for contribution of cumulative
blood lead fractionated over 10 years of
exposure.
Mean blood lead 18 (1.0)
ug/dL
Mean maximum blood lead
48 (3.8) ug/dL
TWA 24 (1.3) ug/dL
Cumulative blood lead 391
(48.2) ug-yr/dL
Mean blood lead 36 (11.7)
ug/dL
Controls
Mean blood lead 6 (2.4)
ug/dL
Mean blood lead 36 (11.7)
ug/dL
Controls
Mean blood lead 6 (2.4)
ug/dL
There were significant increases in sway in all directions at high and
low frequencies with eyes open and eyes closed (p < 0.05).
Regression analysis found blood lead associated with sway in the
anterior-posterior direction, .5-lHz (0.321, p = 0.03), l-2Hz (0.313,
p = 0.04) and TWA associated with right to left sway (0.326,
p = 0.02) after adjustment for the covariates age, height, weight and
alcohol consumption. The authors conclude that change in the
vestibule-cerebellum is affected by blood lead while in the anterior
cerebellar lobe is affected by past lead exposure.
Computerized postural sway measurements found lead workers
have poorer postural stability that increased with eyes closed
(p < 0.01). Regression analysis adjusting for age, height, and
weight found no significant association with blood lead.
The lead exposed group had significantly poorer performance on all
postural sway parameters with eyes closed compared to controls after
adjusting for height, weight, age and drinking habits (p < 0.01).
All postural sway parameters with eyes closed were significantly
associated with IBL for the 2 years prior to testing (n = 23, p < 0.05).
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Table AX6-3.7 (cont'd). Postural Stability, Autonomic Testing, Electroencephalogram, Hearing Thresholds, and
Occupational Lead Exposure in Adults
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
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Asia (cont'd)
Ratzon et al. (2000)
Israel
Teruyaetal. (1991)
Japan
Ishidaetal. (1996)
Japan
63 lead battery workers, mean age 39 (8.7)
years and 48 controls mean age 36 (11.8)
years, matched for age with similar sex and
education, had postural control measured.
Group differences examined with t test.
Dose-effect relations assessed with
Pearson = s correlation coefficients. Linear
regression done with exposure category as
major predictor.
172 lead exposed workers, mean age 34
(18.4-57.4) years had cardiac autonomic
nervous system evaluated by R-R intervals
variation with respiration measured.
128 workers in the ceramic painting industry,
58 men, mean age 55 (11.7) years and
70 women, mean age 52 (9.2) years had
measures of sympathetic function by
variations in R-R interval on EKG and
changes in finger blood flow with postural
changes using Doppler flowmetry.
Correlation analyses and linear regression
examined relationship of finger blood flow
and lead exposure after adjusting for
covariates.
Mean past blood lead
38 ug/dL, mean years
employed 11 and
cumulative lead determined
by average blood lead X
years employed
Mean blood lead 36 (5-76)
ug/dL
Men
Mean lead 17 (2.1) ug/dL
ALAD62 (28.3)5
Women
Mean blood lead 11 (1.7)
ug/dL
ALAD73 (20.8)%
Using a computerized sway measurement system the exposed
workers had significantly increased mean body oscillations with eyes
closed (p < 0.01) and head tilted forward (p < 0.001). Partial
correlation adjusting for education, coffee consumption, hours of
sleep and estimate of health was significant only for total lead
exposure and increased body oscillations with head tilted forward
(p = 2.25, p = 0.0089). In order to maintain balance lead exposed
workers required increased oscillations when visual and vestibular
inputs were altered.
Age adjustment controlled for by use of ratios of predicted to
observed values. A significant dose related decrease of R-R interval
variation during deep breathing was present in 132 workers with
stable blood lead over the past year (p < 0.01). This finding was
more prominent in younger workers with blood lead > 30 ug/dL but a
mild decrease present at blood lead >20 ug/dL. A decrease in R-R
interval variation indicates decreased cardiac parasympathetic
function.
22% had blood lead >20 ug/dL, and 43% had ALAD% <60%. The
46 workers in the lowest group with blood lead <10 ug/dL had
ALAD% >80% equivalent to nonoccupational exposure and
therefore served as the control group. Blood lead (P = 0.205,
p = 0.02), smoking (P = -0.464, p < 0.01), andBMI (P = 0.213,
p = 0.01) were significant predictors of change in finger blood flow
with postural change. Decrease in change of finger blood flow is
compatible with a peripheral sympathetic nerve impairment.
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2! Table AX6-3.7 (cont'd). Postural Stability, Autonomic Testing, Electroencephalogram, Hearing Thresholds, and
^ Occupational Lead Exposure in Adults
o ^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^=^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^=
O Reference, Study
Location, and Period Study Description Lead Measurement Findings, Interpretation
Asia (cont'd)
Niu et al. (2000) 44 lead-exposed workers (17 men, 27 Mean blood lead 29 (26.5) Niu et al. (2000) examined autonomic nervous system in 44 lead
China women) from lead printing houses, mean age ug/dL exposed workers, mean blood lead 29 ug/dL, and 34 controls, mean
35 (4.9) and education 9.3 (no SD) years and (8 workers blood lead blood lead, 13 ug/dL. Linear regression found association between
34 controls (19 men and 15 women), mean exceeded 50 ug/dL) blood lead and decreased R-R interval with valsalva (F/T2.349,
age 33 (7.4) years and education 9.5 (no SD) p < 0.05) and duration of lead exposure and decreased R-R interval
years had autonomic nervous system Controls with deep breathing (F/T 3.263, p < 0.01) after adjusting for age, sex,
examined. ANCOVA controlling for age, Mean blood lead 13 (9.9) education, smoking and drinking. In the same study, quantitative
sex and education examined group ug/dL EEG found significant abnormalities in the lead-exposed workers,
differences and linear regression for dose- (1 control blood lead dominant low amplitude in 59%, dominant beta frequency in 42%
response relationship. exceeded 50 ug/dL) and abnormalities in 81%.
>
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Table AX6-3.8. Occupational Exposure to Organolead and Inorganic Lead in Adults
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Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
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United States
Schwartz etal. (1993)
Stewart etal. (1999)
Two hundred and twenty-two current
employees that manufactured tetraethyl lead
participated in a study to determine if there
was impairment on a neurobehavioral battery
associated with a measure of cumulative
exposure to organic and inorganic lead
derived from 12 years of air sampling.
Mean age was 44 (8.7) years, education
13 (1.7) years.
543 former organolead workers, mean years
since last exposurelS, examined for ongoing
neurobehavioral impairment related to past
lead exposure. Thirty-eight % were age 60
or older, predominantly white, 93% had at
least a high school degree. Linear regression
assessed the relationship between lead dose
and neurobehavioral function adjusting for
the covariates.
Cumulative lead exposure,
inorganic and organic
869 (769) ug-yr/m3
Mean years of exposure
13(9.5)
Meantibialleadl4(9.3)
Peak tibial bone lead
extrapolated back using a
clearance half-time of lead in
tibia of 27 years 24 (17.4)
DMSA chelatable lead level
19 (17.2) ug (urine collected
for 4 hours)
Exposure was divided into 4 groups with the lowest for years of
exposure and cumulative lead exposure serving as the reference
group. After adjustments for premorbid intellectual ability, age, race,
and alcohol consumption, cumulative lead exposure had differential
association poorer performance in many cognitive domains but most
often in manual dexterity and verbal memory/learning. Performance
on tests associated with exposure was 5 to 22% lower in the highest
groups when compared with the low exposure reference group.
Peak tibial lead was a significant predictor of poorer performance on
vocabulary (P = -0.063, p = 0.02), serial digit learning (P = -0.043,
p = 0.04), RAVLT trial 1 (P = -0.054, p = 0.03), RAVLT
recognition (P = -0.019, p = 0.03), Trails B (P = -0.002, p = 0.03),
finger tapping nondominant (P = -0.042, p = 0.02), Purdue pegboard
dominant (P = -0.043, p = 0.00); nondominant (P = -0.49, p = 0.00),
both (P = -0.038, p = 0.00) assembly (P = -0.133, p = 0.00) and
Stroop (P = -0.014, p = 0.00). Current tibial lead had similar
associations Vocabulary (P = 0.103, p = 0.04), Digit Symbol
(P = -0.095, p = 0.05), finger tapping dominant (P = -0.87,
p = 0.02), Finger tapping nondominant (P = 00.102, p = 0.00 ),
Purdue Pegboard dominant (P = -0.065, p = 0.01), nondominant
(P = -0.091, p = 0.00), both (P = -0.068, p = 0.00), assembly
(P = -0.197, p = 0.03 ), Stroop (P = 0.017, p = 0.01). DMSA-
chelatable lead was only significantly associated with choice reaction
time (P=-0.001, p = 0.01).
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Table AX6-3.8 (cont'd). Occupational Exposure to Organolead and Inorganic Lead in Adults
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Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
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United States (cont'd)
Stewart et al. (2002)
Balbusetal. (1997)
Balbusetal. (1998)
From the above group of former organolead
workers 535 were re-examined twice or four
times over a four year period. Also a
nonexposed control group of 118 had repeat
examinations. Mean age at first visit
exposed/controls 56 (7.4)759 (7.0),
percentage with at least a high school
education 66/71.2.
222 organolead manufacturing workers,
mean age 44 (8.7) years and 62 nonexposed
referents, mean age 43 (10) years performed
simple visual reaction time (SVRT). Linear
regression examined relationship between
lead exposure and mean RT, median RT and
standard deviation of RT after controlling for
covariates.
A second publication further examined the
above data for relationship of interstimulus
interval (ISI) and lead exposure.
First examination
Mean blood lead 5 (2.7) ug/dL
Mean tibia lead 14 (9.3) ug/g
Mean peak tibia lead 23
(16.5) ug/g
Mean exposure duration 8
(9.7) years
Mean duration since last
exposure 16 (11.7) years
Mean blood lead 20 (9.5)
ug/dL
Mean peak urine lead level
143 (130)ug/L
Same as above
On 17 of 19 neurobehavioral tests, former organolead workers
demonstrated greater annual decline in adjusted test scores
compared to controls with significant differences for Rey complex
Figure copy, RAVLT Trial 1 and RAVLT recognition. Annual
declines in performance showed greater age-related change in lead
workers compared to controls for block design, digit symbol,
serial digit learning, finger tapping and Trails A. Blood lead did
not predict annual change scores but peak tibial lead did for
symbol digit, Rey Complex Figure delayed recall, RAVLT trial 1,
RAVLT delayed recall, Purdue pegboard (1 measure) and the
Stroop. For these 6 tests it was determined that an increase of
15.7 ug/g bone mineral of peak tibia lead was equivalent in its
effect on annual test decline to 5 more years of age at baseline.
Authors conclude that data supports ongoing cognitive decline
associated with past occupational exposure to lead.
Short ISIs,l-3 seconds, had no relationship with lead exposure
while ISIs of 4-6 seconds were significantly associated with blood
lead (P = 0.06 [SE 0.02], p = 0.02 along with ISIs of 7-10 seconds
(P = 0.05 [SE 0.02], p = 0.03). ISIs 7-10 seconds with peak urine
lead levels (P = 64.29 [SE 21.86], p < 0.01).
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Table AX6-3.8 (cont'd). Occupational Exposure to Organolead and Inorganic Lead in Adults
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Location, and Period Study Description
Lead Measurement
Findings, Interpretation
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United States (cont'd)
Stewart et al. (2002)
Tassleretal. (2001)
Bollaetal. (1995)
Mitchell etal. (1996)
Population as described in Stewart et al.
(1999) and Schwartz et al. (2000). Data on
20 neurobehavioral tests from 529 former
organolead workers were evaluated to
determine if the previously described
relationship with bone lead levels is
influenced by the apolipoprotein E
(ApoE) genotype.
490 former organolead workers, mean age
58 (7.5) years. The peripheral nervous
system was examined with sensory pressure
thresholds, and pinch and grip strength.
190 current workers in organolead
manufacturing (from the 222 described in
Schwartz et al. 1993) mean age 45 (8) years
compared to 52 referents, mean age 45 (8)
years and 144 solvent exposed workers,
mean age 42 (8) years.
58 organolead workers, self-selected for a
clinical evaluation. Mean age 45 (7.1)
years.
Mean blood lead 5 (2.6) ug/dL
Mean DMSA-Chelatable lead
19 (16.3) ng,
Mean current tibia lead 15 (9.4)
Peak tibia lead 24 (17.6) ug/g
IH found organic lead was 65
to 70% of exposure in
production area.
Weighted average blood lead
24 (9.4) ug/dL
Mean blood lead 19 (6. 5)
ug/dL
Mean lifetime blood lead 26
(9.1) ug/dL
Mean lifetime urine lead 5 1
(18.8) ug/L
In 20 linear regression models, coefficients for the ApoE and tibia lead
interaction term were negative in 19 with significance reached for digit
symbol (P = -0.109 [SE 0.054], p < 0.05), Purdue pegboard dominant
(P = 0.068 [SE 0.028], p < 0.05) and complex reaction time
(P = -0.003 [SE 0.001], p<0.05) and borderline significance existed for
symbol digit (P = -0.046 [SE 0.026], p < 0.10), Trails A (P = -0.303,
[SE 0.164] p < 0.10) and Stroop (P = -0.013 [SE 0.008], p < 0.10).
The slope of the relation between tibia lead and neurobehavioral
outcome was more negative in those individuals with at least one s4
allele than individuals without this allele. It is suggested that the
presence of one Apo-s-4 allele increases the risk of persistent central
nervous system effects of lead.
No strong association was found between lead biomarkers and
measures of sensory and motor function after adjusting for age.
The authors attributed the findings to decreased sensitivity of the
peripheral nerves in this dose range of inorganic lead or the possibility
of differential repair in the peripheral nervous system compared to the
central nervous system.
Lead and solvent exposure associated with adverse effects on tests of
manual dexterity. When compared to the solvent group lead exposure
had greater impairment on memory and learning and less on
executive/motor tests. An elevated neuropsychiatric score was present
in 43% of the lead group, 15% of the solvent and 7% of the referent
group.
The most common symptoms were memory loss 74%, joint pain 56%,
trouble sleeping 54%, irritability 51%, paresthesias 49%, fatigue 49%,
nightmares 35%, moodiness 28%, headaches 21% and depression 21%.
Of the 31 workers receiving nerve conduction studies, 29% were
normal, carpal tunnel syndrome 36%, cubital tunnel syndrome 3%,
median neuropathy 3%, ulnar neuropathy 23%, mononeuropathy in
lower extremity 5%, tarsal tunnel syndrome 7% and sensorimotor
polyneuropathy 36%. 39 workers had neurobehavioral evaluation with
64% had abnormal tests of which 46% were considered to be consistent
with a toxic exposure.
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Table AX6-3.9. Other Neurological Outcomes Associated with Lead Exposure in Adults
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Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
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United States
Louis et al. 2005
New York
Louis et al. 2003
New York
Kamel et al. (2002)
Massachusetts
63 cases of essential tremor (ET) and 101
controls, similar for age, 67 (16.6) and 65
(11.1) years, education, gender and
ethnicity were examined for interaction
of blood lead and ALAD gene
polymorphisms and increased odds of
ET.
100 cases of ET and 143 controls
matched for age, sex, and ethnicity.
The relationship between blood lead and
ET was examined.
109 cases of ALS and 256 controls
matched for age, sex and region of
residence examined the relation of lead
and ALS.
ET
Mean blood lead 4 (2.2) ug/dL
Controls
Mean blood lead 3(1.5) ug/dL
2 ET cases but no controls had blood
lead >10 ug/dL
ET
Mean blood lead 3 ug/dL
Controls
Mean blood lead 2 ug/dL
Cases/controls
Mean blood lead 5(0.4)73(0.4) ug/dL
3 cases and no controls had blood lead
>10 ug/dL .
Patella lead 21 (2.1)/17 (2.0) ug/g
5 cases and 1 control had patella lead
levels >50 ug/g
Tibia lead 15(1.6)711(1.6) ug/g
2 cases and no controls had tibia lead
>50 ug/g.
Of the 63 ET cases 18 (29%) vs. 17 (17%) of 101 controls had an
ALAD-2allele(OR1.98[95%CI: 0.93, 4.21]; p= 0.077). When
log blood lead was examined by presence of ALAD2 allele in ET,
log blood lead was highest in ET cases with and ALAD2 allele,
intermediate in ET cases without an ALAD2 allele and lowest in
controls (test for trend, p = 0.10; p = 0.001). When ALAD2 allele
was present, blood lead was significantly associated with odds of
ET (OR 80.29 [95% CI: 3.08, 2.096]; p = 0.008). This increased
odds of ET with an ALAD-2 allele was 30 times greater than in an
individual with only an ALAD-1 alleles. In the highest log blood
lead tertile, ALAD2 allele was present in 22% of ET cases and 5%
of controls. It was proposed that increased blood lead along with
the ALAD2 allele could affect the cerebellum and thereby
increase the risk of tremor.
Ten cases and 7 controls had bone lead levels measured that were
significantly correlated with blood lead suggesting that higher
blood lead may have occurred in the past. Total tremor score was
correlated with blood lead (r = 0.14, p = 0.03). Logistic
regression adjusting for age and current cigarette smoking found
the odds ratio for ET was 1.19(95%CI: 1.03, 1.37) per unit
increase in blood lead. Blood lead was higher in those 39 ET
cases with no family history. Both current and lifetime prevalence
of occupational lead exposure was the same in ET cases and
controls but those with history of occupational exposure did have
a higher blood lead than those without this history (median,
3.1 ug/dL vs. 2.4 ug/dL, p = 0.004).
Increased risk of ALS was found for history of occupational lead
exposure (adjusted OR 1.9 [95% CI: 1.1, 3.3]) increased lifetime
days of exposure (adjusted OR 2.3 [95% CI: 1.1,4.9]).
Association of blood lead and ALS (adjusted OR 1.9 [95% CI:
1.4, 2.6]). Elevation in both blood lead and patella and tibia bone
lead was found in ALS cases though the precision of these
measurements was questioned (Patella lead adjusted OR 3.6 [95%
CI: 0.6, 20.6] and tibia lead adjusted OR 2.3 [95% CI: 0.4,
14.5]). Therefore, this study found lead exposure from historical
questionnaire data and biological markers associated with ALS.
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Table AX6-3.9 (cont'd). Other Neurological Outcomes Associated with Lead Exposure in Adults
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Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
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United States (cont'd)
Kamel et al. (2003)
Massachusetts
Armonetal(1991)
Minnesota
As above, the same data was used to
determine the associations of ALS with
polymorphism in ALAD and the vitamin D
receptor (VDR) and the influence of
genotype.
A case-control design with 47 ALS patients,
mean age 61 with involvement of upper and
lower motor neurons and 201 controls, mean
age 62. For the lead exposure analysis
45 male matched pairs were examined.
Same as above
Lifetime exposure to lead of
200 hours or more (years on
job x hours spent per week)
The ALAD2 allele was associated with a 2-fold increase risk of ALS
after adjustment for the covariates, age, sex, region, education and
physical activity adjusted (OR 1.9 [95% CI: 0.60, 6.3]). Additionally
adjusting for blood lead strengthened the association of ALAD2 and
ALS risk adjusted (OR 3.6 [95% CI: 0.9, 15]). This was not found
for bone lead or occupational history of lead exposure (Patella
adjusted OR 2.1 [95% CI: 0.61, 6.9]; tibial adjusted (OR 2.2 [95%
CI: 0.66, 7.3]; occup his adjusted (OR 2.4 [95% CI: 0.67, 8.7]).
VDR was not associated with lead or ALS risk.
Of 13 discordant pairs for lead exposure, 11 were in ALS patient.
The relative risk was 5.5 (95% CI: 1.44,21.0). A dose-response was
weakened by 3 controls with highest lifetime exposure. Men with
ALS worked more often at blue collar jobs and significantly more
time welding (p < 0.01). These results expanded a prior pilot study
that found a higher incidence of heavy metal exposure in ALS cases.
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Europe
Chancellor et al.
(1993)
Scotland 1990-1991
Gunnarsson et al.
(1992) Sweden 1990
A case-control design 103 ALS patients from
the Scottish Motor Neuron Disease Register
and matched community controls.
Differences in potential occupational
exposures were determined between cases
and controls.
A case-control study of 92 cases of MND
and 372 controls. MND included ALS,
progressive bulbar paresis (PBP), and
progressive muscular atrophy (PMA).
Relation of MND to risk factors including
occupational exposure examined.
Exposure to lead obtained by
lifetime employment history
from Office of Population and
Censuses and Surveys.
Physician's record review and
direct interview questionnaire.
Exposure information
obtained by self-administered
questionnaire.
Odds ration for manual labor in ALS patients was 2.6 (95% CI: 1.1,
6.3). Occupational exposure to lead was more common in ALS
patients (OR 5.7 [95% CI: 1.6, 30]).
Exposure to heavy metals primarily from welding had an increased
Mantel-Haenszel odds ratio of 3.7 [95% CI: 1.1,13.0].
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Table AX6-3.9 (cont'd). Other Neurological Outcomes Associated with Lead Exposure in Adults
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
Europe (cont'd)
Guidetti et al. (1996)
Italy
Vincetietal. (1997)
Italy
A retrospective incidence, prevalence, and
mortality survey of ALS in northern Italy
was performed.
19 ALS cases, mean age 66 (14) years and
39 controls, mean age 64 (12.9) years.
Mean air lead 3|ig/m3 in
1 975 to 1 |ig/m3 in 1 985;
blood lead in monitored
children decreased 18, 14,
and 1 1 ng/dL in same time
period.
Sporadic ALS
Mean blood lead of
1 3 (6.8) |ig/dL
Controls
mean blood lead
ll(4.4)ng/dL
The area studied had documented lead pollution for years. Based
upon 79 cases incidence and prevalence rate were comparable to the
surrounding area.
There were no cases familial ALS. Blood lead between ALS cases
and controls was not significantly different. Blood lead was
associated with disability due to ALS but no support was found for
involvement of lead in the etiology of sporadic ALS.
ON
H
6
o
o
H
O
O
H
W
O
O
HH
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ANNEX TABLES AX6-4
May 2006 AX6-68 DRAFT-DO NOT QUOTE OR CITE
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Table AX6-4.1. Renal Effects of Lead - General Population
to
o
ON
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
X
ON
ON
VO
United States
Kim et al. (1996)
Boston, MA
1979 - 1994
459 men in the Normative Aging Study; periodic exams
every 3-5 years
Mean serum creatinine at baseline
1.2 mg/dL
Random effects modeling, adjusting for baseline age, time
since initial visit, body mass index, smoking status, alcohol
ingestion, education level, hypertension (defined as blood
pressure >160 or 95 mmHg or anti-hypertensive medication
use), and, in longitudinal analysis, baseline serum creatinine
and time between visits.
Mean (SDt blood lead at baseline
9.9(6.1)ug/dL
Blood lead levels from stored red
blood cells were adjusted for
hematocrit; the assay and
adjustment procedure were
validated against freshly collected
samples. Storage tubes were
shown to be lead free.
Cross-sectional
Positive association between log transformed blood lead and
concurrent serum creatinine. 10-fold higher blood lead level
associated with 0.08 mg/dL higher serum creatinine (95% CI:
0.02, 0.13 mg/dL).
Association stronger in participants with lower peak blood lead
levels. P coefficient (95% CI) in the 141 participants whose peak
blood lead < 10 ug/dL: 0.06 (0.023, 0.097)
Longitudinal
Positive association between log transformed blood lead and
change in serum creatinine over subsequent follow-up period in
participants whose peak blood lead was <25 ug/dL
P coefficient (95% CI: 0.027 [0.0, 0.054])
Slope of age-related increase in serum creatinine steeper in group
with highest quartile of time weighted average lead exposure
compared to the lowest quartile
H
6
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O
O
HH
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Table AX6-4.1 (cont'd). Renal Effects of Lead - General Population
to
o
o
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
X
Oi
O
2
O
H
O
c
o
H
W
O
V
O
HH
H
W
United States (cont'd)
Muntner et al. (2003)
U.S.
1988-1994
Blood lead levels measured in 15,211 adult subjects enrolled
in the NHANES III study.
Study cohort representative of U.S. population; non-Hispanic
African Americans, Mexican Americans, the elderly and
children over-sampled to allow stable estimates in these
groups.
Hypertension defined as blood pressure > 140 and/or 90
mmHg and/or current antihypertensive medication use.
Based on evidence of interaction between blood lead and
hypertension, the population was stratified by hypertension
for further analysis.
4,813 hypertensives; 10,398 normotensives.
Elevated serum creatinine (%)
defined as > 99th percentile of each race-gender specific
distribution for healthy young adults [age 20-39 without
hypertension or diabetes]
11.5 % (hypertensives)
1.8 % (normotensives)
Chronic kidney disease (%)
chronic kidney disease defined as GFR <60 mL/min/1.73
m2; estimated by MDRD equation (Levey et al. [1999])
10 % (hypertensives)
1.1% (normotensives)
Multiple logistic regression
Age, race, gender, diabetes, systolic blood pressure, smoking
status, history of cardiovascular disease, body mass index,
alcohol consumption, household income, marital status, and
health insurance
Mean blood lead
4.21 (0.14) ug/dL (hypertensives)
3.30 (0.10) ug/dL (normotensives)
Higher odds ratios of both increased serum creatinine and chronic
kidney disease by quartile of blood lead in hypertensives but not
in normotensives
Hypertensives
Odds ratios for elevated serum creatinine after full adjustment:
Blood lead (range, Ug/dL) %
Quartile 1
Quartile 2
Quartile 3
Quartile 4
(0.7 to 2.4)
(2.5 to 3. 8)
(3.9 to 5.9)
(6.0 to 56.0)
7.2
12.1
12.4
16.3
Odds ratio (95% CD
1.00
1.47(1
1.80(1
2.41 (1
.03,
.34,
.46,
2.10)
2.42)
3.97)
p < 0.001 for chi-squared test for trend
Odds ratios for chronic kidney disease after full adjustment:
Blood lead % Odds ratio (95% CD
Quartile 1 6.1 1.00
Quartile 2 10.4 1.44(1.00,2.09)
Quartile 3 10.8 1.85(1.32,2.59)
Quartile 4 14.1 2.60(1.52,4.45)
p < 0.001 for chi-squared test for trend
Associations were similar when lead was entered as a log
transformed continuous variable.
In non-hypertensives, higher blood lead was associated with a
higher prevalence of chronic kidney disease, but not elevated
serum creatinine, in diabetics.
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Table AX6-4.1 (cont'd). Renal Effects of Lead - General Population
to
o
o
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
X
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2
O
H
O
c
o
H
W
O
United States (cont'd)
Paytonetal. (1994)
Boston, MA
1988-1991
Shadick et al. (2000)
Boston, MA
1991-1996
Blood lead levels measured in 744 men enrolled in the
Normative Aging Study
Serum creatinine
1.3 mg/dL
Measured creatinine clearance
88.2 mL/min
Calculated creatinine clearance
71 mL/min
Multiple linear regression adjusting for age, body mass
index, analgesic and diuretic use, alcohol consumption,
smoking status, systolic/ diastolic blood pressure
777 participants in all male Normative Aging Study
Mean blood lead
8.1 ug/dL
Blood lead levels below the limit
of detection of 5 ug/dL were
receded as 4 ug/dL (n not stated).
Mean blood lead
5.9 ug/dL
Mean Tibia Lead
20.8 ug/g bone mineral
Mean Patella Lead
30.2 ug/g bone mineral
In blood lead negatively associated with In measured creatinine
clearance
B coefficient (95% CD
-0.04 (-0.079, -0.001)
10 ug/dL higher blood lead associated with a 10.4 mL/min lower
creatinine clearance
Borderline significant associations (p < 0.1) between blood lead
and both serum creatinine (P = 0.027; neither SE nor CI provided)
and estimated creatinine clearance (P = -0.022; neither SE nor
CI provided)
A significant association between patella lead and uric acid
(P [95% CI: 0.007 [0.001, 0.013]); p = 0.02) was found, after
adjustment for age, BMI, diastolic blood pressure, alcohol
ingestion, and serum creatinine. Borderline significant
associations between tibia (p = 0.06) and blood lead (p = 0.1) and
uric acid were also observed. Notably these associations were
significant even after adjustment for blood pressure and renal
function, providing further evidence that low level lead increases
uric acid. Fifty-two participants had gout; lead dose was not
associated with risk for gout.
O
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Table AX6-4.1 (cont'd). Renal Effects of Lead - General Population
to
o
o
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
X
Oi
i
160 or 95 mmHg
or physician diagnosis with use of antihypertensive
medication), diabetes (defined as use of oral hypoglycemic
drugs or insulin or reported physician diagnosis), smoking
status, alcohol consumption, analgesic use, and, in
longitudinal models, baseline serum creatinine and its square
Six percent and 26% of subjects had diabetes and
hypertension, at baseline, respectively.
Baseline blood lead
6.5 (4.2) ug/dL
Baseline tibia lead
21.5 (13.5) ug/g bone mineral
Baseline patella lead
32.4 (20.5) ug/g
Mean blood lead levels and serum creatinine decreased
significantly over the follow-up period in the group. Lead dose
not associated with change in creatinine overall
Significant interaction of blood and tibia lead with diabetes in
predicting annual change in serum creatinine
Beta coefficient (95% CI) for natural In baseline blood lead 0.076
(0.031, 0.121) compared to 0.006 (-0.004, 0.016) for non-
diabetics
Beta coefficient (95% CI) for natural In baseline tibia lead 0.082
(0.029, 0.135) compared to 0.005 (-0.005, 0.015 for non-
diabetics
Significant interaction of tibia lead with hypertensive status in
predicting annual change in serum creatinine
Beta coefficient (95% CI) for natural In baseline tibia lead 0.023
(0.003, 0.019) compared to 0.0004 (-0.001,0.002 for non-
hypertensives
Follow-up serum creatinine was also modeled separately in
longitudinal analyses; diabetes modified the association between
baseline tibia lead and follow-up serum creatinine.
O
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Table AX6-4.1 (cont'd). Renal Effects of Lead - General Population
to
o
o
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
X
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O
2
O
H
O
c
o
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W
O
United States (cont'd)
Wu et al. (2003a)
Boston, MA
1991-1995
709 men enrolled in the Normative Aging Study
Serum creatinine
1.2mg/dL
Calculated creatinine clearance
71.3 mL/min
Serum uric acid
6.5 mg/dL
Multiple linear regression, adjusting for age, body mass
index, blood pressure or HTN (depending on model), and
alcohol ingestion. Uric acid models also adjusted for serum
creatinine, other outcome models adjusted for smoking
status and analgesic medication use.
Blood lead
6.2 (4.2) ug/dL
Tibia lead
22 (13.4) ug/g bone mineral
Patella lead
32.1 (19.5) ug/g bone mineral
Significant inverse association between patella lead and
creatinine clearance
Beta coefficient = -0.069, SE not provided
Borderline significant (p = 0.08) inverse association between tibia
lead and creatinine clearance. Borderline significant (p = 0.08)
positive associations between tibia and patella lead and uric acid.
No lead measure significantly associated with serum creatinine.
ALAD gene polymorphism also assessed. 114 participants had
the ALAD2 variant allele (7 were homozygous). None of the
three renal outcomes differed by genotype. Effect modification
by genotype on the association between tibia lead and serum
creatinine was observed; the beta coefficient (and slope) was
greater in the with group with the variant allele (P = 0.002;
p = 0.03 [SE not provided]).
Effect modification of borderline significance (p < 0.1) on
relations between of patella and tibia lead with uric acid was
observed; this was significant in participants whose patella lead
levels were above 15 ug/g bone mineral (P = 0.016; p = 0.04 [SE
not provided]). Similar to the serum creatinine model, patella
lead was associated with higher uric acid in those with the variant
allele. Genotype did not modify lead associations in models of
estimated creatinine clearance.
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Table AX6-4.1 (cont'd). Renal Effects of Lead - General Population
to
o
o
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
X
Oi
Europe
Alfven et al. (2002)
Sweden
OSCAR Study
Date not provided
N = 479 men, 542 women. All resided near two battery Blood lead
plants, 117 participants were current or former workers from 0.16 umolg/L men
plants. 0.11 umolg/L women
Renal outcome = urinary tti microglobulin
Multiple linear regression
Age, smoking status, gender (by stratification), blood
cadmium
Blood lead not associated with urinary tti microglobulin
(regression performed separately in men and women)
O
2
O
H
O
c
o
H
W
O
V
O
HH
H
W
Akesson et al. (2005)
Women's Health in the
Lund Area Study,
1999-2000
N = 820 women
Renal outcomes = GFR (estimated with cystatin C),
estimated creatinine clearance, urinary NAG and tti
microglobulin
Multiple linear regression
Age, body mass index, diabetes, hypertension, and regular
use of nephrotoxic drug, blood and urinary cadmium (in
separate models), smoking status (by stratification)
Blood lead
2.2 ug/dL
Blood lead negatively associated with estimated GFR and
creatinine clearance. No associations with NAG or tti
microglobulin
Beta coefficient (95% CI) for association between blood lead
(ug/dL) and estimated creatinine clearance (ml/min) is —1.8 (-3.0,
-0.7).
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Table AX6-4.1 (cont'd). Renal Effects of Lead - General Population
to
o
o
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
X
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O
2
O
H
O
c
o
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W
O
V
O
HH
H
W
Europe (cont'd)
De Burbure et al. (2003)
France
Study date not provided
600 adults (399 exposed, 201 age and gender matched controls)
400 children (200 exposed, 200 age and gender matched controls).
Age ranged from 8.5 to 12.3 years.
Exposure from residence near smelters
Exclusion criteria for children included obesity, diabetes and puberty;
for adults included pregnancy, cancer, diabetes and kidney disease.
Serum creatinine
1.43 mg/dL (adult male controls)
1.38 mg/dL (exposed adult males)
1.33 mg/dL (adult female controls)
1.26 mg/dL (exposed adult females)
Urinary P^-microglobulin
68.16 ug/g cr (adult male controls)
76.29 ug/g cr (exposed adult males)
63.79 ug/g cr (adult female controls)
71.98 ug/g cr (exposed adult females)
87.8 ug/g cr (boy controls)
97.3 ug/g cr (exposed boys)
88.2 ug/g cr (girl controls)
94.8 ug/g cr (exposed girls)
Urinary NAG
1.12 lU/g cr (adult male controls)
1.24 lU/g cr (exposed adult males)
0.98 lU/g cr (adult female controls)
1.28 lU/g cr (exposed adult females)
2.29 lU/g cr (boy controls)
1.70 lU/g cr (exposed boys)
2.21 lU/g cr (girl controls)
1.07 lU/g cr (exposed girls)
Urinary RBP
82.8 ug/g cr (adult male controls)
85.8 ug/g cr (exposed adult males)
83.42 ug/g cr (adult female controls)
95.81 ug/g cr (exposed adult females)
94 ug/g cr (boy controls) 99 ug/g cr (exposed boys)
110 ug/g cr (girl controls) 109 ug/g cr (exposed girls)
Renal outcome measures also included urinary total protein, albumin,
transferrin, and brush border antigens
Multiple linear regression adjusting for age, sex, body mass index, area
of residence, smoking, alcohol ingestion, mercury, cadmium and
urinary creatinine level
Geometric mean blood lead
7.13 ug/dL (adult male controls)
6.78 ug/dL (exposed adult
males)
4.17 ug/dL
(adult female controls)
5.25 ug/dL
(exposed adult females)
3.42 ug/dL (boy controls)
4.22 ug/dL (exposed boys)
2.74 ug/dL (girl controls)
3.69 ug/dL (exposed girls)
Adults
Mean blood lead level higher in exposed women but not
men. None of the renal outcomes analyzed showed any
significant difference between exposed and unexposed
groups. After adjustment for covariates, blood lead was
not associated with any renal outcomes.
Children
Mean blood lead levels higher in exposed. The highest
geometric mean blood cadmium was 0.52 ug/L. None of
the renal outcomes were significantly higher in exposed.
After adjustment for covariates, blood lead was not
associated with any renal outcomes, however, blood
cadmium was positively associated with NAG. This
association was present in both control and exposed areas.
Participants with extremes of urinary creatinine excluded
from data analyses. As a result, number of subjects in
data tables substantially less than in study.
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Table AX6-4.1 (cont'd). Renal Effects of Lead - General Population
to
o
ON
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
X
ON
Europe (cont'd)
Factor-Litvak et al.
(1993)
Kosovo, Yugoslavia
1985-1986
1447 Yugoslavian women in prospective study of
environmental lead exposure and pregnancy
Exposure from Kosovska Mitrovica with a lead smelter,
refinery and battery plant. Controls from Pristina, 25 miles
away
Renal outcome = Proteinuria assessed with a dipstick
Exclusionary criteria included HTN (n = 37 excluded,
similar blood lead levels to remaining participants)
Multiple logistic regression adjusting for age (linear and
quadratic), height (linear and quadratic), cigarette smoking,
gestational age (linear and quadratic), daily milk
consumption, no. of previous live births, average weekly
meat consumption, hemoglobin level and ethnic group.
Blood Lead
17.1 ug/dL (582 exposed)
5.1 ug/dL (865 controls)
Proteinuria (negative, trace, or > 1+)
Exposed = 16.2% negative, 74.1% trace and 9.7% with > 1+
proteinuria. Controls = 32.4% negative, 60.6% trace and 7.1%
with > 1+ proteinuria. Authors attributed overall high proportion
of proteinuria to pregnancy.
Higher blood lead associated with increased odds ratio for trace
and > 1+ proteinuria.
Comparing women in upper 10th percentile of exposure to lower
10th percentile of exposure, adjusted odds ratios (95% CI) for
trace and > 1+ proteinuria was 2.3 (1.3, 4.1) and 4.5 (1.5, 13.6),
respectively.
Limitations = limited renal outcomes assessed.
O
2
O
H
O
c
o
H
W
O
Staessen et al. (1990)
London, England
Study date not provided
531 London civil servants
(398 male, 133 female)
Exclusionary criteria = occupational exposure to heavy
metals
Serum creatinine
1.10mg/dL(men)
0.88 mg/dL (women)
Mean blood lead
12.4 ug/dL (men)
10.2 ug/dL (women)
After removal of 2 outliers, the study found no significant
correlation between serum creatinine and log blood lead in men.
No correlation between serum creatinine and log blood lead in
women
Limitations = lack of adjustment in data analysis, limited lead
dose and renal outcome assessment, loss of power by analyzing
gender in separate models
O
HH
H
W
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Table AX6-4.1 (cont'd). Renal Effects of Lead - General Population
to
o
o
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
X
Oi
O
2
O
H
O
c
o
H
W
O
Europe (cont'd)
Staessen et al. (1992)
Belgium
1985-1989
Blood lead levels were measured in 1981 adult subjects (965
males, 1016 females) enrolled in the Cadmibel study of
general Belgian population in four cadmium polluted and
unpolluted areas.
Inclusion criteria included age > 20 years and residence in
one of four study areas for >8 years. Participants were
randomly selected from the study areas; participation rates
were 78% in the two rural areas but only 39% in the urban
areas (one area from each category was known to be
cadmium polluted).
Measured creatinine clearance
99 mL/min (males)
80 mL/min (females)
Calculated creatinine clearance
80 mL/min (males)
69 mL/min (females)
Multiple linear regression
Covariates assessed included age, age squared, gender (by
stratifying), body mass index, blood pressure, ferritin level,
smoking status, alcohol ingestion, rural vs. urban residence,
analgesic and diuretic use, blood and urinary cadmium,
diabetes, occupational exposure to heavy metals, and gamma
glutamyl transpeptidase
Blood lead 11.4 ug/dL (males)
7.5 ug/dL (females)
Zinc protoporphyrin also assessed
After adjustment, log transformed blood lead negatively
associated with measured creatinine clearance
P coefficient (95% CD
-9.5 (-0.9, -18.1) males
-12.6 (-5.0, -20.3) females
A 10 fold increase in blood lead associated with a decrease in
creatinine clearance of 10 and 13 mL/min in men and women
respectively
Log transformed blood lead also negatively associated with
calculated creatinine clearance
P coefficient (95% CD
-13.1 (-5.3, -20.9) males
-30.1 (-23.4, -36.8) females
Log transformed zinc protoporphyrin negatively associated with
measured and calculated creatinine clearances and positively
associated with serum (32- microglobulin in both sexes and with
serum creatinine in men
Blood lead positively associated with serum pVmicroglobulin in
men
O
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Table AX6-4.1 (cont'd). Renal Effects of Lead - General Population
to
o
o
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
X
Oi
i
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Table AX6-4.1 (cont'd). Renal Effects of Lead - General Population
to
o
o
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
X
Oi
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Table AX6-4.2. Renal Effects of Lead - Occupational Population
to
o
o
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
United States
Smith etal. (1995)
U.S.
Study date not provided
691 construction workers
96 participants with the ALAD2 allele
Mean blood lead
7.8 ug/dL (ALAD11)
7.7 ug/dL (ALAD12 or 22)
Higher mean BUN (p = 0.03) in participants with the ALAD2
allele compared to those with the ALAD11 genotype.
However, after adjustment for age, alcohol ingestion and
blood lead, the association was no longer significant. Effect
modification was not evaluated.
X
Oi
i
oo
o
Europe
Bergdahl et al. (1997)
Sweden
Study date not provided
89 lead workers; 7 had the ALAD2 allele
34 controls; 10 had the ALAD2 allele
Median blood lead
31.1 ug/dL in lead workers with
ALAD11
28.8 ug/dL in lead workers with
ALAD12or22
3.7 ug/dL in control workers with
ALAD11
3.7 ug/dL in control workers with
ALAD12or22
Higher crude mean serum creatinine (p = 0.11) in participants
with the ALAD2 allele compared to those with the ALAD11
genotype. Adjusted data not presented.
Cardenas etal. (1993)
Belgium
Study date not provided
O
2
O
H
O
c
o
H
W
O
V
O
HH
H
W
N = 41 lead smelter workers, 41 controls (all males)
Study started with 50 lead smelter workers and 50 controls.
Blood lead level >35 |lg/dL and exposure >1 year were required
in exposed workers. Participants with renal disease, renal risk
factors, such as diabetes or regular analgesic medication use, or
urinary cadmium >2 |lg/g creatinine, were excluded.
Multiple linear regression; adjusted for urinary creatinine and, in
some cases, BMI
Serum creatinine
1.02 mg/dL (workers)
1.03 mg/dL (controls)
Battery of more than 20 renal biomarkers obtained including:
RBP
68 ug/L (workers)
64 ug/L (controls)
NAG
1.56 U/L (workers)
1.21U/L (controls)
Mean Blood lead
48.0 ug/dL (workers)
16.7 ug/dL (controls)
Mean duration of lead
exposure = 14 years
Urinary cadmium also measured
as potential confounder
Serum creatinine was not increased in lead workers compared
to controls; associations between lead dose and serum
creatinine, if assessed, were not specifically reported.
In all 82, blood lead:
-associated with thromboxane B2(P = 0.36, p < 0.01)
-negatively associated with 6-keto-prostaglandin F! aiplla
(P =-0.179, p< 0.01)
-neither SE P nor CI provided
Zinc protoporphyrin positively associated with sialic acid
excretion
NAG increased in lead workers but associated with CdU
Limitations = sample size, potential for healthy worker bias,
limited statistical analysis.
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Table AX6-4.2 (cont'd). Renal Effects of Lead - Occupational Population
to
o
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Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
X
Oi
i
oo
O
2
O
H
O
c
o
H
W
O
V
O
HH
H
W
Europe (cont'd)
Coratelli et al. (1988)
Study location and date
not provided; authors
from Italy
Pels etal. (1994)
Study location and date
not provided
Garcon et al. (2004)
France
Study date not provided
20 lead battery factory workers
20 controls
12 month longitudinal study
Renal outcomes = urinary alanine aminopeptidase, NAG and
lysozyme
81 male lead workers; 45 age matched controls
Extensive exclusionary criteria
Renal outcomes
Serum creatinine
Glomerular markers = 6-keto-prostaglandin F! aipha, thromboxane
B2, and fibronectin
Proximal tubular markers = brush border antigens (BBA, BB50,
HF5) and intestinal alkaline phosphatase
Distal nephron markers = prostaglandin E2, prostaglandin F2 alpha
Thirty-five male nonferrous metal smelter workers
Renal outcomes = di-microprotein, (32-microglobulin, retinol
binding protein, a and ji glutathione S transferases (GST)
Oxidative stress markers also measured.
All variables log transformed
Initial mean blood lead
47.9 ug/dL (workers)
23.6 ug/dL (controls)
Median blood lead
42.1 ug/dL (workers)
7.0 ug/dL (controls)
Mean blood lead = 39.6 ug/dL
Mean blood cadmium = 5.8 ug/L
Mean urine cadmium = 4.7 ug/g
creatinine
NAG and lysozyme higher in exposed compared to controls
throughout study. A statistically significant decline in urinary
NAG was noted in association with a one month period of
decreased occupational exposure in the lead workers. NAG
correlated with time of exposure (nonlinear) but not blood
lead. Clinical renal function measures were not studied.
Serum creatinine similar in exposed compared to controls.
Medians of several markers statistically greater in workers
compared to controls. After adjustment for age and
erythrocyte protoporphyrin, several renal marker outcomes
showed "some relation" to blood lead. The table of these data
shows r and r2 but not beta coefficients making the actual
statistical method used unclear.
Study limitations include lack of adjustment in statistical
analysis, potential for healthy worker bias.
Correlations between urine lead and cadmium and the renal
outcomes assessed (not blood lead or cadmium).
Significant positive correlations included:
urine lead and a GST (p < 0.01)
urine cadmium and RBP (p < 0.05)
Also, urine cadmium and 8-OHdG negatively correlated
Limitations = use of urine lead, lack of adjustment for other
covariates, sample size
Significant correlations between blood lead and two markers
of oxidative stress were observed along with a correlation
between blood cadmium and one marker of oxidative stress
-------
Table AX6-4.2 (cont'd). Renal Effects of Lead - Occupational Population
to
o
o
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
X
Oi
i
oo
to
O
2
O
H
O
c
o
H
W
O
V
O
HH
H
W
Europe (cont'd)
Gennartetal. (1992)
Study location and dates
not provided; authors
from Belgium
Gerhardsson et al.
(1992)
Sweden
Study date not provided
98 lead workers and 85 controls from initial group of 221
Renal outcomes = urinary retinol-binding protein, p-2
microglobulin, albumin, NAG, and serum creatinine and p-2
microglobulin and estimated creatinine clearance
Exclusionary criteria included lack of exposure to other metals or
solvents, urinary cadmium <2 ug/g creatinine, neurologic or renal
disease, certain medications, blood lead level >40 ug/dL
(workers) and <40 ug/dL for controls.
70 current lead smelter workers
30 retired lead smelter workers
31 active and 10 retired truck assembly workers (controls)
Renal outcomes = serum creatinine, urinary p-2 microglobulin,
NAG, and albumin, clearances of creatinine, albumin, relative
albumin, p-2 microglobulin and relative p-2 microglobulin
Blood lead measured annually since 1950; time integrated blood
lead index = summation of annual blood lead measurements
Mean Blood lead
51 ug/dL (workers)
20.9 ug/dL (controls)
Mean duration of employment
10.6 years
Median Values
Blood lead
31.9 ug/dL (current lead
workers)
9.9 ug/dL (retired lead workers)
4.1 ug/dL (current control
workers)
3.5 ug/dL (retired control
workers)
Time integrated blood lead index
369.9 ug/dL (current lead
workers)
1496.1 ug/dL (retired lead
workers)
Calcaneus lead
48.6 ug/g bone mineral (current
lead workers)
100.2 ug/g bone mineral (retired
lead workers)
Tibia lead
13.0 ug/g bone mineral (current
lead workers)
39.3 ug/g bone mineral (retired
lead workers)
3.4 ug/g bone mineral (current
control workers)
12.0 ug/g bone mineral (retired
control workers)
Mean renal outcomes were not different in workers compared
to controls. Prevalence of abnormal values was not greater in
workers compared to controls. An analysis of variance, in all
participants, by categorical blood lead, duration of
employment, ZPP, and delta-aminolevulinic acid showed no
relations with any of the outcomes (data were not shown).
Limitations include high lead levels in controls, adjustment
only for age in statistical analysis, potential healthy
worker bias
Creatinine clearance was higher in lead workers; p-values not
reported for this or other median values between lead workers
and controls.
In current lead workers, blood lead was positively correlated
with urinary p-2 microglobulin and time integrated blood lead
index was correlated with NAG (data not shown).
Strengths include assessment of cumulative lead, inclusion of
former workers
Limitations = statistical analysis, lack of power by stratifying
-------
Table AX6-4.2 (cont'd). Renal Effects of Lead - Occupational Population
to
o
ON
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
X
Oi
i
oo
Europe (cont'd)
Pergande et al. (1994)
Study location and date
not provided; research
team is German
Restek-Samarzija et al.
(1996)
Croatia
Study date not provided
82 male lead workers
44 age-matched healthy male volunteers without known exposure
to lead and living "in areas distant from the exposed people"
Renal outcomes = serum creatinine and P2 microglobulin, urinary
albumin and 14 other early biological effect markers
Exclusion criteria included prescription medication use and many
diseases; 11 workers and 3 controls excluded.
74 patients treated between 1951 and 1989 for at least one episode
of lead poisoning (53 occupational, 23 environmental)
Renal outcomes = measured creatinine clearance (collection time
not specified), GFR assessed with 99mTc-diethylenetriaminepenta-
acetic acid (DTPA) clearance
Mean blood lead
42.1 ug/dL (workers)
7.0 ug/dL (controls)
Erythrocyte protoporphyrin also
measured
Serum creatinine and P2 microglobulin not increased in
exposed compared to control participants; correlations with
these outcomes not reported. Blood lead and/or erythrocyte
protoporphyrin correlated with 9 of the urinary renal
outcomes.
Study limitations include lack of adjustment in statistical
analysis, potential for healthy worker bias, potential for
differences between exposed and control groups.
Number of past lead poisonings negatively correlated with
creatinine and DTPA clearances
O
Restek-Samarzija et al.
(1997)
Croatia
Study date not provided
38 patients with occupational lead poisoning, 23 occupationally
exposed workers
Renal outcomes = serum creatinine, measured creatinine
clearance (collection time not specified), hippuran renal flow
Mean blood lead
1.5 umol/L (poisoned workers)
1.6 umol/L (workers)
Creatinine clearance significantly lower in poisoned group.
O
H
O
c
o
H
W
O
O
HH
H
W
-------
Table AX6-4.2 (cont'd). Renal Effects of Lead - Occupational Population
to
o
o
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
Europe (cont'd)
X
Oi
i
oo
O
2
O
H
O
c
o
H
W
O
Roels et al. (1994)
Belgium
Study date not provided
76 lead smelter workers (including 21 participants from Cardenas
etal. [1993] [Dr. Roels, email communication])
68 controls
All males
Matched for age, sex, socioeconomic status, residence, and
workshift characteristics.
Extensive exclusionary criteria included renal disease, analgesic
abuse, chronic medication for gout, diabetes, occupational
exposure to other nephrotoxicants, and prior EDTA chelation.
Renal outcomes included serum creatinine and urea nitrogen,
measured creatinine clearance, NAG, RBP, serum and urinary (32-
microglobulin, as well as other renal early biological effect
markers.
Measured creatinine clearance
121.3 mL/min/1. 73 m (workers)
115.5 mL/min/1 .73 m (controls)
Multiple linear regression, adjusted for age, urinary cadmium,
hypertension, serum gamma-glutamyl transpeptidase, smoking,
exposure status (exposed vs. control), and interaction between
exposure variables and hypertension
Blood lead
43.0 ug/dL (workers)
14.1 ug/dL (controls)
Tibia Lead
66 ug/g bone mineral (workers)
21 ug/g bone mineral (controls)
CdU also measured
Creatinine clearance measured before and after an oral protein
load to determine if eicosanoid changes in Cardenas et al.
(1993) had clinical implications (Acute protein ingestion
causes increased renal perfusion and transient hyperfiltration
thought to be mediated by changes in vasodilator prostanoids.
Therefore, it was hypothesized that, if the changes noted in
Cardenas et al. (1993) were clinically significant, the
hyperfiltration response would be diminished in the lead
workers.)
All participants had normal baseline creatinine clearances
(>80 mL/min/1.73 m2). Both control and lead-exposed
workers showed a similar increment in creatinine clearance
after protein load.
However, mean creatinine clearance was statistically higher in
lead workers compared to controls. Log tibia lead was
positively correlated with log measured creatinine clearance
in the combined group (P = 0.0319, SE not provided).
This was unexpected as the change in eicosanoids found in the
initial study would not seem to result in vasodilatation with
increased GFR. Unfortunately, it was not possible to measure
eicosanoid levels in the follow-up study. No other significant
associations between lead measures and renal outcomes were
observed. CdU associated with NAG.
O
HH
H
W
-------
Table AX6-4.2 (cont'd). Renal Effects of Lead - Occupational Population
to
o
o
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
X
Oi
i
oo
O
2
O
H
O
c
o
H
W
O
V
O
HH
H
W
Europe (cont'd)
Verschoor et al. (1987)
Study location and date
not provided; authors
from The Netherlands
155 lead workers (lead battery and plastic stabilizer)
126 control industrial workers
Workers with renal disease, HTN, prescription medications
excluded
Renal outcomes = BUN, serum creatinine, uric acid,
P2-microglobulin, and RBP, and urinary RBP, NAG, albumin,
uric acid, p2-microglobulin, IgG, and total protein. Urine protein
electrophoresis performed on subset (n = 25)
Cadmium in blood and, in a subset of exposed workers, in urine
was also assessed due to this exposure in one plant each from
which lead exposed and control workers were drawn
Blood lead
47.5 ug/dL (workers)
8.3 ug/dL (controls)
Zinc protoporphyrin also used as
lead dose measure
Mean renal outcomes in all participants shown by categorical
lead levels. NAG and RBP higher at blood lead levels
>21 ug/dL compared to those below this level (statistical
significance not reported). Serum p2-microglobulin and
urinary total protein lower at blood lead levels >21 ug/dL
compared to those below this level (again, statistical
significance not reported).
In simple linear regression models of log transformed urinary
total protein, urinary RBP, NAG and serum p2-microglobulin,
higher log transformed blood lead was significantly associated
with lower serum p2-microglobulin and higher RBP and
NAG.
A matched pair analysis of 55 pairs matched for age within 5
years, smoking, socioeconomic status, and duration of
employment found no differences in renal outcomes between
exposed and controls.
Limitations = lack of adjustment, potential for healthy worker
bias, occupational cadmium exposure (including in controls)
not adequately adjusted for
Latin and South America
Cardozo dos Santos
etal. (1994)
Study location and date
not provided; authors
from Brazil
166 lead battery workers
60 control workers
Renal outcomes = serum creatinine, NAG, urine albumin, and
total urinary protein, y-glutamyl-transpeptidase, alanine-
aminopeptidase
Median blood lead
36.8 ug/dL (workers)
11.6 ug/dL (controls)
Significant results
Median NAG higher in exposed group (p < 0.001). Blood
lead level and duration of exposure correlated with NAG in
combined group (Spearman's correlation coefficients = 0.32
and 0.22, respectively, p < 0.001 for both).
No results mentioned for serum creatinine.
Limitations = statistical analysis (no regression for renal
outcomes)
-------
Table AX6-4.2 (cont'd). Renal Effects of Lead - Occupational Population
to
o
ON
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
X
ON
i
oo
Latin and South America (cont'd)
Pinto de Almeida et al. 52 primary lead smelter workers (had to have worked > 5 years on Mean blood lead
(1987) production line) 64.1 ug/dL (workers)
Northeast Brazil 25.5 ug/dL (controls)
Study date not provided 44 control paper mill workers in same city
Also measured zinc
All males protoporphyrin and delta-
aminolevulinic acid
Renal outcomes = BUN, serum creatinine, uric acid, proteinuria,
creatinine clearance
Only 2 participants excluded for medical reasons
Mean serum creatinine and uric acid higher in exposed than
controls (1.23 vs. 1.1 mg/dL; p < 0.05 and 6.6 vs. 4.7 mg/dL;
p < 0.001, respectively)
Serum creatinine > 1.5 mg/dL present in 32.7% lead workers
compared to only 2.3% controls.
Serum creatinine correlated with duration of employment.
Limitations = data analysis including lack of adjustment,
several outcomes not analyzed.
O
2
O
H
O
c
o
H
W
O
V
O
HH
H
W
Australia
Pollock and Ibels (1988) Thirty-eight bridge workers
Harbor Bridge workers Twenty-four hour urine lead excretion following 1 g of EDTA
in Sydney, Australia Renal outcomes = serum creatinine, creatinine clearance, and 24
Study date not provided hour urine protein excretion
Blood lead mean & range
34.8; 21.8 to 56.2 ug/dL (lead
intoxication)
19.9; 9.5 to 26.1 ug/dL
(nontoxic)
EDTA chelatable lead range
443 to 2366 ug/24 hrs (lead
intoxication)
131 to 402 ug/24 hrs (nontoxic)
No significant differences in renal outcomes by lead exposure
group. Two workers in high exposure group had evidence of
lead nephropathy.
Asia
Chiaetal. (1994b)
Study location not
provided; authors from
Singapore
1982-1992 (blood lead
measurements obtained
every 6 months over
this time)
128 lead workers
152 control workers without lead or cadmium exposure
Renal outcomes = total NAG, NAG-B isoenzyme (released with
lysosomal breakdown assoc with cell membranes, thought to
indicate proximal tubular cell toxicity), NAG-A (released by
exocytosis).
Cross-sectional outcomes but longitudinal exposure data.
Median blood lead
33.8 ug/dL (workers)
8.7 ug/dL (controls)
Median cumulative blood lead
(mean of 3.6 blood lead levels
per worker)
208.3 ug-yr/dL
Change in blood lead
(in 6 months preceding NAG
measurement)
Mean = 5.8%
NAG not different in exposed compared to control workers.
After adjustment for race, recent change in blood lead was
significantly associated with all NAG outcomes (standardized
partial regression coefficients ranged from 0.31 for NAG-A to
0.64 for total NAG; neither SE nor CI provided).
In contrast, current blood lead was inversely associated with
NAG-A and NAG-B separately but, oddly, not with total
NAG. Authors do not comment on these inconsistencies.
NAG not associated with cumulative lead dose.
Strengths = longitudinal exposure data
Limitations = data analysis clarity and adjustment
-------
Table AX6-4.2 (cont'd). Renal Effects of Lead - Occupational Population
to
o
o
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
X
Oi
i
oo
O
2
O
H
O
c
o
H
W
O
V
O
HH
H
W
Asia (cont'd)
Chiaetal. (1994c)
Singapore
Study location not
provided; authors from
Singapore
1982-1992 (blood lead
measurements obtained
every 6 months over this
time)
Chiaetal. (1995a)
Study location not
provided; authors from
Singapore
1982-1993 (blood lead
measurements obtained
every 6 months over this
time)
63 lead workers of >6 months work duration (median = 3 years)
91 lead workers of <6 months work duration were considered
controls
Renal outcomes = urinary BB-50 (brush border antigen in
proximal tubule), total NAG, NAG-B isoenzyme, RBP, a-1-
microgobulin, albumin and urine and serum p2-microglobulin.
Cross-sectional outcomes but longitudinal exposure data.
137 lead stabilizer workers
Control group of 153 postal workers (older than lead workers)
Renal outcomes = serum creatinine, four hour creatinine
clearance, serum (3-2 microglobulin, serum a-1 microglobulin,
urine albumin
Longitudinal blood lead data (mean of 4.5 measurements per lead
worker)
Lead Dose Measures
(means or medians not stated)
Most recent blood lead, time
integrated blood lead index,
relative % change in blood lead,
absolute change in blood lead,
# of times blood lead level >40,
50, and 60 ug/dL.
Lead Dose Measures
(means or medians not stated)
Most recent blood lead, time
integrated blood lead index,
relative % change in recent blood
lead, absolute change in recent
blood lead, # of times blood lead
level >40, 50, and 60 ug/dL.
Urinary BB-50 higher in exposed compared to recent hire
"control" workers. Time integrated blood lead, # times blood
lead >40 ug/dL, and relative change in recent blood lead were
associated with urinary BB-50.
Strengths = longitudinal exposure data
Limitations = data analysis content (lead dose means not
reported), clarity and adjustment.
In analysis of covariance modeling, adjusted for age and race,
mean serum a-1 microglobulin and urine albumin were
significantly higher in control compared to lead workers.
Serum p-2 microglobulin was significantly higher in lead
workers > 30 years of age.
After adjustment for age, race, and smoking, prevalence rates
for abnormal values of serum creatinine and (3-2
microglobulin were higher in the highest category of time
integrated blood lead index in workers > 30 years of age (PRR
[95% CI: 3.8 [1.1, 13.3] and 10.3 [3.9, 26.9], respectively).
Strengths = longitudinal exposure data
Limitations = data analysis content (lead dose means not
reported), clarity and adjustment
-------
Table AX6-4.2 (cont'd). Renal Effects of Lead - Occupational Population
to
o
o
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
X
Oi
i
oo
oo
o
2
o
H
O
c
o
H
W
O
V
O
HH
H
W
Asia (cont'd)
Chiaetal. (1995b)
Study location not
provided; authors from
Singapore
1982-1993 (blood lead
measurements obtained
every 6 months over this
time)
Endo et al. (1990)
Study location not
provided; authors from
Japan
1987
Endo et al. (1993)
Study location and date
not provided; authors
from Japan
128 lead stabilizer factory workers
93 unexposed control subjects (evaluated at pre-employment
examination; all quit within 1 month of hire)
Blood and urinary cadmium also measured on random subset (40
controls and 31 lead workers)
Renal outcomes = serum (3-2 microglobulin and urinary a-1
microglobulin, (3-2 microglobulin, albumin, RBP
39 male workers
7 female workers (none directly exposed to lead)
secondary lead refinery, mean job duration = 10.5 years
Renal outcomes = BUN, serum creatinine and uric acid, urinary
NAG, and tubular reabsorption of phosphate
99 male lead workers
Renal outcomes = serum creatinine and serum and urine alpha-1-
microglobulin
Mean recent blood lead
32.6 ug/dL (workers)
9.0 ug/dL (controls)
Mean time integrated blood lead
index
119.9 (ug/dL) x yr (workers)
0.05 (ug/dL) x yr (controls)
Mean relative change in recent
blood lead
28.2 % (workers)
Mean absolute change in recent
blood lead
6.4 (ug/dL)/year (workers)
# of times blood lead level >40,
50 and 60 ug/dL
Mean blood lead
Ranged from 24.1 to 67.6 ug/dL
(males)
19.6 ug/dL (females)
Other lead measures included
urinary lead, delta-
aminolevulinic acid, and
coproporphyrin.
Median blood lead
Ranged from 7.9 ug/dL in
category I consisting of 16 office
workers who did not work
directly with lead to 76.2 ug/dL
in 16 workers in the highest
exposure group (category V).
Only urinary a-1 microglobulin was significantly higher in
lead workers compared to controls.
In multiple linear regression analysis, adjusted only for
ethnicity and smoking, at least one lead measure was
significantly associated with each of the five renal outcomes.
Outcome
Ua-lMG
Ua-lMG
U P-2 MG
URBP
S P-2MG
UAlb
Lead measure
cum. blood lead
# blood lead >50
cum. blood lead
# blood lead >50
# blood lead >60
# blood lead >60
P (95% CD
0.10(0.06,0.14)
0.43 (0.04, 0.82)
0.05 (0.01, 0.09)
0.35(0.12,0.59)
0.47(0.29,0.65)
0.66(0.13, 1.19)
Cadmium dose measures reportedly not significant in these
models (although power would have been reduced as
cadmium measured only in a subset).
Strengths = longitudinal exposure data
Limitations = data analysis clarity and adjustment. Overlap in
populations between this study and earlier ones possible
Significant correlations of blood lead and delta-amino-
levulinic acid with BUN and NAG were observed. The
correlation between blood lead and NAG was dependent on a
small number of workers whose blood lead levels were above
80 ug/dL.
Limitations include absence of adjustment in statistical
analysis, small sample size.
Median urinary alpha-1-microglobulin significantly higher in
categories III—V compared to the low exposure group of
office workers. This was also the only renal outcome to be
significantly correlated with blood lead (Spearman rank
correlation).
After alpha-1-microglobulin adjusted for age and blood lead
(by stratifying); few significant differences noted. However,
analysis approach resulted in substantial loss of power.
-------
Table AX6-4.2 (cont'd). Renal Effects of Lead - Occupational Population
to
o
o
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
X
Oi
i
oo
VO
O
2
O
H
O
c
o
H
W
O
V
O
HH
H
W
Asia (cont'd)
Hsiao et al. (2001)
Taiwan, PR China
1991-1998
Huang et al. (1988)
Beijing, China
Study date not provided
N = 30 lead battery workers
Mean serum creatinine at baseline
— 1.0 mg/dL (based on figure; exact values not provided)
Longitudinal Analysis, 8 annual evaluations.
Generalized estimating equations used to adjust for
autocorrelation in multiple datapoints from each participant.
Adjusted for age, gender, and, in models of change in serum
creatinine, creatinine at beginning of interval.
Mean blood lead at baseline
—35 ug/dL (based on figure;
exact values not provided)
Mean duration of exposure
at baseline
13.1 years
40 lead workers (4 women)
Control group not described
Renal outcomes = serum beta-2-microglobulin and urinary beta-2-
microglobulin, total protein, IgG
Geometric mean blood lead
Cross-sectional
higher blood lead associated with lower concurrent
serum creatinine.
Longitudinal
Change in blood lead negatively associated with concurrent
change in serum creatinine (p = 0.07).
Blood lead at the beginning of the interval not associated with
change in serum creatinine in the following year.
Associations may represent lead-related hyperfiltration.
However, as noted by the authors, cumulative lead dose may
also be a factor. Mean blood lead declined greatly just before
renal data collection started. Therefore, the inverse
longitudinal associations could be due to persistently elevated
cumulative dose (which was unmeasured but, as evidenced by
the long half-life of bone lead, likely did not decline as much
as blood lead). However, authors did not model cumulative
blood lead or analyze effect modification by time period, age,
or exposure duration to determine if these associations
changed in a pattern consistent with hyperfiltration. The
small sample size also limits conclusions that may be drawn
from these results since a small number of individuals may be
overly influential.
Strengths = longitudinal data
Limitations = data analysis content (lead dose means not
reported), clarity and adjustment
Increased urinary (32 microglobulin in workers compared to
controls
Multiple limitations including lack of information on control
group, data analysis
-------
Table AX6-4.2 (cont'd). Renal Effects of Lead - Occupational Population
to
o
o
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
X
Oi
Asia (cont'd)
Jung et al. (1998)
Korea
Study date not provided
75 randomly selected male lead workers
64 male office workers (controls)
Renal outcomes = BUN, serum creatinine, uric acid and urinary
NAG, albumin, tti microglobulin and |32 microglobulin
Mean Blood lead
Means ranged from 24.3 to
74.6 ug/dL (workers)
7.9 ug/dL (controls)
Other lead measures included
zinc protoporphyrin,
8-aminolevulinic acid activity
and urinary lead, coproporphyrin,
and
8-aminolevulinic acid
Blood lead, zinc protoporphyrin, and urinary 8-aminolevulinic
acid significantly correlated with BUN, NAG, and tti
microglobulin (appears to be combined group analysis)
Limitation = statistical analysis - lack of adjustment
O
2
O
H
O
c
o
H
W
O
^
O
HH
H
W
Konishi, et al. (1994)
Study location
not provided; research
team from Japan
1991
Kumar and
Krishnaswamy (1995)
India
Study date not provided
99 male lead workers, including 16 office workers to serve at
controls
renal outcomes = fractional clearances of tti microglobulin and |32
microglobulin (utilizing serum and urinary levels of both
biomarkers), BUN, serum creatinine, uric acid and urinary NAG
22 auto mechanics volunteers
27 male control workers (from Institute performing study)
Renal outcomes = serum creatinine, 4 hour creatinine clearance
and urinary NAG and (3-2 microglobulin
Renal disease, diabetes, HTN and occupational exposures
excluded in controls, possibly excluded in workers
Median blood lead
Range from 7.9 ug/dL in controls
to 76.2 ug/dL in Category V
Blood lead range
24.3 - 62.4 ug/dL (exposed)
19.4 - 30.6 ug/dL (controls)
Urinary NAG, tti microglobulin and fractional clearance of tti
microglobulin increased with higher blood lead category.
Spearman rank correlation between fractional clearance of tti
microglobulin and blood lead was significant. This relation
also assessed by multiple linear regression with adjustment for
age; both independent variables were significantly associated
with the fractional clearance of tti microglobulin.
Limitation = statistical analysis - lack of adjustment
Urinary NAG and p2 microglobulin levels were significantly
higher in exposed compared to controls. However, only NAG
was significantly correlated with blood lead (r = 0.58,
p<0.01).
Limitations = study size and lack of adjustment in analysis,
values for 4 hour creatinine clearance in abnormal low range
in both exposed and controls
-------
Table AX6-4.2 (cont'd). Renal Effects of Lead - Occupational Population
to
o
o
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
X
Oi
O
2
O
H
O
c
o
H
W
O
V
O
HH
H
W
Asia (cont'd)
Limetal. (2001)
Singapore
1999
Blood lead levels every
6 months from 1982 to
1999
Ongetal. (1987)
Singapore and Japan
Study date not provided
Wang et al. (2002b)
Taiwan
Study date not provided
55 male lead workers
workers followed since 1982, many of same workers as in Chia
etal., (1995b)
Renal outcomes = 4 hour creatinine clearance and urinary
albumin, RBP, oil microglobulin, (32 microglobulin, NAG, NAG-
A, and NAG-B
Exclusionary criteria included diabetes, HTN, recent ingestion of
analgesics, antipyretics, or antibiotics, and thalassemia;
24 participants of the original 80 were excluded as a result.
One female also excluded.
209 lead workers (51 females)
30 control workers from research staff
Renal outcomes = BUN, serum creatinine, calculated creatinine
clearance, and urinary NAG
229 lead battery workers, including 109 females
Renal outcomes = BUN, serum creatinine, serum uric acid
Multiple linear & logistic regression
Adjustment for age, gender, smoking, alcohol ingestion, milk
ingestion.
Mean current blood lead
24.1 ug/dL
Cumulative blood index
880.6 ug x yrs/dL (geometric
mean)
Number of times blood lead
exceeded 40 ug/dL 1.9
(geometric mean)
Mean blood lead
42.1 jig/dL (males) 31.9 ug/dL
(females)
Urine lead also measured
Mean blood lead
67.7 ug/dL (males)
48.6 ug/dL (females)
In separate models, after adjustment for age and smoking,
higher categorical cumulative blood index and number of
times blood lead exceeded 40 ug/dL were associated with
lower creatinine clearance (P < 0.001).
After adjustment, higher number of times blood lead exceeded
40 ug/dL was associated with higher urinary albumin, tti
microglobulin, RBP, NAG, and NAG-B. Similarly,
cumulative blood index was associated with higher urinary
albumin, tti microglobulin, RBP, and p2 microglobulin.
No associations between recent blood lead and any of the
renal outcomes was observed.
Analysis of covariance was used to adjust for smoking and
age
Limitation = statistical analysis - lack of adjustment, small
sample size, potential for healthy worker bias
Blood lead correlated with BUN(r = 0.16; p < 0.01), serum
creatinine (r = 0.26; p < 0.001) and creatinine clearance
(r = -0.16; p < 0.01). Blood lead associated with NAG after
adjustment for age (method not specified).
Higher NAG in exposed compared to controls when stratified
by categorical age.
Strengths = sample size
Limitations = statistical analysis - lack of adjustment, urinary
NAG not adjusted for urine dilution
P coefficient (95% CI) for blood lead in model of BUN, after
adjustment for lead job duration/age = 0.062 (0.042, 0.082).
P coefficient (95% CI) for blood lead in model of uric acid,
after adjustment for gender and weight = 0.009 (0.001, 0.016).
Blood lead not associated serum creatinine
-------
Table AX6-4.2 (cont'd). Renal Effects of Lead - Occupational Population
to
o
o
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
X
Oi
OD
to
O
2
O
H
O
c
o
H
W
O
Asia (cont'd)
Weaver et al. (2003a)
South Korea
1997-1999
N = 803 lead workers including 164 females and 94 former lead
workers
Serum Creatinine:
0.90 mg/dL
Calculated creatinine clearance
94.7mL/min
4-hr measured creat. clearance
114.7mL/min
RBP
63.6 ug/g creatinine
NAG
215.3 |lmol/h/g creatinine
Multiple linear regression, adjusting for age, gender, BMI, work
status (current vs. former worker), HTN or blood pressure
(depending on model), and, for the EBE markers, alcohol
ingestion and diabetes.
42 associations modeled (7 lead measures with 6 renal outcomes)
Interaction models that assessed effect modification by age in
tertiles in 24 associations (4 lead exposure/dose measures with
6 renal outcomes).
Blood lead
32.0 ug/dL
Tibia Lead
37.2 |lg/g bone mineral
DMSA-chelatable lead
767.8 |lg/g creatinine
Lead exposure also assessed with
job duration and three
hematologic measures as
surrogates for lead dose
(aminolevulinic acid in plasma,
zinc protoporphyrin,
and hemoglobin).
Mean CdU measured in
n= 191 subset
1.1 ug/g creatinine
After adjustment, higher lead measures associated with worse
renal function in 9 of 42 models.
Associations in the opposite direction (higher lead measures
associated with lower serum creatinine and higher creatinine
clearances) in five models.
Opposite direction (inverse) associations observed only in
models of the clinical outcomes whereas the associations
between higher lead dose and worse renal function were
predominantly among the biomarker models.
In three of 16 clinical renal interaction models, positive
associations between higher lead measures and worse renal
function in participants in the oldest age tertile were
significantly different from associations in those in the
youngest age tertile which were in the opposite direction
- this pattern was observed at borderline significance (p < 0.1)
in 3 other models
- pattern was not observed in the EBE marker models
CdU associated with NAG.
Authors concluded that occupational lead exposure in the
moderate dose range has an adverse effect on renal function.
Inverse associations may represent hyperfiltration.
Environmental cadmium may have an adverse impact, at least
on NAG.
O
HH
H
W
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Table AX6-4.2 (cont'd). Renal Effects of Lead - Occupational Population
to
o
o
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
X
Oi
Asia (cont'd)
Weaver et al. (2003b)
Korea lead workers
1997-1999
798 lead workers with genotype information in same population
as in Weaver et al. (2003a)
79 (9.9%) participants were heterozygous for the ALAD2 allele
(none was homozygous).
89 (11.2%) had VDR genotype Bb or BB
Blood lead
31.7 ug/dL(ALADI 1)
34.2ug/dL(ALAD12)
31.6ug/dL(VDRbb)
34.8 ug/dL (VDR Bb or BB)
Tibia Lead
37.5 |lg/g(ALADI 1)
31.4ug/g(ALAD12)
37.1 ug/g(VDRbb)
38.1 ug/g (VDR Bb or BB)
Data were analyzed to determine whether polymorphisms in
the genes encoding 8-aminolevulinic acid dehydratase
(ALAD), endothelial nitric oxide synthase (eNOS), and the
vitamin D receptor (VDR) were associated with renal
outcomes or modified relations of lead exposure and dose
measures with renal outcomes.
After adjustment, participants with the ALAD2 allele had
lower mean serum creatinine and higher calculated creatinine
clearance. Effect modification by ALAD on associations
between blood lead and/or DMSA-chelatable lead and three of
six renal outcomes was observed. Among those with the
ALAD 12 genotype, higher lead measures were associated
with lower BUN and serum creatinine and higher calculated
creatinine clearance.
O
2
O
H
O
c
o
H
W
O
No significant differences were seen in renal outcomes by
VDR genotype nor was consistent effect modification
observed.
O
HH
H
W
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Table AX6-4.2 (cont'd). Renal Effects of Lead - Occupational Population
to
o
o
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
X
Oi
O
2
O
H
O
c
o
H
W
O
^
O
HH
H
W
Asia (cont'd)
Weaver et al. (2005a)
Korea
1997-1999
N = 803 current and former lead workers; 164 females
Serum Uric acid
4.8 mg/dL
Other renal outcomes as listed in Weaver et al. 2003a
Multiple linear regression
Interaction models that assessed effect modification by age in
tertiles
Blood lead
32.0 ug/dL
Tibia Lead
37.2 (40.4) ug/g bone mineral
DMSA-chelatable lead
767.8 ug/g creatinine
Work to address whether one mechanism for lead-related
nephrotoxicity, even at current lower levels of lead exposure, is via
increasing serum uric acid. Assessed 1) whether lead dose was
associated with uric acid and 2) whether previously reported
associations between lead dose and renal outcomes (Weaver et al.,
2003) were altered after adjustment for uric acid.
After adjustment for age, gender, body mass index, and alcohol use,
lead biomarkers not associated with uric acid in all participants.
However, in interaction models, both blood and tibia lead were
significantly associated in participants in the oldest age tertile (P
coefficient and 95% CI: 0.0111 (0.003, 0.019) and 0.0036 (0.0001,
0.007) for blood and tibia lead, respectively). These models were
further adjusted for blood pressure and renal function.
Hypertension and renal dysfunction are known to increase uric acid.
However, they are also risks associated with lead exposure.
Therefore, adjustment for these variables in models of associations
between lead dose and uric acid likely results in over-control. On
the other hand, since non-lead related factors contribute to both
renal dysfunction and elevated blood pressure, lack of adjustment
likely results in residual confounding. Therefore, as expected,
associations between lead dose and uric acid decreased after
adjustment for systolic blood pressure and serum creatinine,
although blood lead remained borderline significantly associated (P
(95% CI) = 0.0071 (-0.001, 0.015). However, when the population
was restricted to the oldest tertile of workers with serum creatinine
greater than the median (0.86 mg/dL), likely the highest risk
segment of the population, blood lead remained significantly
associated with uric acid even after adjustment for systolic blood
pressure and serum creatinine (P = 0.0156)
Next, in models of renal function in all workers, uric acid was
significantly (p < 0.05) associated with all renal outcomes except
NAG.
In models in the oldest tertile of workers (266 workers, median age
51.1 years, range 46.0 to 64.8 years), after adjustment for uric acid,
associations between lead dose and NAG were unchanged, but
fewer of the previously significant (p < 0.05) associations noted
between lead dose and the clinical renal outcomes in Weaver et al.
(2003a) remained significant.
-------
Table AX6-4.2 (cont'd). Renal Effects of Lead - Occupational Population
to
o
o
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
X
Oi
O
2
O
H
O
c
o
H
W
O
^
O
HH
H
W
Asia (cont'd)
Weaver et al. (2005b)
South Korea
1999-2001
N = 652 lead workers including 149 females and 200 former
workers
Patella lead measured in the third evaluation of the same study
reported in Weaver et al. (2003a). Data collection performed a
mean of 2.2 years after collection of the data presented in Weaver
et al. (2003a).
Same renal outcomes as Weaver et al. (2003a)
Serum Creatinine:
0.87mg/dL
Calculated creatinine clearance
97.0 mL/min
Multiple linear regression, adjusting for age, gender, BMI, work
status (current vs. former worker), HTN or blood pressure
(depending on model), diabetes, smoking status, and, for the
clinical measures, use of analgesics
Interaction models assessed effect modification by age,
dichotomized at the 67th percentile
Mean blood lead
30.9 ug/dL
Mean Tibia Lead
33.6 ug/g bone mineral
Mean Patella Lead
75.1 ug/g bone mineral
Mean DMSA-chelatable lead
0.63 |lg Pb/mg creatinine
All 4 lead measures were correlated (Spearman's r = 0.51 - 0.76).
Patella, blood and DMSA-chelatable lead levels positively
associated with NAG
Higher DMSA-chelatable lead associated with lower serum
creatinine and higher calculated creatinine clearance
Interaction models
All four lead measures associated with higher NAG among
participants in oldest age tertile
Higher blood, tibia, and patella lead associated with higher serum
creatinine among older participants
-beta coefficients less in the lead workers whose ages were in the
younger two-thirds of the age range; difference between slopes in
the two age groups was statistically significant only for association
of blood lead and serum creatinine
Inverse DMSA associations (higher DMSA-chelatable lead
associated with lower serum creatinine and higher calculated
creatinine clearance) significant in younger workers
Patella lead associations were consistent with those of blood and
tibia lead; DMSA-chelatable lead associations unique.
Authors hypothesized that similarities between patella, blood, and
tibia lead associations could be due, in part, to high correlations
among the lead biomarkers in this population. Despite similar high
correlations, DMSA-chelatable lead associations with serum
creatinine and calculated creatinine clearance were unique. This
biomarker is dependent on renal function and the collection time
was only 4 h. Therefore, the amount of lead that is excreted in this
relatively short time period after chelation may be influenced not
only by bioavailable lead burden, but also by high-normal as well
as actual supranormal glomerular filtration which are more
common in the younger workers.
-------
Table AX6-4.2 (cont'd). Renal Effects of Lead - Occupational Population
to
o
ON
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
Asia (cont'd)
Weaver et al. (2005c)
Korea
1997-1999
798 current and former lead workers.
same population as in Weaver et al. (2003a,b)
Data were analyzed to determine whether polymorphisms in
the genes encoding 8-aminolevulinic acid dehydratase
(ALAD), endothelial nitric oxide synthase (eNOS), and the
vitamin D receptor (VDR) were associated with uric acid or
modified relations of lead exposure and dose measures with
uric acid.
X
ON
O
2
O
H
O
c
o
H
W
O
^
O
HH
H
W
Ye et al. (2003)
Chinese lead workers
Study date not provided
216 lead workers
Renal outcomes = urinary NAG and albumin
Geometric mean blood lead
37.8 (ig/dL (n = 14 workers with
the ALAD12 genotype)
32.4 |lg/dL (n = 212 workers with
the ALAD 11 genotype)
31.9 ug/dL (VDR bb)
41.7 ug/dL (in 20 participants
with VDR Bb or BB)
Uric acid not different by ALAD or VDR genotype. Among
older workers (age > median of 40.6 years), ALAD genotype
modified associations between lead dose and uric acid levels.
Higher lead dose was significantly associated with higher uric
acid in workers with the ALAD1 1 genotype; associations
were in the opposite direction in participants with the variant
ALAD12 genotype.
After adjustment for age, NAG was borderline higher in those
with the ALAD variant allele whose blood lead levels were
>40 |lg/dL (p = 0.06). In all lead workers, after adjustment
for age, gender, smoking and alcohol ingestion, a statistically
significant positive association between blood lead and
creatinine adjusted NAG was observed in the workers with
the ALAD 12 genotype but not in lead workers with the
ALAD 11 genotype (the groups were analyzed separately
rather than in an interaction model).
No effect modification by VDR genotype on associations
between blood lead and urinary albumin and NAG observed
(separate analysis reduced power).
Middle East
Al-Neamy et al. (2001)
United Arab Emirates
Feb-June, 1999
100 "industrial" workers exposed in a range of industries
100 working controls
Blood lead
77.5 ug/dL (workers)
19.8 jig/dL (controls)
Mean BUN and serum creatinine not statistically different
between exposed workers and controls
matched for age, sex, and nationality
Renal Outcomes = BUN, serum creatinine
Limitations = data analysis
-------
Table AX6-4.2 (cont'd). Renal Effects of Lead - Occupational Population
to
o
o
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
X
Oi
O
2
O
H
O
c
o
H
W
O
^
O
HH
H
W
Middle East (cont'd)
Ehrlich et al. (1998)
South Africa
Study date not provided
El-Safty et al. (2004)
Egypt
Study date not provided
382 lead battery factory workers
Mean age = 41.2 years
All males
Multiple linear regression adjusted for age, weight, and height
(Covariates assessed for inclusion also included smoking, alcohol
ingestion, and diabetes)
Clinical renal outcomes included serum creatinine, uric acid,
and BUN.
Mean serum creatinine
1.13mg/dL
Renal early biological effect markers (NAG, RBP, intestinal
alkaline phosphatase, tissue nonspecific alkaline phosphatase,
Tamm-Horsfall glycoprotein, epidermal growth factor, and
microalbuminuria) were measured in 199 participants randomly
selected by tertiles of current blood lead.
45 lead workers with lead job duration <20 years
36 lead workers with lead job duration >20 years
75 control workers
Renal outcomes = urinary oCi-microglobulin, NAG, and
glutathione S-transferase
Mean blood lead
53.5 ug/dL
Mean exposure duration 11.6 years
Mean cumulative blood lead
(defined as sum of the average
blood lead in each year over all
years of employment; done in
subset of 246 with past blood lead
data)
579.0 (ug x yr)/dL
Mean historical blood lead (defined
as cumulative blood lead divided
by years of exposure)
57.3 ug/dL
Mean tibia lead
69.7 |lg/g bone mineral (measured
2 years after initial study on
random sample of 40)
Median urine lead
Ranged from 15.4 ug/g creatinine
in nonsmoking control workers to
250.4 ug/g creatinine in smoking
lead workers with > 20 years lead
job duration
After adjustment for age, weight, and height, categorical
current and historical blood lead and zinc protoporphyrin
were associated with serum creatinine and uric acid, in
separate models. Associations between cumulative blood
lead or exposure duration and the renal outcomes were not
observed.
Among the EBE markers, only current blood lead was
borderline associated with NAG (p = 0.09).
Associations with renal dysfunction were observed at blood
lead levels <40 |lg/dL. Not explained by an effect on blood
pressure since lead measures not associated with blood
pressure. Blood cadmium measured in 56 participants
2 years after the initial study. All low (< 1.2 |lg/L)
suggesting that occupational level cadmium exposure was
not a contributing factor. The authors did implicate lead
body burden which was substantial based on mean tibia lead.
However, cumulative blood lead was not associated in this
study and mean tibia lead in Roels et al. (1994) was similar
(in that study a positive association with creatinine clearance
was observed).
Medians of all 3 renal outcomes significantly higher in lead
workers irregardless of smoking status (analysis stratified by
smoking status).
Urine lead significantly correlated with urinary tti-
microglobulin and glutathione S-transferase in nonsmoking
lead workers and with NAG as well in smoking lead
workers.
Limitations include using urine lead as sole lead dose
measure and data analysis.
-------
Table AX6-4.2 (cont'd). Renal Effects of Lead - Occupational Population
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
Middle East (cont'd)
Mortadaet al. (2001)
Egypt
Study date not provided
43 traffic policemen
52 matched control office workers (similar in terms of age,
gender, smoking, and "social life").
Renal outcomes = serum creatinine, beta-2 microglobulin, BUN
and urinary (3-2- microglobulin, NAG, alkaline phosphatase,
y-glutamyl transferase, and albumin.
Exclusionary criteria included diabetes, HTN, hepatic, renal or
urologic diseases.
Blood lead
32.1 ug/dL (exposed)
12.4 ug/dL (controls)
Lead also measured in hair, urine
and nails
NAG and albumin significantly higher in policemen
compared to controls. NAG positively correlated
(Pearson's) with job duration and blood and nail lead.
Urinary albumin positively correlated with job duration and
blood and hair lead.
Limitations: data analysis - no adjustment, use of parametric
correlation techniques with data likely to be nonparametric;
study size
O
2
O
H
O
c
O
H
W
O
V
O
HH
H
W
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Table AX6-4.3. Renal Effects of Lead - Patient Population
to
o
o
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
X
Oi
United States
Osterloh et al. (1989)
Northern CA
Study date not provided
Steenland et al. (1990)
Michigan
Diagnosis from
1976-1984
40 male subjects with hypertensive nephropathy (hypertension
preceded renal insufficiency; serum creatinine 1.8-4 mg/dL)
24 controls with renal dysfunction from other causes
Patients recruited from the Kaiser Permanente Regional
Laboratory database (large health maintenance organization) in
northern California
325 men with ESRD (diabetes, congenital and obstructive
nephropathies excluded)
controls by random digit dialing, matched by age, race, and place
of residence.
Mean blood lead
7.3 ug/dL (in both hypertensive
nephropathy and controls CRI
from other causes)
Mean EDTA chelatable lead levels
153.3 ug/72 hours (hypertensive
nephropathy)
126.4 jig/72 hours (control CRI)
No significant difference in EDTA chelatable lead levels;
highest chelatable lead level was 609.2 ug/72 hours.
Lead dose and serum creatinine were not correlated.
Blood and chelatable lead levels much lower than those
reported by Wedeen et al. (1983) and Sanchez-Fructuoso
et al. (1996).
Only 17% of their study participants had a history of
possible lead exposure based on questionnaire, again much
lower than the two other studies.
Risk of ESRD significantly related to moonshine alcohol
consumption (OR = 2.43), as well as analgesic consumption,
family history of renal disease, and occupational exposure to
silica or solvents.
O
2
O
H
O
c
o
H
W
O
V
O
HH
H
W
Europe
Behringer et al. (1986)
Germany
Study date not provided
Colleoni and D'Amico
(1986)
Italy
(-1982-1985)
16 patients with CRI (median serum creatinine = 2.2 mg/dL) and
gout
19 patients with CRI (median serum creatinine = 5.1 mg/dL)
without gout
21 healthy controls
Lead excretion in the 96 hours after administration of 1 g EDTA
iv
12 consecutive patients with CRI (mean serum
creatinine = 3.3 mg/dL) and gout, renal diagnosis consistent with
chronic interstitial nephritis in all; 7 had history of occupational
lead exposure
12 controls with chronic glomerulonephritis and no history of
lead exposure or gout
Lead excretion in the 48 hours after administration of 1.5 g
EDTA im
Median blood lead
7.2 ug/dL (controls)
11.5 ug/dL (CRI, no gout)
15.3 ug/dL (CRI & gout)
Median EDTA chelatable lead
(ug/4 days/1.73m2)
63.4 (controls)
175.9 (CRI, no gout)
261.3 (CRI & gout)
Mean EDTA chelatable lead
(ug/48 hrs)
180 (CRI, no gout)
505 (CRI & gout)
EDTA chelatable lead higher in gout patients who developed
gout after CRI than those in which gout preceded CRI
(statistical test results not mentioned or shown). Authors
conclude a role for lead in patients with gout occurring in
setting of CRI and that lead may contributes to renal function
decline in established renal disease from other causes.
Limitations = small groups, limited data analysis
Significantly higher EDTA chelatable lead in the group with
CRI and gout compared to CRI alone. EDTA chelatable
lead significantly correlated with serum creatinine in patients
with CRI and gout but not CRI alone. Authors conclude that
lead is cause of CRI with gout but renal insufficiency alone
not responsible for increased lead body burden (absence of
evidence for reverse causation).
Limitations = small sample size, limited data analysis
-------
Table AX6-4.3 (cont'd). Renal Effects of Lead - Patient Population
to
o
o
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
Europe (cont'd)
Colleoni et al. (1993)
Italy
Study date not provided
All 115 patients on hemodialysis at the time of the study; 41 Mean blood lead
women (corrected for hemoglobin)
Blood lead data from prior study of 383 healthy controls in same 19.9 ug/dL (patients)
geographical area served as comparison 14.7 ug/dL (controls)
Significantly higher mean blood lead in hemodialysis
patients compared to healthy controls. 13% had blood lead
levels >30 ug/dL. Blood lead level was not associated with
duration of hemodialysis. Mean lead levels higher in
smokers and in relation to alcohol ingestion. Lead not
detectable in dialysis fluids.
Limited data analysis
O
o
2
o
H
O
o
HH
H
W
Craswell et al. (1987)
Germany and Australia
Study date not provided
Fontanellas et al. (2002)
Spain
Study date not provided
Jones et al. (1990)
Study location and date
not provided; authors
from UK
See discussion below under Australia
ALAD/restored ALAD as a possible index of lead poisoning in
chronic renal failure patients.
27 dialysis patients
59 healthy controls
Mean blood lead
8.1 ug/dL (patients)
10.0 ug/dL (controls)
Restored ALAD was measured after the addition of zinc
and dithiothreitol (DTT) to the incubation media.
The ALAD/restored ALAD ratio was found to correlate
with the results of the EDTA lead mobilization test.
Patients excreting 1,115 to 3860 ug lead per 72 hours had a
ratio of 0.19 while chronic renal failure patients excreting
an average of 322 ug lead (range 195 to 393) had a ratio of
0.47. In comparison, normal controls had a ratio of 0.5.
Tibia lead levels not correlated with blood lead but were
correlated with lead in bone biopsy measurements
(r = 0.42).
Limitations = data analysis
Kosteretal. (1989)
Study location and date
not provided; authors
from Germany
91 patients with CRI ( median serum creatinine = 2.5 mg/dL)
46 age-matched normal controls.
Lead excretion in the 4 days after
1 g EDTA iv
Mean Blood lead
(corrected for hemoglobin)
11.2 ug/dL (patients)
7.6 ug/dL (controls)
EDTA chelatable lead
164.7 ug/4 days /1.73 m2 (patients)
63.6 ug/4 days /1.73 m2 (controls)
CRI patients had significantly higher blood and EDTA
chelatable lead levels than controls. In 13% of the CRI
patients, EDTA chelatable lead exceeded the highest value
in controls (328.8 ug). EDTA chelatable lead levels were
correlated with serum creatinine in patients (r = 0.37;
p < 0.007).
Limitations = data analysis
-------
Table AX6-4.3 (cont'd). Renal Effects of Lead - Patient Population
to
o
o
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
Europe (cont'd)
Miranda-Carus et al.
(1997)
Spain
1990-1994
27 patients with gout and CRI
50 patients with gout only
26 controls with normal renal function and no gout
Multiple purine metabolism measures including serum urate,
hypoxanthine, and xanthine, as well as their excretion, clearance
and fractional excretion measures
Mean blood lead
17.8 ug/dL (gout & CRI)
14.9 |ig/dL (gout only)
12.4 |ig/dL (controls)
EDTA chelatable lead
845 ug/120 hrs (gout & CRI)
342 jig/120 hrs (gout only)
215 jig/120 hrs (controls)
Lead dose measures significantly higher in patients with gout
and CRI compared to the other two groups. EDTA
chelatable lead inversely correlated with creatinine
clearance. Each of the 2 patient groups were dichotomized
by EDTA-chelatable lead level of 600 Jig/120 hours,
resulting in 3 small groups (n ranging from 6 to 14) and one
group of 44 participants with gout and EDTA chelatable lead
below the cut-off. No significant differences in mean purine
metabolism measures were observed. It is not clear whether
correlations between EDTA-chelatable lead and the purine
measures were assessed and if so whether the small groups
were combined for this analysis. Thus lack of power may be
one reason for the inconsistency with Lin's work. Different
lead body burdens may be a factor as well.
Uric acid parameters were unchanged following chelation in
6 participants with EDTA-chelatable above 600 jig/120
hours. Again higher lead body burdens may be a factor but
the small number and limited details on the group make firm
conclusions difficult.
Nuytsetal. (1995)
Belgium
Study date not provided
Case-control study
272 cases with chronic renal failure (all types) matched to 272
controls by age, sex and residence
Exposure assessed by 3 industrial hygienists blinded to case or
control status
Significantly increased odds ratio for chronic renal failure
with lead exposure (odds ratio = 2.11 [95% CI: 1.23, 4.36])
as well as several other metals. Increased risk with diabetic
nephropathy.
-------
Table AX6-4.3 (cont'd). Renal Effects of Lead - Patient Population
to
o
o
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
O
o
2
o
H
O
o
HH
H
W
Europe (cont'd)
Sanchez-Fructuoso et al.
(1996)
Spain
Study date not provided
Van de Vyver et al.
(1988)
Belgium, France
and Germany
Study date not provided
Winterberg et al. (1991)
Study location and date
not provided; authors
from Germany
296 patients:
Group I = 30 normal control subjects
Group II = 104 patients with essential HTN & normal renal
function
Group III-A = 68 patients with HTN and CRI of uncertain
etiology but presumed nephroangiosclerosis
Group III-B = 64 patients with HTN, CRI, and gout
Group IV = 30 patients with CRI of known etiology
Transiliac bone biopsies obtained from:
11 cadavers without known lead exposure and with normal renal
function
13 patients with CRI, gout and/or HTN 22 lead workers
153 dialysis patients
Iliac crest bone lead measured by biopsy in:
8 controls
8 patients with CRI
14 dialysis patients
Mean blood and EDTA-chelatable
lead levels:
Group I
16.7 ug/dL
324 ug/72 hrs
Group II
16.8 jig/dL
487 jig/72 hrs
Group III-A
18.5 jig/dL
678 jig/72 hrs
Group III-B
21.1 ug/dL
1290 jig/72 hrs
Group IV
16.5 ug/dL
321 ug/72 hrs
Mean transiliac lead levels
5.5 ug/g (153 dialysis pts)
20.6 ug/g (in highest 5% dialysis
pts)
3.7 ug/g (in 10 pts on dialysis due
to analgesic nephropathy)
6.3 ug/g (11 cadavers)
30.1 ug/g (22 lead workers)
Mean iliac crest bone lead levels
1.63 ug/g (8 controls)
2.18 ug/g (8 patients with CRI)
3.59 ug/g (in 14 dialysis pts)
EDTA chelatable lead >600 ug/72 hrs in 16 patients in group
II, 30 patients in group III-A, 44 patients in group III-B, but
no patients in either group I and IV.
Mean blood and EDTA chelatable lead levels in the patients
with CRI of known cause were not statistically different
from controls with normal renal function. However, baseline
urinary lead excretion was lower in group IV. This provides
conflicting evidence regarding the "reverse causality"
hypothesis of increased lead burden due to decreased
excretion in CRI
Significant correlations noted between bone lead levels
(assessed by biopsy) and EDTA chelatable lead level in
12 patients whose chelatable lead levels were >600 ug/72
hours; provides support for validity of chelatable lead levels
in CRI.
A positive correlation was observed between serum
creatinine levels and EDTA-chelatable lead levels
>600 ug/72 hrs but not below this level.
In group III, mean measured creatinine clearance was
significantly lower in those with EDTA chelatable lead
levels >600 |lg/72 hrs compared to participants with
chelatable lead <600 |lg/72 hrs.
In 5% of the hemodialysis patients studied, bone lead
concentrations approximated the levels found in active lead
workers, suggesting lead as a primary cause of their renal
failure. Levels in the 10 patients with analgesic nephropathy
were the lowest (all <7 ug/g), evidence against reverse
causality.
In the combined group of 13 patients with CRI, gout and/or
HTN and 22 lead workers, EDTA chelatable lead correlated
with lead in bone biopsies (r = 0.87).
Noted that the bone lead levels in patients with analgesic
nephropathy and cadaver controls in Van de Vyver et al.
(1988) were much higher than in control groups of other
researchers. They reiterated the concern that lead did
accumulate due to decreased renal excretion.
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Table AX6-4.3 (cont'd). Renal Effects of Lead - Patient Population
to
o
o
O
O
2
O
H
O
o
HH
H
W
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
Latin and South America
Navarro et al. (1992)
Venezuela
Study date not provided
1 8 dialysis patients
14 controls
Bone (biopsy) and blood levels of lead and several other metals
Mean blood lead
5.2 ug/dL (patients)
11.5 ug/dL (controls)
Mean lead in bone
9.7 ug/g (patients)
7.0 ug/g (controls)
Blood but not bone lead significantly higher in patients
compared to controls. Authors concluded that bone
accumulation of aluminum, iron and vanadium, but not lead,
occurred in dialysis patients.
Limitations = sample size, data analysis including lack of
adjustment
Australia
Craswell et al. (1987)
Germany and Australia
Study date not provided
German participants from industrialized area where chronic lead
nephropathy not previously observed
Gp 1 = 8 healthy controls (from hospital staff)
Gp 2a = 12 CRI patients, no gout or lead exposure
Gp 2b = 7 CRI patients, no gout but + lead exposure
Gp 3a = 7 CRI patients with gout but no lead exposure
Gp 3b = 6 CRI patients with gout and lead exposure
Australian participants from Queensland site of known chronic
lead nephropathy
Gp 1=9 healthy controls (from hospital staff)
Gp 2a = 14 CRI patients, no gout or lead exposure
Gp 2b = 11 CRI patients, no gout but + lead exposure
Gp 3a = 25 CRI patients with gout but no lead exposure
Gp 3b = 11 CRI patients with gout and lead exposure
CRI defined as serum creatinine > 1.5 mg/dL
"excess" EDTA chelatable lead defined as lead excreted over 4
days after EDTA minus twice baseline lead excreted pre-EDTA
Median blood lead (hemoglobin
corrected)
Gp 1
German = 6.8 ug/dL
Australian =11.0 ug/dL
Gp2a
German = 6.2 ug/dL
Australian = 9.1 ug/dL
Gp2b
German = 8.5 ug/dL
Australian =16.2 ug/dL
Gp3a
German =10.6 ug/dL
Australian =12.8 ug/dL
German =12.0 ug/dL
Australian = 27.1 ug/dL
Median "excess" EDTA chelatable
lead
Gp 1
German = 68.4 jig
Australian =82.9 ug
Gp2a
German = 126.4 jig
Australian = 393.7 jig
Gp2b
German = 489.0 ug
Australian = 1181.1 jig
Gp3a
German = 227.9 jig
Australian = 808.1 ug
Gp3b
German = 422.7 ug
Australian = 1077.5 ug
Using nonparametric statistical techniques due to skewed
data, German participants excreted statistically less lead than
their Australian counterparts. Mean EDTA chelatable lead
levels were significantly higher in German patients with gout
than in those without gout; the observed increase in the
Australian patients was of borderline significance (p < 0.1).
Limitations = small groups, limited data analysis
-------
Table AX6-4.3 (cont'd). Renal Effects of Lead - Patient Population
to
o
o
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
Australia (cont'd)
Price etal. (1992)
Queensland, Australia
1981-1986
8 renal patients compared with age-matched controls
X-ray fluorescence of finger bone lead conducted twice 5 years
apart
Authors conclude that lead in bone half-life is similar in
renal patients compared to age-matched controls. Study
limitations substantial, however.
Limitations = small numbers (although bone lead measured
in more patients, many were below the limit of detection,
inclusion of outliers without formal statistical analysis.
Asia
O
o
2
o
H
O
o
HH
H
W
Lin and Lim (1992)
Chinese population
(likely in Taiwan)
Study date not provided
Lin and Huang (1994)
Taiwan
Study date not provided
10 healthy controls
10 patients with CRI but no gout
8 patients with gout and subsequent CRI
6 patients with CRI and subsequent gout
Exclusionary criteria included + history of occupational or
environmental lead exposure
Group 1 = 10 patients with normal renal function and no gout;
Group 2 = 10 patients with CRI (serum creatinine >1.4 mg/dL)
and subsequent gout; Group 3 = 20 patients with CRI but no gout
All males
Lead body burden assessed with 1 g EDTA iv followed by 72 hr
urine collection
Mean EDTA chelatable lead in
ug/72hrs/ 1.73m2
90.2 (controls)
98 (CRI, no gout)
171.6 (gout, then CRI)
359.8 (CRI, then gout)
Mean EDTA chelatable lead
Gp 1=60.55 ug/72 hrs
Gp 2 = 252.24 ug/ 72 hrs
Gp3 = 84.86 ug/72hrs
Lead body burden higher in patients with CRI and gout,
especially when CRI precedes gout.
Limitations = small sample sizes, statistical analysis
Mean EDTA chelatable lead and serum urate significantly
higher in the patients with gout. After adjustment for
creatinine clearance, log transformed EDTA chelatable lead
was significantly associated with serum urate levels
(P [95% CI: 0.757 [0.142, 1.372]; p < 0.05), daily urate
excretion (P [95% CI: -60.15 [-118.1, -2.16]); p < 0.05),
urate clearance (P [95% CI: -0.811 [-1.34, -0.282];
p < 0.05), and fractional urate excretion (P [95% CI: -1.535
[-2.723, -0.347]; p < 0.05). EDTA chelatable lead not
associated with creatinine clearance.
Lin and Lim (1994)
Taiwan
Study date not provided
Gp 1 = 12 healthy controls
Gp 2 = 10 patients with HTN
Gp 3 = 12 patients with HTN, then CRI (hypertensive
nephropathy)
Gp 4 = 12 patients with CRI only
Gp 5 = 12 patients with CRI not due to HTN, but subsequent
HTN
Mean EDTA chelatable lead
Gp 1 = 76.6 ug/ 72 hrs
Gp2 = 67.96ug/72hrs
Gp3 = 182.9ug/72hrs
Gp3 = 84.46 ug/72hrs
Gp3 = 92.86 ug/72hrs
Limitations = small sample sizes, limited adjustment in
regression analyses.
Higher mean EDTA chelatable lead level in Gp 3;
5 of 12 had history of gout developing after CRI
Limitations = small sample sizes, limited analyses
-------
Table AX6-4.3 (cont'd). Renal Effects of Lead - Patient Population
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o
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
Asia (cont'd)
Lin et al. (1999)
Taiwan
Study date not provided
32 patients selected from 102 patients with serum creatinine from
1.5 -4.0 mg/dL who were followed in the Institution's outpatient
clinics
Eligibility criteria included serum creatinine from 1.5 — 4.0
mg/dL, stable renal function over 6 months before study entry;
controlled blood pressure and cholesterol; daily protein intake
<1 g/kg body wt; no known history of exposure to lead or other
heavy metals and EDTA chelatable lead >150 but <600 |lg/72
hour.
Exclusionary criteria included potentially reversible or unstable
renal disease (i.e., due to systemic diseases such as lupus and
diabetes), and nephrotoxicant medications.
Patients divided into 16 patients receiving 1 g EDTA i.v. weekly
for two months and a control group of 16 patients who received
no therapy
Mean EDTA chelatable lead levels
pre-chelation
254.9 |lg/ 72 hrs in group
receiving subsequent chelation
279.7 |lg/ 72 hrs in control group
Blood lead levels not mentioned
Rates of progression of renal insufficiency were followed by
reciprocal of serum creatinine during the 12 months prior to
therapy and for 12 months following therapy. Rates of
progression of renal insufficiency were similar in the
treatment group and the control group during the baseline
observation. However, improvement in renal function was
observed during EDTA chelation. Following chelation,
renal function stabilized in the treated group but continued to
decline in the control group. At 12 months after treatment,
the mean difference in the change in the reciprocal of serum
creatinine between the two groups was 0.000042 L/|lmol per
month (95% CI: 0.00001,0.00007). Results using a
sensitivity analysis for patients lost to follow-up (only one in
each group) gave similar results.
-------
Table AX6-4.3 (cont'd). Renal Effects of Lead - Patient Population
to
o
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Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
Asia (cont'd)
Linetal. (200la)
Taiwan
Study date not provided
24 month prospective observational study
110 patients with CRI dichotomized by EDTA chelatable lead
level of 80 ug / 72 hrs into two groups of 55 each
Eligibility criteria included serum creatinine from 1.5 — 4.0
mg/dL, stable renal function (decrease in GFR <5 mL/min over 6
months); blood pressure <140/90 mmHg; cholesterol level <240
mg/dL; daily protein intake <1 g/kg body wt; no known history of
exposure to lead or other heavy metals and EDTA chelatable lead
<600 Hg/72 hour.
Exclusionary criteria included potentially reversible or unstable
renal disease (i.e., due to systemic diseases such as lupus and
diabetes), nephrotoxicant medications, and drug allergies.
196 patients initially screened for study; details on reasons for
non-eligibility not provided.
Primary outcome = 1.5 times increase in the initial creatinine
level or need for dialysis; secondary outcome = change in
creatinine clearance
Cox proportional-hazards model analysis for primary outcome.
Mean differences in creatinine clearance compared at sequential
time points with t or Mann-Whitney U tests.
Adjustment for age, gender, baseline BMI, smoking, proteinuria,
hypertension, hyperlipidemia, daily protein intake, and underlying
renal disease
Intention-to-Treat and sensitivity analyses compared creatinine
clearance a by time period in high and low lead groups.
3 month clinical trial of chelation with 1 year follow-up
At 24 months, 36 patients whose EDTA chelatable lead levels
were 80 - 600 |lg/72 hours and serum creatinine levels of
<4.2 mg/dL were randomized; 24 to a 3-month treatment period
consisting of weekly chelation with 1 g EDTA iv until their
excreted lead levels fell below 80 |lg/72 hours and 12 to placebo
infusion.
Intention-to-Treat and sensitivity analyses compared creatinine
clearance by time period in treated and control groups.
Mean blood lead levels
6.6 ug/dL in high normal lead
body burden group (n = 55)
3.9 ug/dL in low normal lead body
burden group (n = 55)
Mean EDTA chelatable lead levels
pre-chelation
182.9 ug/ 72 hrs in high normal
lead body burden group (n = 55)
37.9 jig/ 72 hrs in low normal lead
body burden group
(n = 55)
24 month prospective observational study
Lead dose measures were only significant differences
between high and low normal lead body burden groups. Of
the 96 participants who completed the observation study, 14
patients in the high normal body lead burden group reached
the primary endpoint compared to 1 patient in the low body
lead burden group (p < 0.001 by log-rank test).
From month 12 to month 24, creatinine clearance in high
normal body lead burden patients was at least borderline
statistically lower than in low body lead burden patients;
from 18-24 months, 95% CI excluded 0. 95% CI for the
difference at 24 months was (-15.0, -3.8); difference in
creatinine clearance between groups was 0.15 mL/s at that
point.
In a Cox multivariate regression analysis, chelatable lead
was significantly associated with overall risk for the primary
endpoint (relative risk = 41.5 [95% CI: 3.9, 440.8];
p = 0.002]). In this model, age, basal BMI, and basal daily
proteinuria were also associated with increased risk.
3 month clinical trial of chelation with 1 year follow-up
The two groups were similar in baseline renal risk factors
(although numbers small so beta error possible).
Mean EDTA dose during the 3 month period was 5 ug.
After three months of lead chelation therapy, the body lead
burden of the patients in the chelation group decreased from
198 to 39.2 ug. After 3 months of chelation and 3 months of
follow-up, creatinine clearance increased by 0.08 mL/s in the
treated group but declined by 0.04 mL/s in the controls.
At the end of the study period, mean creatinine clearance
was 0.68 mL/s in the chelated group compared to 0.48 mL/s
in the control group (p < 0.05; 95% CI for the difference
between groups = -25.0 to -0.2).
-------
Table AX6-4.3 (cont'd). Renal Effects of Lead - Patient Population
to
o
o
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
O
o
2
o
H
O
o
HH
H
W
Asia (cont'd)
Linetal. (200 Ib)
Study location and date
not provided; authors
from Taiwan
Lin et al. (2002)
Study location and date
not provided; authors
from Taiwan
101 patients with CRI (defined as serum creatinine between 1.5
and 3.0 mg/dL)
67 with CRI and gout
34 with CRI only
Eligibility criteria included no known history of lead exposure,
certain diagnoses and medications. CRI must have preceded gout
Randomized chelation trial
30 participants with CRI, gout, and EDTA-chelatable lead levels
between 80.2 and 361 |lg/72 hours randomized to either a
treatment group receiving 1 gram EDTA iv per week for 4 weeks
(N = 20) or to a control group who received glucose in normal
saline iv.
84 healthy participants
27 participants with gout
All with normal renal function (defined as serum creatinine
<1.4 mg/dL)
Participants with a history of occupational heavy metal exposure,
EDTA-chelatable lead levels >600 |lg/72 hours, or systemic
diseases were excluded.
Randomized chelation trial
24 participants with EDTA-chelatable lead levels between 75 and
600 Hg/72 hours randomized to either a treatment group receiving
1 gram EDTA iv per week for 4 weeks (N = 12) or to a control
group who received glucose in normal saline i.v.
Multiple linear regression, adjustment for age, sex, BMI,
daily protein intake, and creatinine clearance.
Mean blood lead
5.4 ug/dL (CRI and gout)
4.4 ug/dL (CRI only)
Mean EDTA-chelatable lead
138.1 ug/ 72 hrs (CRI and gout)
64.2 ug/ 72 hrs (CRI only)
(p<0.01)
Mean blood lead
3.9 ug/dL (controls)
4.2 ug/dL (gout)
Mean EDTA-chelatable lead
45 ug/ 72 hrs (controls)
84 jig/ 72 hrs (gout) (p < 0.0001)
In 101, EDTA-chelatable lead higher in patients with CRI
and gout compared to those with CRI only.
EDTA-chelatable lead, but not blood lead, was associated
positively with serum urate and negatively with daily urate
excretion, urate clearance, and fractional urate excretion.
Randomized chelation trial
The two groups had similar uric acid, renal function, and
lead measures pre-chelation. In the treated group, mean
EDTA-chelatable lead declined from 159.2 to 41 |lg/72
hours; mean serum urate decreased from 10.2 to 8.6 mg/dL
(% change compared to the control group = -22.4; [95% CI:
-46.0, -1.5]; p = 0.02), and mean urate clearance increased
from 2.7 to 4.2 mL/min ((% change compared to the control
group = 67.9; [95% CI: 12.2, 121.2]; p < 0.01). Daily and
fractional urate excretion were also significantly different
between the two groups. Mean measured creatinine
clearance increased from 50.8 to 54.2 mL/min (% change
compared to the control group = 8.0; [95% CI: -0.4,20.1];
p = 0.06).
Significantly higher mean EDTA-chelatable lead and lower
urate clearance were present in patients with gout compared
to those without (3.7 versus 6.0 mL/min/1.73 m2; p < 0.001
for urate clearance)
After adjustment, EDTA-chelatable lead associated with all
four uric acid measures (serum urate, daily urate excretion,
urate clearance, and fractional urate excretion). Blood lead
associated with serum urate. All associations in same
direction as in Lin et al. (2001).
Randomized chelation trial.
The two groups had similar urate, renal function, and lead
measures pre-chelation. In the treated group, mean blood
and EDTA-chelatable lead levels declined (from 5.0 to 3.7
|lg/dL and 110 to 46 |lg/72 hours, respectively). Statistically
significant improvement observed in all four urate measures
in the treated group compared to the control group.
-------
Table AX6-4.3 (cont'd). Renal Effects of Lead - Patient Population
to
o
o
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
Asia (cont'd)
Lin et al. (2003)
Study location and date
not provided; authors
from Taiwan
24 month prospective observational study
202 patients with CRI
Eligibility criteria included serum creatinine from 1.5 - 3.9
mg/dL, stable renal function (decrease in GFR <5 mL/min over 6
months); blood pressure <140/90 mmHg; cholesterol level <240
mg/dL; daily protein intake <1 g/kg body wt; no known history of
exposure to lead or other heavy metals and EDTA chelatable lead
<600 ng/72 hour.
Exclusionary criteria included potentially reversible or unstable
renal disease (i.e., due to systemic diseases such as lupus and
diabetes), nephrotoxicant medications, and drug allergies.
250 patients initially observed, loss due to noncompliance or
unstable renal function, baseline data on the 48 who left or were
removed from the study not provided.
Cox proportional-hazards model analysis for primary outcome.
Generalized estimating equations (GEE) for associations between
baseline chelatable lead or blood lead level and longitudinal
change in GFR (estimated by an MDRD equation [Levey et al.,
1999]) and by measurement of creatinine clearance.
Adjustment for age, gender, baseline BMI, smoking, baseline
serum creatinine, proteinuria, hypertension, hyperlipidemia, daily
protein intake, and underlying renal diseases.
Mean blood lead levels
5.3 ug/dL in total group
(n = 202)
6.1 ug/dL pre-chelation in
chelated group (n = 32)
5.9 ug/dL pre-chelation in control
group
Mean EDTA chelatable lead levels
pre-chelation
104.5 ug/72 hrs in total group
(n = 202)
150.9 ug/72 hrs pre-chelation in
chelated group
144.5 ug/72 hrs pre-chelation in
control group
24 month prospective observational study
Primary endpoint = increase in serum creatinine to 1.5 times
baseline or need for hemodialysis; occurred in 24
participants. Secondary endpoint = change in estimated
glomerular filtration rate (GFR)
In a Cox multivariate regression analysis, chelatable lead
was significantly associated with overall risk for the primary
endpoint (hazard ratio for each 1 |lg chelatable lead was 1.00
[95% CI: 1.00, 1.01]; p = 0.03). In this model, baseline
serum creatinine was also associated (hazard ratio for each
1 mg/dL was 2.75 [95% CI: 1.46, 5.18]; p = 0.002) and, at
borderline significance (p < 0.1), baseline daily protein
excretion and smoking were as well.
The association between baseline chelatable lead and change
in GFR was modeled using GEE. Estimate = -0.003
(p = <0.001) (neither SE nor CI provided). In this model,
gender and daily protein intake were associated with
increased GFR; baseline serum creatinine level, daily urinary
protein excretion, and the presence of polycystic kidney
disease were significant predictors of a progressive decline
in glomerular filtration rate.
Based on this model, a 10 |lg higher baseline chelatable lead
level was associated with a GFR decrease of 0.03 mL per
minute per 1.73 m2 of body-surface area during the 2 year
observation period. Although statistically significant, this
effect is clinically small. Furthermore, it is 40 fold lower
than that reported in Yu et al. (2004) over a follow-up period
that is only two-fold shorter.
-------
Table AX6-4.3 (cont'd). Renal Effects of Lead - Patient Population
to
o
o
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
Asia (cont'd)
Lin et al. (2003) (cont'd)
Study location and date
not provided; authors
from Taiwan
27 month clinical trial of chelation
At 24 months, 64 patients whose EDTA chelatable lead levels
were 80 - 600 |lg/72 hours and serum creatinine levels of <4.2
mg/dL were randomized; half to a 3-month treatment period
consisting of weekly chelation with 1 g EDTA iv until their
excreted lead levels fell below 60 |lg/72 hours and half to five
weeks of placebo infusion.
Intention-to-Treat analysis compared creatinine clearance and
GFR by time period in treated and control groups
27 month clinical trial of chelation
The two groups were similar in baseline renal risk factors
(although numbers small so beta error possible).
After three months of lead chelation therapy, the body lead
burden of the patients in the chelation group decreased from
150.9 to 43.2 ug and their mean blood lead levels decreased
from 6.1 to 3.9 ug/dL. GFR increased by 3.4 mL/min/1.73
m2 in the treated group; in contrast, it decreased by 1.1
mL/min/1.73 m2 in the control group. Mean EDTA dose
during the 3 month period was 5.2 ug.
In the subsequent 24 months, chelation in 19 (59%)
participants was repeated due to increases in serum
creatinine in association with rebound increases in EDTA
chelatable lead levels. Each received one additional
chelation series (mean = 4.1 g EDTA) a mean of 13.7
months after the first chelation period. Control patients
receiving placebo weekly for five weeks every six months.
At the end of the study period, mean estimated glomerular
filtration rate increased by 2.1 mL/min/1.73m ofbody-
surface area in the chelated group compared to a decline of
6.0 in the controls (p < 0.01; 95% CI for the difference
between groups = -11.0 to -5.1).
-------
Table AX6-4.3 (cont'd). Renal Effects of Lead - Patient Population
to
o
o
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
O
o
2
o
H
O
o
HH
H
W
Asia (cont'd)
Yu et al. (2004)
Study location and date
not provided; authors
from Taiwan
121 patients followed over a four year observational period
Eligibility criteria included serum creatinine from 1.5 -3.9
mg/dL, stable renal function (decrease in GFR <5 mL/min
over 6 months); blood pressure < 140/90 mmHg; cholesterol
level <240 mg/dL; daily protein intake <1 g/kg body wt; no
known history of exposure to lead or other heavy metals and
EDTA chelatable lead <600 |lg/72 hour.
Exclusionary criteria included potentially reversible or
unstable renal disease (i.e., due to systemic diseases such as
lupus and diabetes), medical noncompliance (patients were
followed for 6 months to assess compliance before
enrollment in the study), nephrotoxicant medications, and
drug allergies.
Cox proportional hazards model analysis for primary
outcomes and generalized estimating equations (GEE) for
associations between baseline chelatable lead or blood lead
level and longitudinal change in GFR (estimated by an
MDRD equation [Levey et al., 1999])
Adjustment for age, gender, baseline BMI, smoking, baseline
serum creatinine, proteinuria, hypertension, hyperlipidemia,
daily protein intake, use of ACE inhibitor or angiotensin-
receptor antagonists (since not all patients were on these),
and chronic glomerulonephritis (other underlying renal
diseases included in GEE as well)
Mean (SD) blood lead at baseline
3.4 (1.3) |lg/dL in 58 patients with
"low-normal" EDTA chelatable lead
levels (<80 |lg lead/72 hours)
4.9 (2.6) ng/dL in 63 patients with
"high-normal" EDTA chelatable lead
levels (>80 but <600 |lg/72 hours)
The two groups (dichotomized by diagnostic EDTA chelatable
lead of 80 |lg lead/72 hours ) were similar in most baseline risk
factors other than lead body burden. Borderline statistically
significant (p < 0.1) differences included mean older age in the
high chelatable lead group and certain renal diagnoses (chronic
glomerulosclerosis, chronic interstitial nephritis, hypertensive
nephropathy; surprisingly both of the latter two diagnoses were
less common in the lower lead body burden group).
Fifteen patients in the "high-normal" chelatable lead group
reached the primary endpoint (doubling of serum creatinine
over the 4 year study period or need for hemodialysis)
compared to only two in the "low-normal" group (p = 0.001 by
Kaplan-Meier analysis).
In a Cox multivariate regression analysis, chelatable lead was
significantly associated with overall risk for the primary
endpoint (hazard ratio for each 1 |lg chelatable lead was 1.01
[95% CI: 1.00-1.01; p = 0.002]). In this model, the only other
variable reaching at least borderline significance (p < 0.1) was
baseline serum creatinine.
The associations between baseline chelatable lead or blood lead
level and change in GFR were modeled separately using GEE.
Estimates =
-0.1295 (p = 0.002) for lead body burden (neither SE nor CI
provided)
-4.0123 (p = 0.02) for blood lead (neither SE nor CI provided)
Based on these models, a 10 |lg higher baseline chelatable lead
level or l|lg/dL higher blood lead level predicted 1.3 and 4.0
mL/min declines in GFR, respectively, during the four year
study period. Similar to the primary outcome analysis, of the
many traditional renal risk factors adjusted for in these models,
only diagnosis of chronic interstitial nephritis was significantly
associated, in this case with an increase in GFR. Of note,
chronic interstitial nephritis was also a more frequent diagnosis
in the group with the low-normal chelatable lead levels
(p = 0.09).
The authors stated that these patients were not included in
earlier publications (which are described below in Section
6.4.4.3.3 Therapeutic EDTA Chelation in Patients).
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Table AX6-4.4. Renal Effects of Lead - Mortality
to
o
o
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
O
o
2
o
H
O
o
HH
H
W
United States
Cooper (1988);
Cooper, Wong and
Kheifets (1985)
16 U.S. plants
Employment between
1946 and 1970; mortality
from 1947 to 1980
Steenland et al. (1992)
Idaho
Employed between 1940
and 1965; mortality up to
1988
4519 male battery plant workers
2300 male lead production workers
Employed for at least one year between 1946 and 1970
Mean blood lead
63 ug/dL in n = 1326 battery
workers
80 ug/dL in n = 537 production
workers
Past lead exposures poorly
Cause of death per death certificate (extrapolated when missing) documented prior to 1960
Standardized mortality ratios (SMRs) compared with national
age-specific rates. PMR also assessed
Analyzed separately by battery and lead production, by hire date
before and after 1/1/1946, and by cumulative years of
employment
(1-9, 10-19, 20+)
1990 male lead smelter workers
employed in a lead-exposed department for at least one year
between 1940 and 1965
Vital status was determined using records from the Social
Security Administration and the National Death Index.
Mean blood lead
56.3 ug/dL (n = 173, measured in
1976)
High lead exposure defined as
workers from departments with an
average >0.2 mg/m3 airborne lead
or >50% of jobs had average
levels more than twice that level
(1975 survey). In this category,
n= 1,436.
Follow-up >90% in both groups; 2339 deaths observed
"chronic or unspecified nephritis" SMR
222 (95% CI: 135, 343) in battery workers
265 (95% CI: 114, 522) in lead production workers
"other hypertensive disease" SMR ("includes HTN and
related renal disease without mention of heart disease)"
320 (95% CI: 197, 489) in battery workers
475 (95% CI: 218, 902) in lead production workers
Race adjusted proportionate mortality ratios analyses similar.
Nephritis deaths observed primarily in workers hired before
1946.
Limitations = due to mortality analysis (inaccuracies of death
certificates, exposure assessment generally limited)
Compared to the U.S. white male population, the
standardized mortality ratio (SMR) for chronic kidney
disease, based on only 8 deaths, was 1.26 (95th CI = 0.54,
2.49). SMR = 1.55 in high lead exposure group, also not
significant. The SMR for chronic kidney disease increased
with duration of exposure from 0.79 in workers exposed 1-5
years to 2.79 in workers exposed >20 years; however SMR
was not significant.
Europe
Fanning (1988)
UK
Deaths from 1926-1985
Deceased males identified through pension records of lead battery
and other factory workers
867 deaths of mean with high lead exposure compared to 1206
men with low or no lead exposure
Range of blood lead
40-80 ug/dL since -1968 in high
lead exposure group; thought not
to have had clinical lead poisoning
due to medical surveillance
<40 |ig/dL since -1968 in little or
no exposure group
Odds ratio for renal disease = 0.62, not significant, based on
only 11 deaths. Similar for diagnosis of nephritis. Possible
decreasing odds ratio over time of deaths with mention of
nephritis on death certificate but not significant and numbers
still quite small.
Limitations = standard mortality study issues although
deaths compared with other workers and not general
population which is a strength in this type of study.
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Table AX6-4.5. Renal Effects of Lead - Children
to
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o
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
O
o
2
o
H
O
o
HH
H
W
United States
Hu(1991)
U.S.
Study date not provided
Loghman-Adham (1998)
Chicago, IL
Study date not provided
McDonald and Potter
(1996)
Boston, MA
1991
Moel and Sachs (1992)
Chicago, IL
1974-1989
21 of 192 adults who were hospitalized at Boston Children's
Hospital between 1932 to 1942 for childhood lead poisoning
were traced to a Boston area address. Matched on age, sex, race,
and neighborhood to 21controls.
Mean (SD) blood lead 6.0 ug/dL
(lead poisoned)
7.5 ug/dL (controls)
134 children and young adults, 8 to 13 years after chelation
therapy for severe lead poisoning
Mean age at poisoning = 2.3 years
Mean age at follow-up = 13.4 years
Mean peak blood lead level
121 ug/dL
Mean blood lead level at time
of study
18.6 ug/dL
454 pediatric hospital patients who were diagnosed with lead
poisoning between 1923 and 1966 were traced through 1991
Mortality study, comparison with U.S. population
62 participants with blood lead >100 ug/dL, diagnosed and
chelated between 1966 and 1972, together with 19 age-matched
control siblings with initial blood leads less than 40 ug/dL. Mean
age at follow-up = 22 years.
Renal outcomes = serum creatinine, uric acid, and (32-
microglobulin, fractional excretion of p2-microglobulin, urinary
protein:creatinine ratio, and tubular reabsorption of phosphate.
Mean initial blood lead
150.3 ug/dL (highly poisoned as
children)
Data for siblings not available as
levels <40 ug/dL not quantified.
No significant differences in blood lead level, serum
creatinine, or BUN. Mean measured creatinine clearance
higher in the previously lead poisoned group compared to
controls (112.8 vs. 88.8 mL/min/1.73 m2 [p < 0.01]). Mean
in the lead exposed group was also higher than the predicted
value of 94.2 mL/min/1.73 m2 from the nomogram of Rowe
et al. (1976). Suggests lead-related hyperfiltration. As noted
in section 6.4, one survivor, identified but not included in the
study, had disease consistent with lead nephropathy.
Limitations = small study size and concern for survivor bias
in the study group.
Mean serum creatinine was normal (0.8 mg/dL). Calculated
creatinine clearance normal in all but 3 children. No
correlation between either initial or current blood lead and
serum creatinine or calculated creatinine clearance.
Urinary a-amino nitrogen concentrations were significantly
increased compared with 19 healthy age matched controls
and were correlated with current blood lead levels. Thirty-
two children (24%) had glycosuria. Fractional excretion of
phosphate, however, was normal in all children. The author
concluded that a partial Fanconi syndrome could persist for
up to 13 years after childhood lead poisoning. The author
notes that the prognostic significance of this is unknown at
present.
Chronic nephritis was not a significant cause of death.
Mortality from all cardiovascular disease was elevated
(observed/expected = 2.1 [95% CI: 1.3, 3.2]) and cerebral
vascular deaths were particularly common among women
(observed/expected = 5.5 [95% CI: 1.1, 15.9]).
There were no statistical differences in either renal function
or blood pressure between study subjects and control
siblings. Initial blood lead level was not associated with
serum creatinine, after adjustment for age, gender and body
mass index. A modest increase in serum creatinine values
was observed over a nine-year period in four of the 62 study
subjects (up to 1.6 mg/dL).
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Table AX6-4.5 (cont'd). Renal Effects of Lead - Children
to
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o
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
O
o
2
o
H
O
o
HH
H
W
Europe
Bernard et al. (1995b)
Czech Republic
Study date not provided
De Burbure et al. (2006)
France, the Czech
Republic, and Poland
Study date not provided
Factor-Litvak et al.
(1999)
Kosovo, Yugoslavia
1985-1993
144 children living close to a lead smelter (exposed groups 1 and
2)
51 controls living in a rural area presumed to be relatively
unpolluted with lead.
Mean age = 13.5 years.
Renal outcome measures included urinary albumin, RBP, NAG,
Clara cell protein and p2-microglobulin.
Retinol binding protein
73.8 ug/g cr (controls)
109.4 ug/g cr (exposed group 1)
117.8 ug/g cr (exposed group 2)
P2-microglobulin
60.3 ug/g cr (controls)
89.1 ug/g cr (exposed group 1)
66.4 ug/g cr (exposed group 2)
NAG
1.56 lU/gcr (controls)
2.32 lU/g cr (exposed group 1)
1.46 lU/g cr (exposed group 2)
Multiple linear adjusting for age and gender.
804 exposed and control children
Exposed children recruited from residents near historical
nonferrous smelters, must have lived > 8 years near smelters
Mean age = 10 yrs; range = 8.5-12.3 yrs.
Renal outcome measures included serum creatinine, cystatin C
and 62-microglobulin as well as urinary RBP, NAG, Clara cell
protein.
577 children followed at 6 month intervals through 7.5 years
of age.
Divided into a high exposure and a low exposure group, based
on residence in Kosovska Mitrovica with a lead smelter, refinery
and battery plant or in Pristina, 25 miles away.
Renal outcome = Proteinuria assessed with a dipstick.
Multiple logistic regression modeling of proteinuria
dichotomized as either any or none, adjusting for socioeconomic
status, maternal education/ intelligence, and quality of
childrearing environment.
Blood lead
8.7 ug/dL (control boys)
8.39 ug/dL (control girls)
10.9 ug/dL (exposed boys 1)
9.4 ug/dL (exposed girls 1)
14.9 ug/dL (exposed boys 2)
12.9 |ig/dL (exposed girls 2)
Mean blood lead
ranged from 2.8 to 4.2 mg/dL in
various control and exposed
groups
Urinary cadmium, arsenic and
mercury as well as blood cadmium
also assessed
Mead blood lead from graph
peaked at —38 ug/dL between ages
3-5 in Kosovska Mitrovica and at
—10 ug/dL in controls. Blood lead
level (not means) range = 1 to
70 ug/dL
Mean blood lead levels significantly higher in both exposed
groups compared to the control group. In contrast, blood
cadmium levels were similar among all groups. After
adjustment for age, sex, blood cadmium, and zinc
protoporphyrin, log transformed blood lead was associated
with log transformed RBP (P coefficient = 0.302, p = 0.005
[SE nor CI provided]).
Serum concentrations of creatinine, cystatin C, and 62-
microglobulin negatively correlated with blood lead levels
Authors state suggestive of an early renal hyperfiltration that
averaged 7% in the upper quartile of PbB levels
(>5.5 ug/dL; mean, 7.84 ug/dL)
In higher exposed group, adjusted OR for proteinuria was
3.5 (CI = 1.7 - 7.2); adjusted odds of proteinuria increased
by 1.15 (CI = 1.1-1.2) per unit increase in blood lead in the
higher exposed group. Proteinuria unrelated to blood lead in
lower exposed control group.
Limitations = limited renal outcomes assessed, high dropout
rate in the study.
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Table AX6-4.5 (cont'd). Renal Effects of Lead - Children
to
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Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
Europe (cont'd)
O
o
2
o
H
O
o
HH
H
W
Pels etal. (1998)
Poland
1995
Oktem et al. (2004)
Turkey
Study date not provided
Price etal. (1999)
Belgium, Poland,
Germany and Italy
Study date not provided
112 children (50 controls, 62 exposed)
Mean age = 9.9 years and 10.6 years in controls and exposed
group, respectively.
Numerous (29) renal outcome measures were examined
including serum creatinine and p2-microglobulin, and urinary
NAG, RBP, Clara cell protein, p2-microglobulin, 6-keto-
prostaglandin FIO
(6-keto-PGFi0), prostaglandin E2 (PGE2) and thromboxane B2
(TXB2).
Urinary RBP
46 ug/g cr (exposed)
42 ug/g cr (controls)
Urinary P^-microglobulin
89 ug/g cr (exposed)
37 ug/g cr (controls)
Serum creatinine
0.63 mg/dL (exposed)
0.63 mg/dL (controls)
79 adolescent auto repair workers (mean age 17.3 years)
71 rural adolescents as negative controls (mean age 17.0 years)
Renal outcomes = urinary NAG, p2-microglobulin, uric acid,
and calcium; blood urea nitrogen (BUN), serum creatinine and
uric acid
Urinary lead measured in 481 European children (236 controls,
245 exposed) aged 6-14 years.
Several renal outcome measures assessed including urinary
NAG and p2-microglobulin; values not reported
Blood lead
13.3 ug/dL (exposed)
3.9 |ig/dL (controls)
Blood lead
7.79 ug/dL (exposed workers)
1.6 ug/dL (controls)
Mean urinary lead
Range from 3.9 to 7.2 ug/g cr
(controls)
Range from 5.2 to 24.6 ug/g cr
(exposed)
Significantly higher mean serum p2-microglobulin, and
urinary transferrin, 6-keto-PGFi0, thromboxane B2,
epidermal growth factor, (32-microglobulin, PGE2, and Clara
cell protein in the exposed children. In contrast, NAG-B
was lower in the exposed group. Categorical blood lead
associated with prevalence of values above the upper
reference limits for several biomarkers. Urinary 6-keto-
PGFi0, TXB2, p2-microglobulin, Clara cell protein,
epidermal growth factor and PGE2 positively correlated with
blood lead (r = 0.441, 0.225, 0.203, 0.261, 0.356, and 0.23,
respectively; all with significant p-values)
Limitations = data analysis, limited adjustment
No difference in mean BUN, serum creatinine, uric acid, or
GFR (apparently estimated) between workers and controls.
Urinary NAG and calcium significantly higher in workers
compared to controls. Urinary NAG positively correlated
blood lead (r = 0.427).
Limitations = data analysis, lack of adjustment
Urinary lead generally higher in exposed children as
compared to controls. Authors unexpectedly found
substantial differences in renal biomarkers by study site.
Authors note several renal biomarkers differed between
exposed and control groups. Also questioned the use of
"control" groups in ubiquitous exposures.
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Table AX6-4.5 (cont'd). Renal Effects of Lead - Children
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Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
O
o
2
o
H
O
o
HH
H
W
Europe (cont'd)
Schiireretal. (1991)
Germany
1988-1989
Sonmez et al. (2002)
Turkey
Study date not provided
Staessen et al. (2001)
Belgium
1999
22 children, age 5-14 years, with CRI
20 siblings or neighbors as lower exposed group
16 control children without known lead exposure
39 adolescent auto repair workers (mean age 16.2 years)
13 adult battery workers as positive controls (mean age 32 years)
29 rural adolescents as negative controls (mean age 14.8 years)
Serum creatinine
0.99 mg/dL (exposed group)
0.99 mg/dL (positive/ adult controls)
0.89 mg/dL (negative/ adolescent controls)
Urinary NAG
4.7 lU/g cr (exposed group)
7.4 lU/g cr (positive/ adult controls)
3.1 lU/g cr (negative/ adolescent controls)
100 exposed and 100 control children
Mean age = 17 years
Two exposed groups were recruited from industrialized suburbs
while the control group was recruited from a rural area.
Bymicroglobulin
5.22 ug/mmol cr (controls)
5.3 ug/mmol cr (exposed group 1)
9.09 ug/mmol cr (exposed group 2)
Cvstatin-C
0.65 mg/L (controls)
0.63 mg/L (exposed group 1)
0.71 mg/L (exposed group 2)
Mean blood lead
2.9 ug/dL in children with CRI,
not tested in other groups
Mean dental lead content
2.8 ug/g in children with CRI
1.7 ug/g in sibs/neighbors
1.4 ug/g in controls
Blood lead
8.13 ug/dL (exposed group)
25.3 ug/dL
(positive/adult controls)
3.49 ug/dL
(negative/ adolescent controls)
Blood lead 1.5 ug/dL (controls)
1.8 ug/dL (exposed group 1)
2.7 ug/dL (exposed group 2)
Lead levels in teeth significantly higher in both the patient
and sibling/neighbor control groups compared to the
unexposed control group.
All participants had normal blood urea, creatinine, and uric
acid levels as well as normal routine urinalysis
Blood lead level and urinary NAG significantly higher in
adolescent auto repair workers compared to the negative
control group
Limitations = data analysis, lack of adjustment
Blood lead, p2-microglobulin, and Cystatin-C levels higher
in exposed group 2 as compared to controls and exposed
group 1
After adjustment for sex and smoking status, blood lead was
associated with both pVmicroglobulin and cystatin-C.
A two-fold increase in blood lead was associated with a
3.6 % increase in Cystatin-C ([95% CI: 1.5, 5.7];
p < 0.0001) and a 16% increase in p2-microglobulin ([95%
CI: 2.7, 31]; p = 0.02). Blood cadmium was not associated
with either outcome.
Multiple linear regression adjusting for sex and smoking status
-------
Table AX6-4.5 (cont'd). Renal Effects of Lead - Children
to
o
o
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
Europe (cont'd)
Verberketal. (1996)
Romania
1991-1992
151 children who resided at different distances from a lead
smelter
Mean age = 4.6 years.
Renal outcomes = urinary RBP, NAG, ai-microglobulin,
albumin and alanine aminopeptidase.
Geometric means
Urinary RBP
49.4 ug/g cr
Urinary NAG
6.9 U/g cr
Urinary q^-microglobulin
2.4 mg/g cr
Urinary alanine aminopeptidase
19.8U/gcr
Multiple regression analysis adjusting for age and gender
Blood lead
34.2 (22.4) ug/dL
After adjustment for age and gender, a 10 ug/dL increase in
blood lead was associated with a 13.5% increase in NAG
excretion (90% CI = 10.2-17%). No threshold was
observed. No other significant associations noted.
Africa
Dioufetal. (2003)
Senegal
1998
38 Senegalese children
(19 exposed, 19 controls)
Age range = 8-12 years old.
Renal function assessed by measuring urinary alpha-glutathione
S-transferase (aGST)
Mean (SD) blood lead
10.7 (1.7) jig/dL (exposed)
6.1 (1.8) ug/dL (controls)
Blood lead significantly higher in exposed group (urban
dwellers) as compared to controls (rural dwellers).
Unclear as to whether aGST was higher or lower in controls
as compared to exposed group (stated to be higher in
controls in the results section BUT stated to be higher in the
exposed group in the discussion). Regardless, the difference
was not statistically significant.
Limitations = small sample size, data analysis
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ANNEX TABLES AX6-5
May 2006 AX6-117 DRAFT-DO NOT QUOTE OR CITE
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Table AX6-5.1. Cardiovascular Effects of Lead
to
o
o
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
Blood Pressure, Hypertension
United States
O
o
2
o
H
O
o
HH
H
W
Chen et al. (2006)
U.S.-Baltimore, MD;
Cincinnati, OH;
Newark, NJ;
Philadelphia, PA
-1998-2004
780 children from 12-33 months participated in a
randomized clinical trial of oral succimer
chelation in four clinical centers. Half the
children had up to three 26-day treatments, the
other half were given placebo. 75% got two
treatment sessions and 81% of those with two
treatments received a third.
Blood pressure was measured pre-treatment, at 7,
28 and 42 days after each treatment, then every 3
to 4 months for five years of follow up. Cross-
sectional multiple regression models adjusting
for clinic location, baseline linear lead, race, sex,
parents' education, single parent, age attest,
height at test, and BMI at test for each period of
the study tested the difference of diastolic and
systolic blood pressure between placebo and
succimer groups. Cross-sectional multiple
regression models for the effect of linear blood
lead at each period on blood pressure adjusted for
clinic location, treatment group, race, sex,
parents' education, single parent, age attest,
height and BMI. Two mixed models, one from
start of treatment to 9-month follow up, the other
from 12 to 60 months follow up, adjusted for the
same variables, and tested the effect of treatment
group over time.
Blood lead ranged from
20-44 ug/dL at pre-treatment
and from 1-27 ug/dL at
5 year follow up. Succimer-
treated group had
significantly lower blood lead
than placebo group only for
9-10 months following the
end of treatment. Blood lead
did not differ significantly
beyond that period.
Adjusted systolic blood pressure was significantly higher in the succimer group
than the placebo group at 36 months (1.27 mm Hg [95% CI: 0.06, 2.48]) and at
60 months follow up (1.69 mm Hg [95% CI: 0.34, 3.04]). Systolic blood pressure
was not significantly different at any other time period; diastolic blood pressure
was never significantly different between groups.
Concurrent linear blood lead was not associated with blood pressure in cross-
sectional models at any time point in the study. Adjusted coefficients for linear
blood lead and systolic blood pressure ranged from 1.36 mm Hg (95% CI: -0.58,
3.30) at pre-treatment to -0.72 mm Hg (95% CI: -1.91, 0.48) at 36 months of
follow up. Diastolic pressure coefficients were generally lower but followed the
same pattern.
Mixed model analysis for start of treatment through 9 months follow up showed
succimer treatment effect of 0.24 mm Hg (95% CI: -0.79, 1.28) for systolic and
0.46 mm Hg (95% CI: -0.44, 1.36) for diastolic blood pressure. The treatment
effect from 12 through 60 months follow up was 1.09 mm Hg (95% CI: 0.27,
1.90) systolic and 0.15 mm Hg (95% CI: -0.45, 0.75) for diastolic blood pressure.
The only reliable effect of succimer treatment was an elevation of systolic blood
pressure, especially notable between three and five years post treatment. The
authors could not account for the apparent increase in blood pressure in the
succimer-treated group 3-5 years after treatment ended. It is notable that the two
groups had different mean blood lead for less than a year after succimer treatment
ended, a period perhaps too short to observe any beneficial effect of treatment.
Failure to find cross-sectional effects of blood lead on blood pressure, especially
pre-treatment, may indicate that lead exposure for a period of less than three years
after birth is not sufficient to affect blood pressure. It could also mean that blood
pressure measurements in the first three years of life are highly variable, as could
be seen from scatter plots of blood pressure versus blood lead at pre-treatment
compared 60 month follow up. The use of linear lead term may have reduced
sensitivity to finding a significant blood lead effect on blood pressure. No model
diagnostics mentioned.
The same study group also showed no effect of succimer treatment on IQ or
neurobehavioral test scores at 36 and 60 months follow up in other publications.
-------
Table AX6-5.1 (cont'd) Cardiovascular Effects of Lead
to Reference, Study
Q Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
Blood Pressure, Hypertension
United States (cont'd)
Cheng etal. (2001)
U.S.-Boston, Normative
Aging Study (VA)
1991-1997
833 males (—97% white), average age
(SD):
65.5 (7.2) Normotensive subjects,
N = 337
68.3(7.8) Borderline hypertension
subjects, N= 181
67.9 (6.8) Defnite hypertension subjects,
N = 314
474 males with no history of
hypertension at first measurement,
returning up to 6 years later for
hypertension study.
Linear multiple regression models of
blood pressure and Cox proportional
hazard models of new cases of
hypertension after up to 7 years, with one
group of covariates forced into models
based on biological plausibility and
another group forced based on significant
univariate or bivariate results or >20%
effect modification of lead variable
coefficient in multiple models. Linear
blood lead, tibia lead, and patella lead
forced in separate models.
Arithmetic mean (SD) blood
lead: 5.9-6.4 ug/dL (3.7-4.2),
depending on hypertension group
(only data shown).
Multiple regression models of blood pressure always included age, age-squared, BMI,
family history of hypertension, daily alcohol consumption, and daily calcium
consumption. Increasing tibia lead concentration was associated with increased
systolic blood pressure (diastolic not addressed) in baseline measurements in subjects
(n = 519) free from definite hypertension (systolic >160 mmHg, diastolic >95 mmHg,
or taking daily antihypertensive medication). Each increase of 10 ug/g tibia lead
concentration was associated with an increase in systolic blood pressure of 1.0 mmHg
(95% CI: 0.01, 1.99). Patella and linear blood lead were not significant.
Cox proportional hazard models always included age, age-squared, BMI, and family
history of hypertension. In follow up (n = 474), only increasing patella lead predicted
increasing risk of definite hypertension in those classified as normotensive at baseline.
For every 10 ug/g increase in patella lead risk ratio increased 1.14(95% CI: 1.02,
1.28). Combining borderline hypertension (systolic 141-160 mmHg or diastolic 91-95
mmHg) with definite hypertension (n = 306), the relative risk ratio of becoming a
combined hypertensive associated with a 10 ug/g increase in patella lead was 1.23
(95% CI: 1.03, 1.48). Linear blood lead and tibia lead were not significant.
Linear blood lead is not indicated for blood pressure models due to strong likelihood of
significant residual heteroscedasticity and non-normality. Relatively small sample size
may have prevented tibia blood lead significance in the Cox proportional hazard
models.
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Table AX6-5.1 (cont'd). Cardiovascular Effects of Lead
to
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o
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
Blood Pressure, Hypertension
United States (cont'd)
Den Hond et al. (2002)
U.S.-NHANES III
1988-1994
O
o
2
o
H
O
o
HH
H
W
Gerr et al. (2002)
U.S.-Spokane WAand
area around Silver
Valley ID
1994
4,685 white males, 5,138 white females,
1,761 black males, 2,197 black females,
from 20 years up. Log-transformed
blood lead, systolic and diastolic blood
pressure measured at survey time and
analyzed with forward, stepwise multiple
regression with covariates.
Avg. Age:
White
Male: 44.3
Female: 46.2
Black
Male: 40.5
Female: 41.5
502 young people, age 19-29 years, 53%
female, nearly evenly divided into the
Spokane group (no unusual childhood
exposure) and the Silver Valley group,
where a lead smelter operated during
their childhood. Multiple regression
models of systolic blood pressure and
diastolic blood pressure. All covariates
forced into model as block with both
linear blood lead and tibia bone lead in
each model.
Geometric Mean
(25th-75th percentile)
blood lead:
White Male Mean 3.6 ug/dL
(2.3-5.3)
White Female Mean 2.1 ug/dL
(1.3-3.4)
Black Male Mean 4.2 ug/dL
(2.7-6.5)
Black Female Mean 2.3 ug/dL
(1.4-3.9)
Mean (SD) blood lead only given
stratified on tibia lead category:
(Tibia <1 ug/g) blood lead mean
1.9 ug/dL (1.6)
(Tibia 1-5 ug/g) blood lead mean
2.3 ug/dL (2.1)
(Tibia 6-10 ug/g) blood lead
mean 2.4 ug/dL (2.4)
(Tibia <10 ug/g) blood lead mean
3.2 ug/dL (2.3)
No other descriptive tibia lead
data given.
After adjusting for age, age-squared, BMI, hematocrit, smoking, alcohol, and an
indicator variable for use of antihypertensive medications, each model was further
modified by a unique mix of other covariates, including: coffee consumption, dietary
calcium, dietary sodium/calcium ration, total serum protein, total serum calcium,
diabetes, and poverty index. Log lead was forced in last.
In stratified analyses, only blacks had significant positive blood pressure associations
with log blood lead. Each doubling of blood lead was associated with increase of black
male systolic blood pressure of 0.9 mmHg (95% CI: 0.04, 1.8), black female systolic
blood pressure of 1.2 mmHg (95% CI: 0.4, 2.0), and female diastolic blood pressure of
0.5 mmHg (95% CI: 0.01, 1.1). In white males only, each doubling of blood lead was
significantly associated with a decrease in diastolic blood pressure of-0.6 mmHg (95%
CI: -0.9, -0.3).
Stepwise models can rely on chance associations due to multiple testing and usually
lead to a different pattern of covariate adjustment in different models. Inclusion of
likely confounding variables such as serum calcium could have affected estimated lead
effects. No testing for significant lead coefficient differences between each stratum.
No model diagnostic tests reported. No explanation offered for inverse relationship
between lead and diastolic blood pressure in white males. No adjustment for survey
design.
Adjusting for sex, age, height, BMI, education, income, current smoker, current
alcohol use, childhood residence (the two recruitment areas), current birth control pills,
hemoglobin, and serum albumin, only tibia lead, and not linear blood lead, was
significantly related to systolic and diastolic blood pressure. Compared to the <1 ug/g
tibia lead category, subjects in the >10 ug/g category had 4.3 mmHg (95% CI: 1.4,
6.7) higher systolic blood pressure and 2.8 mmHg (95% CI: 0.4, 5.2) higher diastolic
blood pressure.
Linear blood lead is not indicated for blood lead-blood pressure models. No diagnostic
testing reported. Insufficient descriptive data given for tibia lead.
-------
Table AX6-5.1 (cont'd). Cardiovascular Effects of Lead
to Reference, Study
Q Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
Blood Pressure, Hypertension
United States (cont'd)
Gump et al. (2005)
U.S.-Oswego, NY
Dates of study not given
122 9.5-year old children participated.
Multiple regression models of percent
changes in systolic and diastolic blood
pressure, heart rate, stroke volume,
cardiac output, and total peripheral
resistance due to acute stress were
adjusted by stepwise entry of up to
50 possible control variables with
quartile blood lead forced in last.
A linear contrast was used to test dose-
response effects of quartile lead.
Linear lead terms were also used.
Contemporary blood lead quartile:
1st quartile: 1.5-2.8 ng/dL
2ntl quartile: 2.9-4.1 ug/dL
3rd quartile: 4.2-5.4 ug/dL
4th quartile: 5.5-13.1 ug/dL
All betas represent percent change in outcome from nonstress to stress condition for
each change in 1 ug/dL blood lead.
Systolic BP beta: -0.009 (95% CI: -0.74,0.055)
Diastolic BP beta: 0.069 (95% CI: -0.001,0.138)
Heart rate beta: 0.013 (95% CI: -0.046,0.072)
Stroke volume beta: -0.069 (95% CI: -0.124,-0.015)
Cardiac output beta: -0.056 (95% CI: -0.113,0.001)
Total peripheral resistance beta: 0.088 (95% CI: 0.024,0.152)
Mean successive difference of cardiac interbeat interval beta:
-0.028 (95% CI: (-0.098,0.042)
Despite low power to detect significant effects, blood pressure, cardiac output, and
total peripheral resistance change to stress were associated with contemporary blood
lead. Stepwise modeling creates unique models for each outcome. Some models had
up to 12 control variables plus lead, an excessive number for only 122 subjects.
Scatter plots of regression with linear lead and bar charts of response to quartile lead
showed obvious non-linearity, though all lead effects were modeled as linear effects.
Probability of contemporary exposure to mercury and PCB's was very high.
No model diagnostic testing reported.
-------
Table AX6-5.1 (cont'd). Cardiovascular Effects of Lead
to Reference, Study
Q Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
Blood Pressure, Hypertension
United States (cont'd)
Glenn et al. (2003) 496 males, mean (SD) (range) age 55.8
U.S.-New Jersey (7.4, (40-71) years, working or formerly
1994-1998 working at a plant producing tetraethyl
or tetramethyl lead until 1991, were
followed from 10 months to 3.5 years
during which blood pressure was
repeatedly tested. Blood lead was tested
only at baseline. Tibia lead was tested in
1991 (at the end of organic lead
production at the plant) and called "peak
tibia lead" and again during 1997
(year 3). Generalized estimating
equations with an exchangeable
correlation structure for repeated
measurements were used for systolic and
diastolic blood pressure. One group of
covariates was forced into the model as a
block (age at baseline, race, BMI,
indicator variable for technician, lead
variable (linear blood lead, peak tibia
lead, and tibia lead each tested
separately), duration of follow up, and
the interaction between the lead variable
and the duration term. Potential
confounding variables were entered
stepwise and retained in the model if
significant. Alternate models not using
linear time were constructed, using
quartile of follow up time to avoid
assuming a linear relationship of change
in blood pressure with time.
Arithmetic mean (SD, range)
blood lead at baseline:
(4.6,2.6, -l-20)ng/dL.
Tibia lead at year 3:
14.7(9.4,-1.6-52) ug/g
Peak tibia lead:
24.3(18.1, -2.2-118.8)
Controlling for baseline age, BMI, antihypertensive medication use, smoking,
education, technician and number of years to each blood pressure measurement, each
1 |ig/dL increase in linear baseline blood lead was associated with average systolic
blood pressure increase of 0.64 mmHg/year (95% CI: 0.14, 1.14), each 10 ug/g
increase in year 3 tibia lead with an average increase of 0.73 mmHg/year (95% CI:
0.23, 1.23), and each increase of 10 ug/g of peak tibia lead with an average increase of
0.61 mmHg/year (95% CI: 0.09, 1.13). Similar results were obtained using the follow
up time quartile designation for systolic blood pressure with all subjects and with
subjects not taking antihypertensive medications.
This was one of the few studies using a prospective design and that used a statistical
technique accounting for repeated measures. No justification given for using an
exchangeable correlation structure instead of an alternate one. Only examined cortical
bone lead (tibia) and not trabecular bone lead (patella or calcaneus). Linear blood lead
may not be indicated for use in blood lead-blood pressure models. Stepwise modeling
involves multiple testing of the same data set with no control for altered probabilities.
No model diagnostics presented.
-------
Table AX6-5.1 (cont'd). Cardiovascular Effects of Lead
to Reference, Study
Q Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
Blood Pressure, Hypertension
United States (cont'd)
O
o
2
o
H
O
o
HH
H
W
Glenn et al. (2001) 213 males (92% white), mean (SD) age
U.S.-New Jersey 58.0 (7.4) years, working or formerly
1996-1997 working at a plant producing tetraethyl
or tetramethyl lead until 1991, were
genotyped for ATP1A2(5') and
ATP1A2(3') polymorphism. ATPase is
thought to play a role in regulating blood
pressure and lead inhibits its activity.
Blood pressure, blood lead, and tibia
lead were measured. Multiple linear
regression models were used for systolic
and diastolic blood pressure. Logistic
regression model was reported for
hypertension (systolic >160 mmHg,
diastolic > 96 mmHg, or taking
antihypertensive medications).
Covariate entry methods not specified,
but were likely stepwise. Covariates for
the blood pressure model were age, use
of antihypertensive medications, alcohol,
smoking, season of year, linear blood
lead, tibia lead (the two lead measures
apparently tested separately),
ATP1A2(5') and ATP1A2(3')
polymorphism (each tested separately),
and an interaction term between
polymorphism and lead. Covariates for
the hypertension models were age, BMI,
lifetime alcoholic drinks, linear blood
lead and tibia lead, and polymorphism,
each lead measure and polymorphism
tested separately.
Arithmetic mean (SD, range)
blood lead:
5.2ug/dL(3.1, 1-20).
Mean (SD) tibia lead:
16.3 ug/g (9.3)
None of the relationships between the ATP1A2(5') polymorphism and either blood or bone
lead or blood pressure were significant.
The ATP1A2(3') polymorphism was homogenous for the 10.5 kilobase allele (10.5/10.5) in
11 subjects, heterogeneous for the 10.5 and 4.3 kilobase allele (10.5/4.3) in 82 subjects, and
heterogeneous (10.5/4.3) in 116 subjects. Prevalence of the 10.5 allele was significantly
higher in blacks than in whites.
Regression coefficients of 4.3/4.3 and 10.5/4.3 genotypes were not significantly different
and all subsequent analyses compared the 10.5/10.5 genotype with the combined 4.3/4.3-
10.5/4.3) genotype. The significant interaction between linear blood lead and the 10.5/10.5
genotype showed that for every 1 ug/dL of blood lead systolic blood pressure increased 5.6
mmHg (95% CI: 1.2, 9.9) more than the blood pressure of the combined genotype group.
Blood lead range of the combined genotype group was twice that of the 10.5/10.5 group.
When data were truncated to make blood lead of both groups cover the same range,
coefficients of the genotype-linear blood lead interaction term did not change appreciably.
Authors state that tibia lead interacted with genotype on blood pressure but showed no data
to estimate either type or size of effect. Diastolic blood pressure was not related to
genotype, to lead or to the interaction between lead and genotype.
Prevalence of hypertension (30% in total sample) was significantly higher among the
10.5/10.5 group (63.4 %) than among the combined group (28.3 %). Adjusting for age,
BMI, and lifetime alcohol, the odds of hypertension in the 10.5/10.5 group were OR 7.7
(95% CI: 1.9, 31.4) compared to the 4.3/4.3 group. The heterogeneous group was not
significantly different from the 4.3/4.3 group.
Linear blood lead specification not indicated for blood lead-blood pressure modeling.
Examination of partial residual plot for systolic blood pressure and linear blood lead shows
typical heterogeneity of residuals as a function of predicted values. Thus, presented
coefficients may be inefficient and biased. Only 9 subjects were homogenous for 10.5/10.5
in the multiple regression model. Only cortical bone lead was tested, not trabecular bone
lead. Cortical bone lead models not shown or quantitatively described. Blood lead rounded
to nearest unit ug/dL. Mixed organic-inorganic lead exposure. Relatively small sample
size may have prevented detection of other significant effects. No model diagnostics
described.
-------
Table AX6-5.1 (cont'd). Cardiovascular Effects of Lead
to Reference, Study
Q Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
Blood Pressure, Hypertension
United States (cont'd)
O
o
2
o
H
O
o
HH
H
W
Hu et al. (1996) 590 males (over 98% white), mean age around
U.S.-Boston- 67 years, divided into 146 hypertensives
Normative Aging (systolic >160 mmHg, diastolic >95 mmHg, or
Study-VA daily antihypertensive medication) and 444
1991-1994 non-hypertensives. Linear blood lead, tibia
and patella bone lead added separately to
logistic regression model containing forced
covariates of age, race, BMI, family history of
hypertension, pack-years smoking, alcohol
ingestion dietary sodium and calcium. Then, a
backward elimination procedure starting with
all covariates, including all lead variables,
resulted in a model in which only significant
covariates were retained.
Korrick et al. 284 women, from 47-74 years, mean age (SD)
(1999) 5 8.7 (7.2), were divided into 97 cases
U.S.-Boston- (systolic >140 mmHg, diastolic >90 mmHg, or
Nurse Health physician-diagnosed hypertension) and
Study 195 controls. Controls were further classified
1993-1995 as low normal (<121/75 mmHg) and high
normal >121/75 mmHg). Three ordinal
regression models were constructed, each
containing either blood lead, tibia lead or
patella lead with forced entry of all other
covariates. A final backwards elimination
ordinal regression model started with all
covariates, including all lead variables,
excluding each until only significant variables
were left. Interactions were tested in the final
model between patella lead and alcohol use,
age, and menopausal status.
Hypertensives:Arithmetic mean
(SD) blood lead:6.9 ug/dL (4.3)
Mean tibia lead:
23.7 ug/g (14.0)
Mean patella lead:
35.1 ug/g (19.5)
Non-hypertensives:
Arithmetic mean (SD) blood lead:
6.1 ug/dL (4.0)
Mean tibia lead:
20.9 ug/g (11.4)
Mean patella lead:
31.1 ug/g (18.3)
Mean blood lead (SD, range):
3.1 ug/dL (2.3, <1-14)
Mean tibia lead (SD, range):
13.3 ug/g (9.0, -5-69)
Mean patella lead (SD, range):
17.3 ug/g (11.1, -5-87)
Logistic regression model with all forced covariates revealed no significant lead effects
when the three lead variables were forced into the model separately. After backward
elimination, the only significant covariates left were BMI and family history of
hypertension. Of all the lead variables, only tibia lead remained in the model.
With each increase of 10 ug/g of tibia lead, odds of being classified hypertensive rose
(OR 1.21; 95% CI: 1.04, 1.43).
Stepwise regression, backward or forward, involves multiple testing with the same
data set, capitalizes on chance occurrence in the data set, and gives over-optimistic
probability values. No model diagnostic testing reported.
Only patella lead was significantly related to increased odds of hypertension in the
preliminary models, adjusted for age, BMI, alcohol, dietary calcium and sodium, ever
smoke, and family hypertension. Each 10 ug/g increase in patella lead was associated
with increased odds of hypertension OR 1.28 (95% CI: 1.03, 1.60). In the backward
elimination model adjusted for age, BMI dietary sodium and family hypertension, only
natural log transformed patella lead remained in the model. Identical odds ratios from
patella lead were obtained in both models. None of the interaction tests were
significant.
Small study size may have limited power to detect significant interactions. The
proportional odds assumption of the ordinal regression model was verified. Note that
the odds ratios above are for movement from one of the two lower categories, low
normal and high normal, to the next higher category as patella lead increased. No other
model diagnostic tests reported.
-------
Table AX6-5.1 (cont'd). Cardiovascular Effects of Lead
to Reference, Study
Q Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
Blood Pressure, Hypertension
United States (cont'd)
Morris etal. (1990)
U.S.-sampled from
general population
around Portland, OR
responding to ads to
participate in clinical
trials of non-
pharmacological
management of blood
pressure.
1984-1989?
145 males and 106 females, 73% with
arterial pressures >105 mmHg, provided
blood pressure measurements once a week
over four consecutive weeks. Blood for
lead analysis was collected during this
period. Stepwise multiple regression was
used to construct separate models of
systolic and diastolic blood pressure
stratified by sex. Covariates available to be
entered were age, BMI, dietary calcium and
"other nutrient intakes," ionized serum
calcium, erythrocyte protoporphyrin and
natural log transformed blood lead
Arithmetic mean (SD) blood
lead:
Males: 8.0 ug/dL (4.4)
Females: 6.9 ug/dL (3.6)
Natural log blood lead was only a significant predictor of blood pressure in males.
Adjusting for age and ionized serum calcium, every one natural unit increase in blood
lead was significantly associated with a 4.58 mmHg (neither SE nor CI stated)
in systolic blood pressure and, adjusting for hemoglobin, age, and current smoking,
a 1.90 mmHg (neither SE nor CI stated) in diastolic blood pressure.
The usual precautions regarding multiple testing and different covariate patterns in
stratified models constructed with stepwise regression apply. Reporting of effects not
complete. Small sample size limits conclusions about non-significant effects. High
prevalence of hypertensives in sample due to study recruitment design. Blood lead
technique, as represented by presented graph, had a detection limit of 5 ug/dL.
No model diagnostics.
-------
Table AX6-5.1 (cont'd). Cardiovascular Effects of Lead
to
o
o
Reference, Study
Location, and
Period
Study Description
Lead Measurement
Findings, Interpretation
O
o
2
o
H
O
o
HH
H
W
Blood Pressure, Hypertension
United States (cont'd)
Nash et al. (2003)
U.S.-NHANES III
1988-1994
1084 premenopausal and 633
postmenopausal women, from 40 to
59 years. Multiple linear regression
models with covariates, including
linear blood lead, entered as a block
for systolic and diastolic blood
pressure. Logistic regression models
with same covariates and lead quartile
added last for hypertension.
Mean (range) blood lead by
lead quartile:
1st quartile mean 1.0 ug/dL
Range: 0.5, 1.6
2ntl quartile mean 2.1 ug/dL
Range: 1.7,2.5
3ri quartile mean 3.2 ug/dL
Range: 2.6, 3.9
4th quartile mean 6.4 ug/dL
Range: 4.0,31.1
Linear blood lead was entered last after forcing in age, race/ethnicity, alcohol use, cigarette
smoking, BMI, and kidney function (serum creatinine) in multiple regression models for all
women and women stratified by menopause status for systolic and diastolic blood pressure. Lead
quartile was added to logistic regression models of hypertension (systolic >140 mmHg,
diastolic >90 mmHg or taking antihypertensive medication with the same covariates as the blood
pressure models, in all women and stratified by menopausal status. Tested additional models in
which women treated for hypertension were excluded from models. All models were adjusted
for sample design and weighting.
Each increase of 1 ug/dL of blood lead was significantly associated with a 0.32 mmHg (95% CI:
0.01, 0.63) increase of systolic blood pressure and a 0.25 mmHg (95% CI: 0.07, 0.43) increase
of diastolic blood pressure in all women without respect to menopausal status. In analyses
stratified by menopausal status, only postmenopausal women showed a significant blood lead
effect. For each 1 ug/dL increase of blood lead was associated with significantly increased
diastolic blood pressure of 0.14 (95% CI: -0.11, 0.39 sic.) only in postmenopausal women.
Referenced to the first blood lead quartile, no other quartile showed significantly increased odds
for hypertension in all subjects or in subjects stratified by menopausal status. With further
analyses stratified by systolic and diastolic hypertension without women taking antihypertensive
medications, in the combined group of pre and postmenopausal women the odds of diastolic
hypertension were significant when the 4th lead quartile was compared to the 1st quartile (OR 3.4
[95% CI: 1.3, 8.7]). In a model of only postmenopausal women untreated for hypertension, odds
of diastolic hypertension were significantly increased in the higher three quartiles of blood lead
(OR 4.6 [95% CI: 1.1, 19.2]); OR 5.9 [95% CI: 1.5, 23.1]); OR 8.1 [95% CI: 2.6, 24.7]),
respectively) and odds of systolic hypertension were significant only for the two middle lead
quartiles (OR 3.0 [95% CI: 1.3, 6.9]; OR 2.7 [95% CI: 1.2, 6.2]), respectively.
Linear blood lead is suspect in linear regression models of blood pressure as it is usually
associated with biased and inefficient estimation of lead coefficients due to probable
heteroscedasticity and non-normal distribution of residuals. No model diagnostics were reported.
No statistical testing for differences in lead coefficients according to strata. Nine stratified
models overall. Not all stated significance levels and standard errors in the blood pressure model
table corresponded for certain variables.
-------
Table AX6-5.1 (cont'd). Cardiovascular Effects of Lead
to
o
o
Reference, Study
Location, and
Period
Study Description
Lead Measurement
Findings, Interpretation
Blood Pressure, Hypertension
United States (cont'd)
Nawrot et al. (2002)
31 U.S. and
European studies,
community and
occupationally
exposed, published
between 1981 and
2001.
48 different groups, 32 of which were
only of men, 15 of which were only of
women, and one studying both sexes.
Total meta-analysis N > 58,490.
Age ranged from 15 to 93 years,
depending on the study. Two
methods of meta-analysis were used,
subject-weighted and non-weighted,
using study-reported effect sizes and
standard errors, transformed from the
original study specification of blood
lead (linear, logarithmic, or blood lead
group) to a single effect size for
doubling of blood lead. Also did
analyses stratified by race and sex.
Mean blood lead
concentration across studies
ranged from 2.3 to 63.8
ug/dL. Total range of blood
lead across studies was 0 to
97.9 ug/dL.
Each doubling of blood lead was associated with a significant 1.0 mmHg (95% CI: 0.5, 1.4)
increase in systolic blood pressure and a significant 0.6 mmHg (95% CI: 0.4, 0.8) increase in
diastolic blood pressure. Stated that differences in lead effect were not statistically different
between sexes, but did not describe test nor give statistics other than p-values. Presented black
and white differences as a trend for blacks to be "more susceptible than whites", but presented no
tests.
Statistically examined assumptions of homogeneity of effect and found no significant
heterogeneity. Tested for publication bias (statistically significant results tend to be published
more than non-significant results) and found no evidence. Found no significant effects of
removing one study at a time in sensitivity analysis. It appears that the presented results of effect
sizes and confidence intervals were calculated by the subject-weighted method, but this was not
made explicit. Included some studies that presented no lead coefficients or standard errors,
assuming effect size of zero, though the reported effect sizes without these studies did not appear
to be different from overall effect sizes. For studies using a linear lead measure, effect sizes were
calculated by doubling the arithmetic mean blood lead. If the concentration-response curve for
the lead-blood pressure relationship was really better characterized by a log-linear function, the
authors' use of studies with a linear blood lead term with high average blood lead led to over-
estimation of the slope of the relationship and those studies with low blood lead averages
produced an under-estimation of the slope of the relationship.
-------
Table AX6-5.1 (cont'd). Cardiovascular Effects of Lead
to
o
o
Reference, Study
Location, and
Period
Study Description
Lead Measurement
Findings, Interpretation
O
o
2
o
H
O
o
HH
H
W
Blood Pressure, Hypertension
United States (cont'd)
Proctor etal. (1996)
U.S.-Boston-
Normative Aging
Study (VA)
1992-1993
Rothenberg et al.
(1999)
U.S.-Los Angeles
1995-1998
798 men from 17 to 44 years.
Multiple linear regression models of
natural log blood lead on systolic and
diastolic blood pressure. All
covariates forced into model.
Arithmetic mean (SD, Range)
blood lead:
6.5 ug/dL (4.0, 0.5-35)
1188 immigrants and 439
nonimmigrants, from 15 to 43 years,
all women in third trimester of
pregnancy. Multiple regression
models of natural log blood lead on
systolic and diastolic blood pressure
with all covariates forced into models.
Covariates selected from larger set
based on significant univariate or
bivariate tests.
Geometric Mean (SD) blood
lead:
Immigrants: 2.3 ug/dL(1.4)
Non-immigrants: 1.9 ug/dL
(1.3)
Natural log blood lead, age, age-squared, BMI, adjusted dietary calcium, exercise, indicator
variables for current and former smoker, daily alcohol consumption, sitting heart rate, and
hematocrit were entered into multiple regression models without regard for significance.
Increased diastolic, but not systolic, blood pressure was significantly associated with increased
blood lead. Each natural log increase in blood lead was associated with a 1.2 mmHg (95% CI:
0.1, 2.2) increase in diastolic blood pressure.
Interactions between dietary calcium and blood lead on blood pressure were not significant.
Further analyses stratified on use of antihypertensive medication and those older than or equal to
74 years still revealed significant blood lead-diastolic blood pressure relationships.
Blood lead in over half the study group (n = 410) was determined by analyzing previously frozen
erythrocytes collected several years prior to the blood pressure measurements used in the study
and corrected by using hematocrit values also measured when blood was originally collected.
Combining both groups means that nearly half the group was tested for the effects of blood lead
on blood pressure measured at the same time, the other half measured several years apart. There
was no correction in models for this potential effect. The effect of taking antihypertensive
medication could have been assessed in a single model by using an indicator variable.
No statistical testing for the effects of stratification on the blood lead-blood pressure relationship.
No model diagnostics.
Natural log blood lead, age, BMI, coffee drinking, iron supplementation, and job stress were
entered as a block without regard to significance in linear multiple regression models of systolic
and diastolic blood pressure stratified by immigration status.
Increased blood lead was significantly associated with increased blood pressure only in
immigrants. Each natural log unit increase in blood lead was associated with a 1.7 mmHg (95%
CI: 0.7, 2.8) increase in systolic blood pressure and a 1.5 mmHg (95% CI: 0.5, 1.9) increase in
diastolic blood pressure in immigrants.
Used and reported model diagnostic tests, as evidenced by the use of standard error calculations
robust to residual heteroscedasticity. Stated reasons for stratification on immigrant status were
significant differences between the two groups in blood lead, blood pressure, age, BMI, and
education. Did not statistically test difference in lead coefficients between the immigration
strata. Did not correct for potential non-linearity in age effects on blood pressure.
-------
Table AX6-5.1 (cont'd). Cardiovascular Effects of Lead
to
o
o
Reference, Study
Location, and
Period
Study Description
Lead Measurement
Findings, Interpretation
Blood Pressure, Hypertension
United States (cont'd)
Rothenberg et al. 668 women, 15 to 44 years, studied in third
(2002a) trimester pregnancy and again a mean of 10
U.S.-Los Angeles weeks postpartum. Exclusion criteria were
1995-2001 diabetes, renal or cardiovascular disease,
extreme postnatal obesity (BMI > 40), and
subjects using stimulant drugs. Multiple
linear regression models of natural log
blood lead, tibia and calcaneus lead on
systolic and diastolic blood pressure with
all covariates and all lead variables forced
into model. Separate models for third
trimester and postpartum, excluding all
women with hypertension (see below)
during each specific period. Logistic
regression for hypertension (systolic >140
mmHg or diastolic >90), specific to third
trimester and postpartum periods with the
same covariates and lead variables.
Geometric mean blood
lead (SD):
3ri trimester: 1.9 ug/dL
(1.7)
postpartum: 2.3 ug/dL
(2.0)
Tibia mean lead (SD):
8.0 ug/g (11.4)
Calcaneus mean lead
(SD):
10.7 ug/g (11.9)
Multiple linear regression models for normotensives adjusted for postnatal hypertension (3ri
trimester model only), BMI, age, parity, smoking, alcohol, immigrant status, and educational
level plus all three lead indices. Only calcaneus lead was associated with blood pressure in 3ri
trimester models. Every 10 ug/g increase in calcaneus lead was associated with 0.70 mmHg
(95% CI: 0.04, 1.36) increase in systolic blood pressure and a 0.54 mmHg (95% CI: 0.01, 1.08)
increase in diastolic blood pressure. In postpartum models, natural log blood lead was the only
variable statistically associated with blood pressure. Every natural log unit increase in blood lead
was associated with -1.52 mmHg (95% CI: -2.83, -0.20) decrease in systolic blood pressure
and a -1.67 mmHg (95% CI: -2.85, -0.50) decrease in diastolic blood pressure.
In logistic models, only calcaneus lead was significantly associated with increased odds for
hypertension. Each 10 ug/g increase in calcaneus lead was associated with an OR 1.86 (95% CI:
1.04, 3.32) of 3ri trimester hypertension. None of the lead variables was associated with
postpartum hypertension.
Models did not use age-squared covariate. Models did not use repeated measures statistics. No
statistical comparisons between 3ri trimester and postpartum models. Model diagnostic tests
reported.
-------
Table AX6-5.1 (cont'd). Cardiovascular Effects of Lead
to
o
o
Reference, Study
Location, and
Period
Study Description
Lead Measurement
Findings, Interpretation
O
o
2
o
H
O
o
HH
H
W
Blood Pressure, Hypertension
United States (cont'd)
Schwartz et al. 543 mostly former organolead workers,
(2000c) predominantly white (92.8%), at a
U.S.-Eastern tetraethyl/tetramethyl plant, mean (SD)
1996-1997 [range] age 7.6 (7.6) [41.7-73.7] years had
blood lead, DMSA-chelatable lead (4-hr.
urinary lead excretion after a single 10
mg/kg dose of DMSA) measured for
modeling systolic and diastolic blood
pressure and hypertension (systolic >160
mmHg or diastolic > 96 mmHg or taking
antihypertensive medications. Tibia lead
~2 years later was also used as a lead index.
For blood pressure, linear multiple
regression with backward elimination of
non-significant covariates or covariates that
"had important influence on the coefficients
for the lead-dose terms." Each lead
variable was tested in a separate model.
Potential covariates for these models were
age, BMI, current tobacco use, and current
use of antihypertensive medications. Other
models were constructed taking out those
subjects using antihypertensive
medications. Both linear and linear +
quadratic blood and tibia lead terms were
tested. Logistic regression analyses were
used to test the effect of the lead variables
on hypertension, controlling for age,
diabetes, lifetime alcohol consumption, and
BMI. Logistic models also tested each lead
measure in interaction with age.
Blood lead arithmetic
mean (SD, range)
4.6 ug/dL (2.6, 1-20)
DMSA-chelatable lead
mean (SD, range)
19.0 ug (16.6, 1.2-136)
Tibia lead mean (SD,
range) 14.4 ug/g
(9.3,-1.6-52)
Adjusting for age, BMI, current smoking, and current use of antihypertensive medications, each
1 ug/dL increase in blood lead-squared was significantly associated with 0.189 mmHg (95% CI:
0.087, 0.330) increase in systolic blood pressure with three outliers removed. With the same
covariates, each 1 ug/dL increase in linear blood lead was significantly associated with 0.310 mmHg
(95% CI: 0.028, 0.592) in diastolic blood pressure taken over a 2-year period (n = 525). No other lead
variables were significant.
For the hypertension models, only the interaction of linear blood lead by age was significant, with
subjects showing significant decrease in odds ratio of hypertension with every joint increase of 1
ug/dL blood lead and 1 year increase in age (linear blood lead X age OR 0.98; [95% CI: 0.97, 0.99]).
The interaction suggested a concentration-response relationship between linear blood lead and
hypertension only up to —58 years of age.
Authors note that blood pressure findings "were not affected by exclusion or inclusion of subjects
using antihypertensive medications," but do not present either the data or the statistical tests to
evaluate that conclusion. No other model diagnostics were reported. Although blood lead was also
modeled as a quadratic lead term for systolic blood pressure, no analysis was shown for non-linear
blood lead terms for diastolic blood pressure.
Trabecular bone lead was not tested, though other studies indicate that it is a better lead index than
cortical lead for cross-sectional blood pressure and hypertension study.
Although the backward procedures described could have resulted in less than the full set of considered
covariates entering the models, all model presentations were limited to showing the lead coefficients
and all models indicated in a footnote that the lead coefficients were adjusted for each possible
covariate for that model. While this is possible with the short list of covariates, given the 14 models
presented one might expect to see at least one model where one of the covariates did not remain.
Schwartz (1995) Total subjects not specified, men and
15 prior U.S. and women ages 18 to 74 years. Random
European studies effects meta-analysis with inverse variance
published between weighting of lead-blood pressure
1985 and 1993 coefficients from each study. Sensitivity
analysis performed by dropping study with
largest or smallest effect.
Blood lead levels not
stated.
Each doubling of natural log blood lead level was associated with an increase of 1.25 mmHg (95% CI:
0.87, 1.63) systolic blood pressure. Sensitivity analysis showed negligible change in meta-analysis
coefficient. Concluded that adding newer studies would not change calculated coefficient. Noted
lead-blood pressure slope was larger at lower lead levels than at higher lead levels
The study only analyzed systolic, not diastolic, blood pressure. Superseded by Nawrot et al. (2002).
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Table AX6-5.1 (cont'd). Cardiovascular Effects of Lead
to
o
o
Reference, Study
Location, and
Period
Study Description
Lead Measurement
Findings, Interpretation
O
o
2
o
H
O
Blood Pressure, Hypertension
United States (cont'd)
Schwartz (1991) Under 10,000 subjects (exact number not
NHANES II reported), males and females, aged 25 to 74
U.S. years for left ventricular hypertrophy
1976-1980 results with logistic regression. Linear
blood lead used for LVH. For blood
pressure results, multiple linear regressions
stratified by sex, with one block of
variables forced, another block of variables
entered with stepwise regression, aged 6
months to 74 years, exact number not
given. Natural log blood lead used for
linear regression. Both logistic and linear
regressions adjusted for survey design.
No blood lead
descriptive data given.
Used logistic regression to study lead effect on left ventricular hypertrophy (LVH) determined by
a combination of electrocardiogram parameters and body habitus, controlling for age, race, and
sex. Every 10 ug/dL blood lead increase was associated with increased odds of LVH of 1.33
(95% CI: 1.20, 1.47). Interaction terms for race by blood lead and sex by blood lead were not
significant.
Blood pressure models stratified by sex always included BMI, age and age-squared, race, and
natural log blood lead. Male blood pressure model also included family history of hypertension,
cholesterol, height, cigarette use, serum zinc, and tricep skin fold. Female model also included
serum zinc, family history of hypertension, tricep skin fold, and cholesterol. Every 1 natural log
unit of blood lead increase was associated with an increase in diastolic blood pressure of 2.93
mmHg(95%CI: 0.93, 4.98) in males and 1.64 mmHg (95% CI: 0.27,3.01). Used interaction
terms for race-blood lead and sex-blood lead in a non-stratified model and found no significant
effect of race or sex on the blood lead-blood pressure coefficient.
Incomplete reporting of subject size for models and for descriptive statistics for all variables in
models. Tested both linear and log transformed lead in preliminary testing. Found log lead had
lower probability values than linear lead for blood pressure, and linear lead had lower probability
values than log lead for LVH. No testing of significant difference between the two blood lead
specifications. No model diagnostics reported. Only reported diastolic blood pressure results.
o
HH
H
W
Sokas et al. (1997) 264 active or retired construction workers,
U.S.-Maryland over 99% men, who were not involved in
1989-1990 lead work at time of testing, mean age
(range) 43 years (18-79). Multiple
regression modeling of systolic and
diastolic blood pressure adjusted for
covariates of BMI, age, hematocrit,
erythrocyte protoporphyrin, race, linear
blood lead and a race-linear blood lead
interaction. Method of covariate entry not
made explicit, though it appeared to be
forced.
Mean blood lead Linear blood lead was not significantly related to either systolic or diastolic blood pressure,
(range): 8.0 ug/dL though the race by linear blood lead interaction was marginally significant (p = 0.09). Each
(1-30). 1 ug/dL increase in blood lead increased black systolic blood pressure 0.86 mmHg (no SE or
95% CI reported) more than white systolic blood pressure.
Linear blood lead term may not be appropriate. Small sample compromises interpretation of
non-significant results. By using erythrocyte protoporphyrin and blood lead in the same model,
these two measures of lead exposure may have been confounded. Incomplete reporting of
procedures and results. No model diagnostic tests reported.
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Table AX6-5.1 (cont'd). Cardiovascular Effects of Lead
to
o
o
Reference, Study
Location, and
Period
Study Description
Lead Measurement
Findings, Interpretation
Blood Pressure, Hypertension
United States (cont'd)
O
o
2
o
H
O
Sorel et al. (1991) 2056 females, 2044 males, 473 blacks and
U.S.-NHANES II 3627 whites, from 18-74 years, were used
1976-1980 in survey design and weight adjusted
multiple linear regressions stratified by sex,
with separate models for systolic and
diastolic blood pressure. Covariates
included age, BMI, race, and poverty
income ratio and linear blood lead. Method
of covariate entry not specified but may
have been forced. Different covariate
groups were used for different models.
Primary test for the effect of race on the
lead-blood pressure relationship was to note
the change in the race coefficient in models
with and without the linear blood lead
variable.
Age-adjusted arithmetic
mean blood lead:
Black female: 13.2
ug/dL (no variance
information for any
blood lead)
White female: 12.1
ug/dL
Black male: 20.1 ug/dL
White male: 16.8
ug/dL
Linear blood lead was significantly related only to diastolic blood pressure in males, adjusting for
age and BMI. For every 1 ug/dL blood lead increase diastolic blood pressure increased 0.13
mmHg (95% CI: 0.04, 0.21). Adding race to the model with and without linear blood lead terms
did not appear to change the race coefficient. Adding poverty index to the models with and
without blood lead produced the same small change in poverty index coefficient.
Linear blood lead may not be appropriate. Only confidence intervals were used to assess the
significance of changes in race and poverty index coefficients across models with and without
lead, instead of using interaction terms of these two variables with lead. Incomplete reporting of
procedures and results. No model diagnostic tests reported.
o
HH
H
W
Sharp et al. (1990) After exclusion of subjects under treatment
U.S. -San Francisco, for hypertension, 249 male bus drivers, 132
CA of whom were black, age from 31 to 65
1986 years, were used in race stratified multiple
regression models of systolic and diastolic
blood pressure with covariate forced entry
of age, age-squared, BMI, caffeine use,
tobacco use, and natural log blood lead.
Alcohol use was added in other models.
Other models stratified by caffeine use.
Geometric mean (range)
blood lead:
Black males: 6.5 ug/dL
(3-21)
Non-black males:
6.2ug/dL(2-15)
Significant log blood lead effects were noted in blacks. In models excluding alcohol use, for
every one natural log unit increase of blood lead, systolic blood pressure rose 7.53 mmHg (95%
CI: 0.86, 14.2) and diastolic blood pressure rose 4.72 mmHg (95% CI: 0.15,9.29). Stratified by
infrequent/frequent caffeine users, only black infrequent caffeine users showed a significant
response to blood lead. For every one natural log unit increase of blood lead, systolic blood
pressure rose 16.69 mmHg (95% CI: 3.83, 29.5) and diastolic blood pressure rose 10.43 mmHg
(95% CI: 1.26, 19.6). Non-black blood pressure was decreased with increasing natural log lead
but was marginally significant. In all non-black subjects, for every unit increase in natural log
blood lead, systolic blood pressure decreased -5.71 mmHg (95% CI: -12.0, 0.6). Addition of
alcohol to the models decreased all coefficients a small amount. Progressive addition of age,
BMI, caffeine, and tobacco, in that order, progressively increased the coefficient of natural log
blood lead in models of systolic and diastolic blood pressure in blacks. Removal of two black
outliers did not materially change the results for blacks.
No statistical tests for comparing stratified models, models with and without caffeine use, effect
of progressive addition of covariates, or addition of alcohol. Influence diagnostics reported for
detecting the two outlying subjects. No other diagnostic tests reported. Small differences in text
and table reports of the same coefficients. Small sample size limits interpretation of non-
significant results.
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Table AX6-5.1 (cont'd). Cardiovascular Effects of Lead
to
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Reference, Study
Location, and
Period
Study Description
Lead Measurement
Findings, Interpretation
O
o
2
o
H
O
o
HH
H
W
Blood Pressure, Hypertension
United States (cont'd)
Tepperetal. (2001)
U.S.-Cincinnati, OH
After 1991 to before
2001
Vupputuri et al.
(2003)
U.S.-NHANES III
1988-1994
43 females and 57 males, current or former
workers at a lead-acid battery factory,
between 36 and 73 years of age, with at
least 10 years working in battery
production, participated. Multivariate
regression models and logistic regression
models were constructed to assess lead
exposure effect on outcome (hypertension:
>140/90 mm Hg and >160/95 mm Hg or
taking antihypertensive meds; diastolic and
systolic blood pressure, and left ventricular
mass/body surface area (g/m ).
Echocardiograms were used to determine
left ventricular mass. Variables used to
adjust all models were age, BMI, sex, and
family history of hypertension.
5188 white women, 2300 black women,
5360 white men, and 2104 black men, aged
18 years and older. Survey adjusted
multiple linear and logistic regression were
used to assess linear blood lead effect on
systolic and diastolic blood pressure and
hypertension in race and sex stratified
models.
Plant blood lead records were used
to calculate cumulative blood lead
index (CBLI) used as a tertile
measure, a linear continuous
measure, and a log transformed
measure.
CBLI Hg/dL-yr
1st tertile: 138-504
2ntl tertile: 505-746
3rd tertile: 747-1447
Time averaged blood lead TABL)
was treated the same way:
TABL
1st tertile:
2ntl tertile:
3rd tertile:
ug/dL
12-25
26-33
34-50
Arithmetic mean (SD) blood lead:
White women
3.0 ug/dL (7.2)
Black women 3.4 ug/dL (4.8)
White men 4.4 ug/dL (7-3)
Black men 5.4 ug/dL (9.3)
No odds ratios were given for hypertension and any lead variable for hypertension defined as
>140/90 mm Hg but ORs were claimed not significant. Odds ratios were 2.71 and 1.44 for the
third tertile CBLI and TABL lead measures compared to first tertile, apparently significant, but
no probabilities, SEs or CIs given.
With the 81 subjects not taking anti-hypertensive meds, neither CBLI tertile nor TABL tertile
were significantly associated with either diastolic or systolic blood pressure (coefficients, SB's or
95% CIs not given). Using log transformed CBLI probability of a positive association with
diastolic blood pressure was 0.06. Using log transformed TABL, probability of a positive
association with diastolic blood pressure was 0.10. No coefficients, SEs, or CIs given.
Left ventricular mass adjusted for body surface area was not significantly related to any lead
measure. No coefficients, SEs or CIs given.
Despite the certainty of the authors that "we found no convincing evidence of an association...",
the very low power of this study gives certainty to none of the findings. Very poor reporting of
results further reduces the possibility of evaluation. No model diagnostic testing was reported.
Multiple linear regression models were all adjusted for age, education, BMI, alcohol
consumption, leisure time physical activity, dietary sodium and potassium, and total calories.
Only black women and men showed significant linear lead effects. Every 1 ug/dL increase in
blood lead was associated with an increase of 0.47 mmHg (95% CI: 0.14, 0.80) in systolic and
0.32 mmHg (95% CI: 0.11, 0.54) diastolic blood pressure in black women, and 0.25 mmHg
(95% CI: 0.06, 0.44) systolic and 0.19 mmHg (95% CI: 0.02, 0.36) diastolic blood pressure in
black men.
Odds of hypertension (systolic >140 mmHg, diastolic >90 mmHg, or taking antihypertensive
medication) significantly increased for every SD (3.3 ug/dL) of blood lead level in black women
(OR 1.39 [95% CI: 1.21, 1.61]), in white women (OR 1.32 [95% CI: 1.14, 1.52]), in black men
(OR 1.26 [95% CI: 0.99, 1.19]), but not in white men.
Linear blood lead terms are usually not appropriate in multiple linear regression models of blood
pressure. Furthermore, they reported their results in terms of change in 1 SD unit of lead. Linear
SD of lead is incorrect for log-normal distributions of blood lead. No model diagnostic tests
reported. Discrepancy between Methods report of race-lead and sex-lead interactions in simple,
not multiple, analyses, but Results reports significant interactions for race-lead and sex-lead in
multiple regression models for both linear regression and logistic regression models, without
showing the results of the interaction analyses. The probability of the stated interactions
(p < 0.001) appears extremely low, given the degree of 95% CI overlap in lead coefficients
among the stratified models. No model diagnostics reported.
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Table AX6-5.1 (cont'd). Cardiovascular Effects of Lead
to
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o
Reference, Study
Location, and
Period
Study Description
Lead Measurement
Findings, Interpretation
O
o
2
o
H
O
o
HH
H
W
Blood Pressure, Hypertension
Europe
Bost et al. (1999) 2763 women and 2563 men from a multi-
Europe-England- stage stratified probability survey
Health Survey for representative of the English population
England living in private residences, mean (SE) age
1995 for men 47.5 years (0.34) and for women
47.7 years (0.33) (all subjects 16 years and
older) were used in an analysis of blood
lead association with systolic and diastolic
blood pressure. Stepwise multiple
regression analysis were used testing
natural log blood lead against common log
systolic blood pressure and non-
transformed diastolic blood pressure, with
the following potential covariates: age,
BMI, smoking status, region of residence,
social class, and alcohol consumption.
Models were stratified by sex, with and
without adjustment for alcohol, including or
excluding those taking antihypertensive
medications.
Factor-Litvak et al. 281 5.5-year old children studied since
(1996) pregnancy participated. Multiple linear
Europe-Kosovska regression models of systolic blood
Mitrovica and pressure, adjusted for height, BMI, gender,
Pristina, Kosovo ethnic group, and birth order, and diastolic
—1992-1993 blood pressure, adjusted for waist
circumference, ethnic group, and birth order
were constructed by stepwise elimination
from a larger pool of potential confounding
variables and retained if they modified the
linear blood lead coefficient by more the
10%.
Geometric mean blood
lead:
Men: 3.7 ug/dL(no
stated measure of
variance)
Women: 2.6 ug/dL
(no stated measure of
variance
Blood lead arithmetic
mean: 22.7 ug/dL
Range: 5-55 ug/dL
Model tables presented only standardized variable coefficients. The most consistent results were
reported on common log lead association with men's diastolic blood pressure. Every doubling of
blood lead was significantly associated with an increase of 0.78 mmHg (95% CI: 0.01, 1.55) diastolic
blood pressure, adjusted for age, log BMI, and alcohol, but excluding men on antihypertensive
medication. Every doubling of blood lead was significantly associated with an increase of 0.88 mmHg
(95% CI: 0.13, 1.63) in the same model with men on antihypertensive medication. Every doubling of
blood lead was significantly associated with an increase of 0.96 mmHg (95% CI: 0.23, 1.70) in the
same model excluding men on antihypertensive medication and not adjusting for alcohol. Every
doubling of blood lead was significantly associated with an increase of 1.07 mmHg (95% CI: 0.37,
1.78) including men taking antihypertensive medication and not accounting for alcohol. None of the
multiple regression models had significant lead terms for women.
This report was not sufficiently detailed. Stepwise regression modeling is prone to the usual pitfalls.
Survey design adjusted analysis not used. Lead was not entered in models in which criterion
probability was exceeded (p > 0.05). No rationale given for stratifying. No testing of differences
among lead coefficients for the different models was made, which would have been especially
valuable to compare models adjusted and not adjusted for alcohol use. No explanation for using log
systolic blood pressure as dependent variable. No model diagnostics reported.
For each increase of 1 ug/dL of blood lead:
Systolic beta = 0.054 (95% CI: -0.024, 0.13)
Diastolic beta = 0.042 (95% CI: -0.01, 0.090)
Despite low power to detect significant effects, there was a marginally significant tendency for blood
pressure to be positively associated with blood lead. Stepwise multiple regression may have
capitalized on chance results. The linear lead term may have reduced the ability to detect significant
effects of lead if the modeled relationship were non-linear. Log lead was tested but not reported. A
quadratic lead term was reported non-significant. No model diagnostics reported.
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Table AX6-5.1 (cont'd). Cardiovascular Effects of Lead
to
o
o
Reference, Study
Location, and
Period
Study Description
Lead Measurement
Findings, Interpretation
Blood Pressure, Hypertension
Europe (cont'd)
O
o
2
o
H
O
o
HH
H
W
Fewtrell et al. (2004)
Global
1988-2002
Maheswaran, et al.
(1993)
Europe-England-
Birmingham
1981
Using available global figures on
categorized blood lead ranges by age group,
authors calculated relative risk ratios
relating increased blood pressure to
ischemic heart disease, cerebrovascular
disease, hypertensive disease, and other
cardiac diseases. They used a calculation
of "impact fraction," based on the
proportion of the population within the
particular lead exposure category and the
relative risk at that exposure category
compared to the risk at the reference level.
They used the meta-analysis of Schwartz
(1995) to derive an accumulating 1.25
mmHg increase in blood pressure in men
for 5-10, 10-15, and 15-20 ug/dL, and an
increase of 3.75 mmHg for blood lead
levels above 20 ug/dL. Comparable blood
pressure increases in women for each lead
category was 0.8 mmHg for each of the
first three categories and 2.4 mmHg for
blood lead >20 ug/dL.
809 out of 870 workers, mean (SD) age
43.3 (10.4) years, at an lead acid battery
plant were used in the study. Women and
workers taking antihypertensive
medications were excluded. Used multiple
linear regression analyses of systolic and
diastolic blood pressure, forcing age, BMI,
alcohol use, linear blood lead, zinc
protoporphyrin, years of work exposure,
cigarette smoking as covariates.
See left for blood lead
categories used.
Geometric mean (SD)
blood lead was:
31.6ug/dL(5.5)
The largest risk ratios were for hypertensive disease populations at ages 15-44, calculated at 1.12,
1.41, 1.78, and 2.00 for each of the four lead categories for men, and 1.08, 1.25, 1.45, and 1.56
for women. Risk ratios for all disease categories increased with increasing lead category and
decreased for populations older than 44 years.
The authors assumed a linear relationship between blood pressure and blood lead, whereas
available evidence suggests it may be non-linear. If blood lead-blood pressure concentration-
response function is log-linear, as implicitly accepted by over half the reviewed studies, the
calculated global risk ratios for all cardiovascular disease will be overestimated at higher blood
lead levels and underestimated at lower blood lead levels.
Linear blood lead was not significant for either systolic or diastolic blood pressure.
Authors used two indices of lead exposure in the same models. Over much of the studied blood
lead range, zinc protoporphyrin was likely collinear with blood lead. Linear blood lead may not
be the appropriate metric to use in blood pressure models. Did not use age-squared to adjust for
non-linear relationship of blood pressure with age. Did not report model diagnostics.
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Table AX6-5.1 (cont'd). Cardiovascular Effects of Lead
to
o
o
Reference, Study
Location, and
Period
Study Description
Lead Measurement
Findings, Interpretation
Blood Pressure, Hypertension
Europe (cont'd)
Menditto et al. (1994)
Europe-Rome-New Risk
Factors Survey
1989-1990
1319 males, mean (range) age
63 (55-75) years, not treated for
hypertension, were used in
forward stepwise multiple linear
regression models of systolic
and diastolic blood pressure
with available covariates of age,
BMI, heart rate, serum high
density lipoprotein, non-high
density lipoprotein,
triglycerides, glucose, cigarette
use, alcohol use, sum of five
skinfold thicknesses (triceps,
biceps, subscapular,
suprascapular, and suprailiac),
and natural log transformed
blood lead.
Median (2.5th-97.5th
percentiles, range) blood lead
11.3 ug/dL (6.2-24.7, 4-44.2)
Only BMI, heart rate, and serum glucose were not simultaneously and significantly correlated
with both natural log blood lead and blood pressure. In a systolic blood pressure model adjusted
for BMI, age, heart rate, high and non-high density lipoprotein, triglycerides, glucose, and
cigarettes, each unit increase in natural log blood lead was significantly associated with a
5.6 mmHg (95% CI: neither SE nor CI stated) increase in blood pressure. In a diastolic blood
pressure model adjusted for BMI, heart rate, age, cigarettes, triglycerides, and high density
lipoprotein, each unit increase in natural log blood lead was significantly associated with a
1.7 mmHg (95% CI: neither SE nor CI stated) increase in blood pressure. In stratified models
for alcohol drinkers (n = 1068) and non-drinkers (n = 251) only alcohol drinkers showed
significant natural log blood lead associated blood pressure increase, with lead coefficients
similar to those of the entire group.
Authors observed change in natural log blood lead coefficient produced by successive addition of
covariates to models. In no case did the coefficients change by more than 30% after addition of a
covariate. Authors noted that wine was the predominant drink in alcohol users and that the
correlation between alcohol consumption and natural log blood lead level was the highest among
all correlations reported (p < 0.001; correlation coefficient not stated).
No statistical tests were made to determine if the change in lead coefficients with addition of
covariates was significant, nor were statistical tests made to determine if the lead coefficients in
the alcohol use stratified models were significantly different. Small size of the non-alcohol
drinking group in stratified analysis precludes interpretation of non-significant effects.
Incomplete reporting of results. Paper published in a supplement issue reporting meeting papers
may indicate that it received less that the normal peer-review scrutiny for published research
articles. No model diagnostic tests were reported.
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Table AX6-5.1 (cont'd). Cardiovascular Effects of Lead
to
o
o
Reference, Study
Location, and
Period
Study Description
Lead Measurement
Findings, Interpretation
O
o
2
o
H
O
o
HH
H
W
Blood Pressure, Hypertension
Europe (cont'd)
M011er and
Kristensen (1992)
Europe-Denmark-
Copenhagen County-
Glostrup Population
Studies
1976-1990
A cohort born in 1936 was followed at age
40 (women n = 546, men n = 504), age 45
(women n = 430, men n = 463) and again at
age 51 (men only n = 439). Reported no
difference in results if subjects taking
antihypertensive medications were
excluded. Reported results included these
subjects. Linear multiple regression models
of systolic and diastolic blood pressure of
follow up, stratified by sex and by year,
used a sequence of forced entry of
covariates: natural log blood lead was
tested alone (unadjusted), then adjusted for
tobacco, cholesterol, physical activity, and
sex (Model 1), then adjusted for the above
covariates plus systolic blood pressure
(Model 2), and then adjusted for the above
covariates plus alcohol (Model 3). Another
group of linear multiple regression models
of change of systolic and diastolic blood
pressure from age 40 to 51 years in men
only, following the same covariate entry
scheme as above, but used change in
covariates instead of the original covariates.
All subjects were followed until 54 years of
age (from 1976 to 1990) to assess lead
association with total mortality and with
coronary heart disease (CHD; ICD-8 410-
414) and cardiovascular disease (CVD;
ICD-8 430-435) combined morbidity and
mortality using Cox proportional hazards
models (n = 1050). Cox models were
adjusted as above.
Arithmetic mean (SD, range)
blood lead by age and sex:
Women 40 years: 9.6 ug/dL
(3.8) [4-39]
Women 45 years: 6.8 ug/dL
(3.5)[2-41]
Men 40 years: 13.6 ug/dL (5.7)
[5-60]
Men 45 years: 9.6 ug/dL (4.3)
[3-39]
Men 51 years: 8.3 ug/dL (4.1)
[2-62]
In women, each one unit increase in natural log blood lead was associated with a
significant increase in systolic blood pressure of 4.93 mmHg (p = 0.002; neither SE nor
CI stated) at age 40 and an increase of 2.64 mmHg (p = 0.06; neither SE nor CI stated) at
age 45, in models adjusted for tobacco, BMI, and physical activity (Model 1). When
alcohol (Model 2) or alcohol plus hemoglobin (Model 3) were added to the models lead-
blood pressure relationships were not significant at either age. With each one unit
change in natural log blood lead, diastolic pressure increased 4.26 mmHg (p = 0.002;
neither SE nor CI stated) at 40 years and 3.26 mmHg (p = 0.002; neither SE nor CI
stated) at 45 years in Model 1. In Model 2, the increase in diastolic blood pressure was
3.21 mmHg (p = 0.02; neither SE nor CI stated) at 40 years and 2.86 mmHg (p = 0.01;
neither SE nor CI stated) at 45 years. In Model 3, the increase in diastolic blood
pressure was 2.65 mmHg (p = 0.07; neither SE nor CI stated) at 40 years and 2.78
mmHg (p = 0.01; neither SE nor CI stated) at 45 years.
In men, the only significant association between natural log blood lead and blood
pressure was at 45 years. For every increase of one unit of natural log blood lead the
increase in systolic blood pressure was 2.73 mmHg (p = 0.05; neither SE nor CI stated).
The change in blood lead between 40 and 51 years was not significantly associated with
change in systolic or diastolic blood over the same period in any of the models.
None of the relative hazard ratios for CHD and DVD combined morbidity and mortality
between 40 and 54 years were significantly related to blood lead concentration. Total
mortality, however, was significantly increased with increased blood lead. In Model 1,
every increase of one natural log unit of blood lead was associated with an increased
relative hazard of mortality of 1.96 (p = 0.009; neither SE nor CI stated). For Model 2,
every increase of one natural log unit of blood lead was associated with an increased
relative hazard of mortality of 1.82 (p = 0.03; neither SE nor CI stated). There were 40
cases of CHD recorded, of which 13 were fatal. There were 54 cases of CVD recorded,
of which 19 were fatal. Of the total of 46 subjects who died during the period, 32 (70%)
died of cardiovascular problems. It was not clear if blood lead at a particular age or a
mean blood lead across ages was used in the Cox proportional hazards models.
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Table AX6-5.1 (cont'd). Cardiovascular Effects of Lead
to
o
o
Reference, Study
Location, and
Period
Study Description
Lead Measurement
Findings, Interpretation
Blood Pressure, Hypertension
Europe (cont'd)
M011er and
Kristensen (1992)
(cont'd)
Though this study was one of the few to use a longitudinal design, it did not take
advantage of that design feature in blood pressure modeling. Cross-sectional multiple
regression modeling at each age loses valuable information available in repeated
measures modeling. Power to detect significant effects is much higher in repeated
measurement modeling than in cross-sectional modeling. Analyzing only change in
blood pressure loses information regarding starting and ending blood pressure.
Including change in blood lead is problematical due to the unknown history of lead
exposure prior to the start of the study, the resultant bone lead load as a result of past
exposure, the unknown lead contribution of bone to blood, and the unknown relative
contributions of past exposure and present exposure to alteration in blood pressure.
Modeling other covariates as change is also questionable. BMI, to pick a covariate with
known and strong effects on blood pressure, may be high and relatively constant over the
study period or low and relatively constant over the study. In both cases, the change in
BMI will be small, but the high BMI will be associated with higher blood pressure than
will the low BMI. Thus, both cases modeled as change in BMI should have the same
effect on blood pressure when the high BMI subject has expected higher blood pressure
than the low BMI subject. Using difference scores for the dependent and the exposure
variables also risks confounding secular trends in either or both of these variables, for
whatever reasons, with independent difference variable effect on dependent difference
variable effect.
The Cox proportional hazards model, however, is longitudinal in nature. Failure to
detect significant associations between lead and cardiovascular morbidity/mortality
could have been due to the small sample size used for this type of analysis. The blood
pressure part of the study did not take mortality into account during the study, which
could have produced a progressively increasing "healthy subject" effect. Since subjects
taking antihypertensive medications were included in analyses, an indicator variable
should have been used to account for them, whether or not their exclusion in preliminary
testing produced no apparent change in results. This paper contained a good discussion
of confounding variables. Incomplete reporting of results and procedures. No model
diagnostic tests were reported.
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Table AX6-5.1 (cont'd). Cardiovascular Effects of Lead
to
o
o
Reference, Study
Location, and
Period
Study Description
Lead Measurement
Findings, Interpretation
O
o
2
o
H
O
o
HH
H
W
Blood Pressure, Hypertension
Europe (cont'd)
Staessen et al. 359 men and 369 women participated
(1996a) at baseline (between 1985 and 1989)
Europe-Belgium- and again about 5 years later (median
PheeCad study. 5.2 years) at follow up (between 1991
1985-1995 and 1995), mean age (range) at
baseline 46 years (20-82), about half
of whom were recruited from towns
surrounding a non-ferrous smelter
(targeted to produce high cadmium
exposure) and half from towns
without heavy metal production.
Over half the men had occupational
exposure (59.0% from the near
smelter towns, 17.4% from the other
towns).
Four different outcomes were
explored: time-integrated
conventional blood pressure (average
of 10 baseline and 5 follow up blood
pressure measurements), 24-hour
ambulatory blood pressure only
during the follow up period (average
of readings every 20 minutes from 8
AM to 10 PM and every 45 minutes
from 10 PM to 8 AM, weighted by
interval between measurements),
difference in conventional blood
pressure over the five year follow up
period, and incidence of developing
hypertension during follow up.
Geometric mean (5%-95%
percentile) by sex and time
period:
Baseline women:
6.6 ug/dL (3.3-14.5)
Follow up women:
4.8 ug/dL (1.7-11.8)
Baseline men: 11.4ug/dL
(5.6-28.8)
Follow up men: 7.7 ug/dL
(3.7-20.1)
The study was one of the few prospective longitudinal studies reported and was innovative in its use
of 24-hour ambulatory blood pressure as one of its outcome variables.
Time-integrated conventional blood pressure models:
In 187 peri- and post-menopausal women, after adjusting for age, BMI, gamma-glutamyltransferase
activity, and hematocrit, each increase of one unit of natural log blood lead was associated with an
increase in diastolic blood pressure of 7.49 mmHg (95% CI: 1.48, 13.50). No other time-integrated
conventional blood pressure measurements were significantly associated with time-integrated
natural log blood lead in either men or women, nor in stratified groups within sex.
Ambulatory 24-hour blood pressure models:
In all 345 women, after adjusting for age, hematocrit, gamma-glutamyltransferase activity, and oral
contraceptive use, each one unit increase in natural log blood lead was associated with an increase
of diastolic blood pressure of 3.49 mmHg (95% CI: 0.02, 6.96). When the group was limited to the
174 premenopausal women each unit increase in natural log blood lead was associated with an
increase of diastolic blood pressure of 5.48 mmHg (95% CI: 0.56, 10.40).
Difference in blood pressure between baseline and follow up:
After adjustment for change in BMI, beginning use of antihypertensive medication and
contraceptive medication during the follow up period, and starting smoking there was no significant
relationship between difference in either systolic or diastolic blood pressure and blood lead in
women. After adjustment for change in BMI, change in exposure at work, change in smoking,
beginning use of antihypertensive medication in men there was no significant relationship between
difference in either systolic or diastolic blood pressure and blood lead in men.
Incidence of hypertension:
At baseline 107 (14.7%) and 120 (16.5%) subjects had borderline and definite hypertension,
respectively. At follow up 98 (13.5%) and 186 (25.5%) had borderline and definite hypertension,
respectively. 51 of 501 initially normotensive subjects became borderline hypertensive and 47 of
the 501 became border line hypertensive during the follow up period. After adjusting for sex, age,
and BMI, natural log baseline blood lead was not related to significant risk ratios of becoming
hypertensive (not stated, but presumably combined definite and borderline hypertension) or
becoming a definite hypertensive.
-------
Table AX6-5.1 (cont'd). Cardiovascular Effects of Lead
to
o
o
Reference, Study
Location, and
Period
Study Description
Lead Measurement
Findings, Interpretation
O
o
2
o
H
O
o
HH
H
W
Blood Pressure, Hypertension
Europe (cont'd)
Staessen et al. Multiple regression models were used to test the
(1996a) (cont'd) association between natural log transformed blood lead
(mean of baseline and follow up lead) and blood
pressure (systolic and diastolic), stratified by sex, then
further stratified by use of antihypertensive
medications in men and menopausal status in women.
Age and age-squared (calculated in quintiles) were
forced into the models, then remaining covariates were
stepwise added to the model. Though not explicitly
stated, natural log blood lead (mean of baseline and
follow up) was likely forced in last. Other candidate
covariates were BMI, hemoglobin or hematocrit,
serum gamma-glutamyltransferase activity (an index of
alcohol use) and serum calcium, 24 hour urinary
sodium and potassium excretion, energy expenditure,
exposure to heavy metals (at the workplace), social
class, smoking and drinking habits, menstrual status in
women, and use of antihypertensive medications, oral
contraceptives, and hormone replacement therapy. In
ambulatory blood pressure models, differences
between baseline and follow up blood pressure models
were constructed in the same way. For the difference
models "concurrent variations in blood lead
concentrations" were used, presumably difference in
baseline and blood lead. For the hypertension
incidence model two definitions of hypertension were
used: definite hypertension (systolic >160 mmHg,
diastolic >95 mmHg or taking antihypertensive
medications) and borderline hypertension (systolic
between 141 to 159 mmHg and diastolic between 91 to
94 mmHg). Method of covariate entry into
hypertension incidence models not stated. Baseline
natural log blood lead was used as the exposure index.
The study does not use the full power of repeated measurements in the analyses. For
problems encountered when collapsing repeated measurements to difference measures, see
M011er (1992) above. Stepwise regressions are prone to capitalizing on chance results due
to multiple testing of the same data and almost always produce a different mix of covariates
when they are stratified. Thus, it was puzzling to find that where information on the effects
of stepwise covariate addition to models was available in this article, that the same
covariates were listed for both models based on the stratification variable. There is
excessive reliance on fractionation of the data set due to multiple stratification, sometimes
reducing the number of subjects in a model to as few as 171. Even the models using the
most subjects had only 359 subjects. Low power to detect significant effects cautions
against any interpretation of non-significant results. The time-integrated model used
10 baseline blood pressure measurements and 5 follow up blood pressure measurements,
thus weighting the average toward baseline blood pressure. The entry of the biochemical
correlate of alcohol use in most of the models suggests that lead effects and lead-containing
alcohol effects on blood pressure were confused, especially given the European setting and
the time period during which the study was conducted. Control for use of hypertensive
medication rarely entered models and partial control for this variable was achieved only by
stratified analyses, further reducing power to detect significant effects in the remaining
subgroup. No justification was given for stratified analyses. Incomplete information in
statistical methods and results complicates interpretation. It was uncertain if stepwise
regression was used for logistic models. No comparisons were performed to assess
possible bias due to subject attrition over the course of the study. The over six decades of
age represented in the sample was modeled by linear and quadratic terms based on age
quintiles rather than continuous age, making it likely that adequate control for age effects
on blood pressure was not achieved and that the "healthy subject" effect seen in older
groups was not controlled. If stepwise addition of significant covariates was used in the
blood pressure difference models, were covariates in those models that were marked in the
coefficient column as non-significant not included in the models, and if that were so, it is
unclear from where the probability values that substitute for the coefficients of those
variables were derived. There were no model diagnostic tests reported.
-------
Table AX6-5.1 (cont'd). Cardiovascular Effects of Lead
to
o
o
Reference, Study
Location, and
Period
Study Description
Lead Measurement
Findings, Interpretation
O
o
2
o
H
O
o
HH
H
W
Blood Pressure, Hypertension
Europe (cont'd)
Staessen et al. (1993)
Belgium-Cadmibel
Study
1985-1989
827 males and 821 females recruited from two areas
in Belgium, one of them surrounding a non-ferrous
smelter, mean age (SD) 46 (15) and 44 (15) years,
in men and women respectively. Subjects taking
antihypertensive medication were excluded from the
analyses. Stepwise multiple regression models of
systolic and diastolic blood pressure were stratified
by sex. Covariates available for entry were age and
age-squared, BMI, pulse rate, log protoporphyrin,
log gamma-glutamyltranspeptidase, serum calcium,
log serum ferritin, log serum creatinine, log serum
zinc, urinary calcium, urinary sodium, and urinary
potassium. Natural log blood lead was the only
variable forced into the models. Additional models
tested the interaction of serum calcium and blood
lead on blood pressure.
Geometric mean
blood lead (range),
stratified by sex:
Male blood lead
10.4 ug/dL
(2.7, 84.9)
Female blood lead
6.2 ug/dL (1.3, 42.4)
In men, adjusting for age and age-squared, BMI, pulse rate, log gamma-
glutamyltranspeptidase, serum calcium, and log serum creatinine, every unit natural log
blood lead increase was significantly associated with a -5.2 mmHg (95% CI: -0.5, -9.9)
decrease in systolic blood pressure. Natural log blood lead was not significant in the model
for diastolic blood pressure for men nor the systolic or diastolic blood pressure for women.
Adjusting for age and age-squared, BMI, pulse rate, and log gamma-
glutamyltranspeptidase, the interaction term between natural log blood lead and serum
calcium was only significant for systolic blood pressure in women. Every doubling of
blood lead was associated with a 1.0 mmHg decrease in systolic blood pressure at serum
calcium concentration of 2.31 mmol/L (25th percentile) and an increase in systolic blood
pressure of 1.5 mmHg at serum calcium concentration of 2.42 mmol/L (75th percentile).
Stepwise multiple regression analyses run risks of accepting chance associations due to
multiple analyses of the same data set. The role of alcohol use or alcohol use markers in
confounding lead effect on blood pressure in this setting has already been noted. The
unexplained interaction between serum calcium and blood lead highlights the potential
confounding role of serum calcium with lead in blood pressure studies. The study shows
graphs indicating distinct differences in the age-serum calcium and age-blood lead
relationships for men and women. From 50-70 years of age serum calcium is higher than
from <29-49 years in women and exceeds serum calcium of men at those older ages. The
steepest rise in women's blood lead with age occurs between the 40-49 and 50-59 year
decades. The timing of these changes in women suggests that menopause may be a factor,
which was accounted for only in the model for diastolic blood pressure. It also suggests
that serum calcium level and age were also confounded in the blood pressure models.
As serum creatinine clearance and blood lead are inversely related, and serum creatinine is
a significant covariate in the systolic blood pressure model for men with a significant
negative blood lead coefficient, it is possible that serum creatinine and blood lead are
confounded with blood pressure in the men's systolic blood pressure model. There were no
assessments of subject selection bias due to exclusions. The authors note examining
quintile blood pressure relationships with all covariates to determine the acceptability of the
linear relationship implied by the linear modeling technique. No other model diagnostic
tests were reported.
-------
Table AX6-5.1 (cont'd). Cardiovascular Effects of Lead
to
o
o
Reference, Study
Location, and
Period
Study Description
Lead Measurement
Findings, Interpretation
Blood Pressure, Hypertension
Europe (cont'd)
Telisman et al.
(2004)
Europe-Croatia-
Zagreb
Date of data
collection not given.
100 workers from factories producing lead-based
products, mean (range) age 30 (20-43) years.
Exclusion criteria were absence of psychological
stress (e.g., death in family) over last 4 months,
absence of verified diabetes, coronary heart disease,
cerebrovascular and peripheral vascular disease,
renal disease, hyperthyroidism, androgenital
syndrome, primary aldosteronism, and "other
diseases that could influence blood pressure or
metal metabolism." Linear or natural log blood lead
were considered for stepwise entry in models of
systolic and diastolic blood pressure, forcing in all
other covariates: blood cadmium, BMI, age, serum
zinc, serum copper, hematocrit, smoking, and
alcohol.
Arithmetic mean Neither linear nor natural log blood lead entered as significant in multiple regression
(range) blood lead: models of systolic and diastolic blood pressure.
36.7 ug/dL Very small sample size limited power to detect significant effects; non-significant effects
(9.9-65.9) should not be interpreted as lack of effect. Too many covariates for a small study. Almost
no subjects below 10 ug/dL. Taking hypertensive medications not controlled, likely a
problem with top systolic and diastolic blood pressure in the group 170 mmHg and
110 mmHg, respectively. No model diagnostic testing reported.
-------
Table AX6-5.1 (cont'd). Cardiovascular Effects of Lead
to
o
o
Reference, Study
Location, and
Period
Study Description
Lead Measurement
Findings, Interpretation
Blood Pressure, Hypertension
Asia
O
o
2
o
H
O
o
HH
H
W
Lee et al. (2001) 798 workers from various lead-using or producing
Korea-Chonan factories, mean (SD, range) age 40.5 years (10.1)
1997-1999 [17.8-64.8], 79.4% male, were classified as to Vitamin
D receptor genotype (VDR: bb or Bb/BB) and delta-
aminolevulinic acid dehydratase (ALAD: 1-1 or 1-2)
genotype, as VDR polymorphism has been implicated
in modifications of lead absorption and lead uptake
and release from bone as well as risk for elevated
blood pressure and hypertension, and ALAD
polymorphism affects lead binding to it in the
erythrocyte, the major storage depot of lead in blood.
The hypothesis was that polymorphism type could
influence the effect of lead on blood pressure and
hypertension.
Multiple linear regression models of linear blood lead,
DMSA-chelatable lead, and tibia lead effect on systolic
and diastolic blood pressure with potential covariates
of age and age-squared, sex, creatinine clearance,
hemoglobin, weight, height, BMI, job duration,
tobacco and alcohol consumption, pack-years of
tobacco, and cumulative life time alcoholic drinks.
Stepwise procedure allowed retention of covariates
only if they were significant or "there were substantive
changes in the coefficients of predictor variables after"
their inclusion. In the models shown, Appearance of
multiple lead variables and the interaction between
lead variables and genotype for each gene depended
upon the specific model. Both ALAD and VDR
receptor polymorphism were sometimes tested
simultaneously in each model containing
polymorphism terms and sometimes VDR appeared
without ALAD.
Arithmetic mean With simple t-tests, subjects with VDR Bb/BB allele were significantly older, had more
(SD, range) blood DMSA-chelatable lead, and had higher systolic and diastolic blood pressure than subjects
lead 32.0 ug/dL with VDR bb allele.
(15.0,4-86)
In multiple regression models of systolic blood pressure, controlling for age and age-
Mean (SD, range) squared, sex, BMI, antihypertensive medication use, and cumulative life-time alcoholic
DMSA-chelatable drinks, adding tibia lead, VDR type, and ALAD type, each increase of 10 ug/g of tibia lead
lead 186 ug was associated with an increase of 0.24 mmHg (95% CI: -0.01, 0.49) and VDR BB/Bb
(208.4, 4.8-2103) type was associated with an increase of 3.24 mmHg (95% CI: 0.18, 6.30) blood pressure
compared to the VDR bb type. ALAD genotype was not significant. In the same model,
Mean (SD, range) but substituting linear blood lead for tibia lead, each increase in 1 ug/dL of linear blood
tibia lead 37.2 ug/g lead was associated with an increase of 0.07 mmHg (95% CI: 0.00, 0.14) and VDR BB/Bb
(40.4, -7to 338) type was associated with an increase of 2.86 mmHg (95% CI: -0.22, 5.94) blood pressure
compared to the VDR bb type. ALAD genotype had no significant effects on blood
pressure.
When both tibia and linear blood lead were entered simultaneously along with VDR
genotype, adjusting for the same covariates, only VDR Bb/BB was significant; compared to
VDR bb, blood pressure was 3.51 mmHg (95% CI: 2.41, 8.61) higher. ALAD genotype
had no significant effects on blood pressure.
In a model without any lead terms, VDR genotype was interacted with the age and the age-
squared terms. The VDR Bb/BB genotype interaction with the linear age term was
significant for systolic blood pressure. Compared with the bb genotype the VDR Bb/BB
genotypes' blood pressure increased 0.36 mmHg (95% CI: 0.06, 0.66) per year faster with
increasing age.
There were no significant effects of any lead variable with diastolic blood pressure, though
the VDR Bb/BB genotype had significantly higher blood pressure (1.9 mmHg; not enough
information given to calculate CI) than the bb genotypes.
There were no significant interactions of the lead measures with the genotypes for either
ALAD or VDR.
-------
Table AX6-5.1 (cont'd). Cardiovascular Effects of Lead
to
o
o
Reference, Study
Location, and
Period
Study Description
Lead
Measurement
Findings, Interpretation
Blood Pressure, Hypertension
Asia (cont'd)
Lee etal. (2001)
(cont'd)
O
o
2
o
H
O
o
HH
H
W
Lustberg et al. (2004)
Korea-Chonan
1997-1999 (period of
enrollment; no
statement on dates of
data collection)
Logistic regression analysis was used to test the effect of
the lead indices on hypertension (systolic >160 mmHg
or diastolic >96 mmHg or taking antihypertensive
medications) using the same group of potential
covariates, testing the lead terms and the lead-genotype
interaction terms separately. The hypertension models
tested both gene polymorphisms separately.
793 (number given for genotype analysis; numbers in Lead according
models not given) current and former lead workers, to genotype:
mean (SD) age 40 (10) years and 80% male, were
genotyped for the three polymorphisms of endothelial Arithmetic mean
nitric oxide synthase (eNOS) (GG, GT, TT), an enzyme (SD) blood lead,
that is a modulator of vascular resistance. The effect of GG: 32(15)
genotype and the interaction of genotype with blood Hg/dL
lead and tibia lead on systolic and diastolic blood
pressure were evaluated by multiple linear regression Arithmetic mean
analyses, forcing covariates of age (modeled as a 2 (SD) blood lead,
degree of freedom spline with knot at 45 years), sex, TG/TT: 32(15)
natural log BMI, smoking and alcohol consumption, Hg/dL
high school education, and job duration. Both blood
lead and tibia lead were entered as percentiles and Mean (SD) tibia
entered together. Logistic models of hypertension lead, GG: 37
(systolic >140 mmHg or diastolic >90 mmHg or (42) ug/g
reported use of antihypertensive medication) used the
same covariates. Interaction terms between each of the Mean (SD) tibia
lead measures (plus a lead-squared term) and genotype lead, TC/TT: 36
was used to determine differential effect of lead (34) ug/g
according to genotype.
Subjects with the Bb/BB genotypes had a significantly higher odds hypertension prevalence (OR 2.1
[95% CI: 1.0, 4.4]) than subjects with the bb genotype, adjusting for age, sex, BMI, tibia lead, and
current alcohol use. There were no significant effects of any lead variable nor of ALAD on
hypertension status.
Linear blood lead may not give efficient and unbiased estimates of blood lead effect on blood pressure.
The descriptive data shows highly skewed distributions for blood lead, DMSA-chelatable lead, and
tibia lead in this group, suggesting that coefficients of all lead effect on blood pressure may not have
been efficient and unbiased. Stepwise models usually produce different covariate patterns for different
models, though the tables indicate that the covariates used for all the models discussed above were the
same. No model diagnostic tests were reported.
85% (673/793) of the group were typed GG, 14% (114/793) were TG, and 1% (6/793) were TT. TG
and TT groups were combined for analysis (TG/TT).
Mean systolic and diastolic blood pressures, adjusted for all covariates, were not significantly different
between GG and TG/TT groups.
In multiple regression models for systolic and diastolic blood pressure, neither percentile blood lead
nor percentile tibia lead, entered together, were significant predictors. Interaction terms between the
lead variables and genotype were not significant.
In the logistic regression model for hypertension, neither percentile blood lead nor percentile tibia
lead, entered together, were significant predictors.
Reporting was incomplete: number of subjects entering the models was not stated; no comparisons
between recruited subjects and subjects not used in models. Despite reporting non-significant
interactions, the paper showed both loess plots and tables of analyses stratified by genotype, reporting
significant associations between both tibia and blood lead in the GG genotype, insignificant in the
other. Inspection of the loess plots revealed striking non-linearity for both adjusted blood lead-systolic
blood pressure and adjusted tibia lead-systolic blood pressure relationships. Small group size of the
TG/TT genotypes and highly unbalanced terms of the interaction may have contributed to the non-
significant interactions. Although the interaction lead term was also probed as a quadratic function,
the tibia lead interaction was not, suggesting that poor concentration-response specification in the
model may also have contributed to the lack of significant main effects and interactions.
-------
Table AX6-5.1 (cont'd). Cardiovascular Effects of Lead
to
o
o
Reference, Study
Location, and
Period
Study Description
Lead
Measurement
Findings, Interpretation
Blood Pressure, Hypertension
Asia (cont'd)
Nomiyama et al. 123 female lead-exposed leaded crystal toy workers, Blood lead mean
(2002) mean age (range) 27.3 (17-44) years, and 70 female (SD, range) in
China, Beijing sewing workers (reference group), mean age (range) lead workers:
No statement on 24.2 (16-58) years were tested. Forward stepwise 55.4
dates of data multiple regression models of systolic and diastolic (13.5, 22.5-99.4
collection blood pressure of the combined groups were used with Hg/dL
linear blood lead and a set of covariates. Variables with
p < 0.2 were allowed to enter. The covariate set was Blood lead mean
selected from a larger set of potential covariates by (SD, range) in
factor analysis, and a representative variable from each non-lead
factor was selected for possible entry in the regressions. workers:
6.4(1.6,3.8-
Alternate models were constructed using four ordered 11.4) ug/dL
categories of blood lead, instead of the linear continuous
blood lead variable. Logistic regressions were used to
determine the odds of elevated systolic (>125 mmHg)
and elevated diastolic (>80 mmHg) blood pressure as a
function of blood lead category.
Adjusted for age, urine protein, and plasma triglyceride, each 1 ug/dL increase in linear blood lead
significantly associated with a 0.13 mmHg increase in systolic blood pressure (no SE or CI given;
p = 0.0003). Adjusted for plasma triglyceride, age, urine protein, plasma low density lipoprotein, and
hypertension heredity, each 1 ug/dL increase in linear blood lead was associated with a 0.10 mmHg
increase in diastolic blood pressure (no SE or CI given; p = 0.0001).
Using the ordered categories of blood lead and the same covariates for systolic and diastolic blood
pressure, the 40-60 ug/dL group had 4.2 mmHg (95% CI: 0.0, 8.5) higher systolic blood pressure and
4.1 mmHg (95% CI: 1.3, 6.8) higher diastolic blood pressure than the reference group (blood lead
(<11.4ug/dL). The group with >60 ug/dL blood lead had 7.5 mmHg (95% CI: 3.0, 12.0) systolic
blood pressure and 6.3 mmHg (95% CI: 3.4, 9.1) diastolic blood pressure higher than the reference
group.
Logistic regression models for "elevated" blood pressure, modeled using the same covariates were
similar. In the 40-60 ug/dL group odds of systolic blood pressure >125 mmHg and diastolic blood
pressure >80 mmHg were 4.26 (95% CI: 1.07, 17.04) and 2.43 (95% CI: 0.97, 6.04), respectively,
higher than the reference group. The odds of "elevated" systolic and diastolic blood pressure in the
group with blood lead >60 ug/dL were 7.48 (95% CI: 1.86, 30.12) and 3.31 (95% CI: 1.29, 8.50),
respectively.
Incomplete reporting in paper: no model N, no SEs for linear blood lead regressions, no description of
type of factor analysis used or dates of data collection. Innovative use of factor analysis to select
covariates that, depending on how the factor analysis was run, could have produced a set of orthogonal
variables for model entry. However, BMI was not included in the original set of covariates or in the
models. Small sample size limits conclusions based on nonsignificant results. Stepwise regression
produced a different covariate pattern for each component of blood pressure. The linear blood lead
variable may be inappropriate given the marked skewness of blood lead in descriptive analysis. The
11 |ig/dL gap in blood lead between lead workers and non-lead workers could have introduced
problems in analyses with continuous blood lead. Larger age spread in non-exposed group than in
exposed group could have caused misspecification of age variable. No control for antihypertensive
medication use. No model diagnostics reported.
-------
Table AX6-5.1 (cont'd). Cardiovascular Effects of Lead
to
o
o
Reference, Study
Location, and
Period
Study Description
Lead
Measurement
Findings, Interpretation
Blood Pressure, Hypertension
Asia (cont'd)
Wu et al. (1996) 222 workers in two lead battery plants, 112 men, mean
Central Taiwan (range) age 36.2 (18-67) years, and 110 women, mean
No statement on (range) age 36.2 (18-71) years were tested for blood
dates of data lead relationships with systolic and diastolic blood
collection pressure in multiple regression models, using a fixed,
forced set of covariates: age, sex, BMI, working
history, years of work, noise exposure, natural log
ambient air lead concentration, and ordered categorical
blood lead concentration.
Arithmetic mean (SD,
range) blood lead:
Women: 44.6(18.4)
[8.3-103.1] ug/dL
Men: 60.2(26.8)
[17.0-150.4] ug/dL
Using four ordered blood lead categories (<25 ug/dL [n = 16/222; 6.8%],
25-40 ug/dL [58/222; 26.1%], 41-60 ug/dL [63/222; 27.9%], and >60 ug/dL [85/222;
38.3%]) adjusted systolic and diastolic blood pressure were not significantly related to
the top three blood lead categories compared to the lowest, natural log ambient lead.
Years in work environment was a significant predictor of both systolic and diastolic
blood pressure, but age was only marginally significant for systolic blood pressure and
not significant for diastolic blood pressure.
Small study size limits any conclusions drawn from non-significant results. Three
measures, all related to lead exposure, were simultaneously tested in the models. While
blood lead may only be weakly correlated with years of work, ambient air lead would
be expected to be much better correlated with blood lead. There is a clear possibility of
collinearity among those three variables, which would inflate standard errors and reduce
coefficients. Authors selected ordered categories of lead to "avoid unnecessary
assumption of linearity." The use of natural log air lead concentration suggests that
some diagnostics were run, but no model diagnostic tests were reported. No control for
antihypertensive medication use.
-------
Table AX6-5.2. Cardiovascular Morbidity Effects of Lead
to
o
o
Reference, Study
Location, and
Period
Study Description
Lead Measurement
Findings, Interpretation
Cardiovascular Morbidity
United States
Cheng etal. (1998)
U.S.-Boston,
Normative Aging
Study (VA)
1991-1995
775 males (97% white), mean age (SD)
[range] 67.8 years (7.3) [48-93].
Multiple linear regression models of heart
rate-corrected QT and QRS
electrocardiogram intervals were adjusted
by stepwise entry of covariates, retaining
only those that remained significant at
p < 0.10. Linear blood lead, tibia, and
patella bone lead were apparently (not
described in text) entered separately.
Logistic regression models for Minnesota
ECG Coding Center diagnoses of
intraventricular conduction deficit (IVCD),
atrioventricular conduction deficit (AVCD),
and arrhythmia were adjusted by covariates
the same way. Only analyses stratified by
age (<65 years, n = 277; >65 years, n = 498)
were presented
Arithmetic mean (SD) blood
lead: 5.8 ug/dL (3.4)
Mean (SD) tibia lead:
22.2 ug/g (13.4)
Mean (SD) patella lead:
30.8 ug/g (19.2)
Multiple regression models of QT intervals, adjusted for age, alcohol intake, BMI, and
diastolic blood pressure, found that only tibia and patella lead were significantly related to
outcome in the under 65 group. Every 10 ug/g increase of tibia and patella lead was
associated with a 5.0 ms (95% CI: 0.8, 9.2) and 3.0 ms (95% CI: 0.2, 5.8) increase in QT
interval, respectively. Multiple regression models of QRS intervals, adjusted for age,
fasting glucose level, and diastolic blood pressure, also found that only tibia and patella
lead were significantly related to outcome in the under 65 group. Every 10 ug/g increase of
tibia and patella lead was associated with a 4.8 ms (95% CI: 1.8, 7.8) and 2.2 ms (95% CI:
0.1, 4.4) increase in QRS interval, respectively. There were no significant effects of lead in
the 65 and over group.
Logistic regression models of IVCD, adjusted for age and serum HDL level, found that
only tibia lead was significantly related to outcome in the under 65 group. Every 10 ug/g
increase of tibia lead was associated with increased odds of IVCD, OR 2.23 (95% CI: 1.28,
3.90). There were no significant lead effects in the 65 and over group for IVCD. Logistic
regression models of AVCD, adjusted for age and serum HDL level, found that both tibia
and patella lead were significantly related to outcome in the 65 and over group. Every 10
ug/g increase of tibia lead and patella lead was associated with increased odds of AVCD,
OR 1.22 (95% CI: 1.02, 1.47) and OR 1.14 (95% CI: 1.00, 1.29), respectively. Leadwas
not significantly related to AVCD in the under 65 group. There were no significant effects
of lead on arrhythmia in either age group.
Stepwise models may capitalize on chance associations. Linear blood lead specification
may not be appropriate in some or all of these models. Not clear if three models were
constructed for each stratified analysis for each outcome, each based on a different lead
index. No statistical comparisons across strata. No model diagnostics were presented.
Gump et al. (2005) See Gump et al. (2005) entry in Blood
U.S.-Oswego, NY Pressure/Hypertension section for heart rate,
Dates of study not stroke volume, cardiac output, total
given peripheral resistance, and cardiac interbeat
interval data.
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Table AX6-5.2 (cont'd). Cardiovascular Mordibity Effects of Lead
to
o
o
Reference, Study
Location, and
Period
Study Description
Lead Measurement
Findings, Interpretation
Cardiovascular Morbidity
United States (cont'd)
O
o
2
o
H
O
o
HH
H
W
Navas-Acien (2004)
U.S.-NHANES IV-
Phase 1
1999-2000
Schwartz (1991)
NHANES II
U.S.
1976-1980
2125 subjects (1070 males, 1055 females),
age 40->70 years were tested for peripheral
arterial disease (PAD; n = 139) by taking
the ratio of the ankle mean systolic blood
pressure to the arm mean systolic blood
pressure. Any subject with the ratio <0.90
was classified as PAD. Logistic regression
analysis was weighted and adjusted by
sample design. Covariates forced into the
models were age, sex, race, education, and
lead quartile (Model 1); Model 1 covariates
plus BMI, alcohol intake, hypertension,
diabetes, hypercholesterolemia, glomerular
filtration rate, and C-reactive protein
(Model 2); Model 2 covariates plus self-
reported smoking status and serum cotinine
(Model 3); and Model 3 covariates plus
cadmium quartile (Model 4). Tested
interactions between lead and cadmium on
PAD, and between lead and sex, race,
smoking status, renal function, and c-
reactive protein on PAD. Tested for trend
of OR as a function of lead quartile.
See Schwartz (1991) entry in Blood
Pressure/Hypertension for left ventricular
hypertrophy results.
Geometric mean blood lead
(25%-75% percentile):
2.1 ug/dL (1.4, 2.9)
Lead quartile 1: <1.4ug/dL
Lead quartile 2: 1.4-2.1
ug/dL
Lead quartile 3: 2.1-2.9
ug/dL
Lead quartile 4: >2.9 ug/dL
Odds for PAD significantly increased with lead quartile (1st quartile used as comparison)
for all four models. Only models 1 and 2, however, showed a significant increase in odds
of PAD for the 4th lead quartile compared to the 1st lead quartile, OR 3.78 (95% CI: 1.08,
13.19) and OR 4.07 (95% CI: 1.21, 13.73). None of the tested interactions with blood lead
quartile were significant.
Well-designed study with sound statistical analysis. Including two variables for smoking in
Models 3 and 4 (smoking status and cotinine) may have over-controlled for smoking).
There was a trend toward increased blood lead level with increased smoking status and with
increased cotinine levels, though no statistical tests of trend were reported. Thus the two
smoking variables and lead may have been confounded with PAD. No model diagnostic
tests reported.
Tepperetal. (2001)
U.S.-Cincinnati, OH
After 1991 to before
2001
See Tepper et al. (2001) entry in Blood
Pressure/Hypertension for left ventricular
mass results
-------
Table AX6-5.2 (cont'd). Cardiovascular Mordibity Effects of Lead
to
o
o
Reference, Study
Location, and
Period
Study Description
Lead Measurement
Findings, Interpretation
Cardiovascular Morbidity
Europe
Gustavsson et al. Study base was all Swedish citizens 45-70
(2001) years old from Stockholm County free of
Europe-Stockholm, previous myocardial infarction. Cases who
Sweden survived at least 28 days after infarct (1,105
1992-1994 males and 538 females) of which 937 men
(85%) and 398 women (74%) had sufficient
information on occupational exposures and
"main confounders", were compared
against referents (1,120 men and 538
women) matched to cases by sex, age, year,
and hospital catchment area. Risk ratios for
the case group compared to referent group
were adjusted on the basis of the matching
variables and smoking, alcohol drinking,
hypertension, overweight, diabetes mellitus,
leisure physical "inactivity", and were
calculated for a number of exposure factors
separately, including lead.
Lead exposure was classified
as none, low or high
corresponding to airborne
dust levels of 0, >0 to 0.03,
and >0.04 mg/m3,
respectively, for the highest
intensity of exposure during
at least one year of work.
The same three classifications
were used for 0, >0 to 0.04,
and >0.05 mg/m3 for
cumulative exposure.
All risk ratios were calculated relative to the "no exposure" groups. Adjusted risk ratios for
surviving a myocardial infarction were 0.88 (95% CI: 0.69, 1.12) and 1.03 (95% CI: 0.64,
1.65) for low and high exposure groups for peak lead exposure, and were 0.81 (95% CI:
0.60, 1.11) and 1.00 (0.74, 1.34) for the low and high cumulative exposure groups.
This study of myocardial morbidity was compromised by poor lead exposure
characterization (occupational air dust lead concentration) and by including a covariate
collinear with lead exposure and confounded with the outcome, hypertension, in the
adjusted models.
No model diagnostics were reported.
-------
Table AX6-5.3. Cardiovascular Mortality Effects of Lead
to
o
o
Reference, Study
Location, and
Period
Study Description
Lead Measurement
Findings, Interpretation
Cardiovascular Mortality
United States
Lustberg and 4190 persons, 30 to 74 years, 929 of whom died
Silbergeld (2002) during follow up, had baseline blood lead
U.S.-NHANES II measurements during the NHANES II period.
1976-1980, follow Proportional hazard models for circulatory
up to 1992 disease-related death (ICD-9 codes 390-459)
were based on the complex survey design, but
not weighted. Presented models were
unadjusted, adjusted for age and sex, and
adjusted for age, sex, location, education, race,
income, smoking, BMI, and exercise. Blood lead
was entered as an ordinal three-category variable.
Blood lead <10 ug/dL,
n=818
Blood lead 10-19 ug/dL,
n = 2735
Blood lead 20-29 ug/dL,
n = 637
Blood lead >30 ug/dL,
n = 102, excluded from
analysis
Crude, sex and age adjusted, and multivariate adjusted circulatory disease mortality were
all significantly increased in the 20-29 ug/dL group compared to the <10 ug/dL reference
group. Risk ratio for the highest lead group for crude circulatory mortality was 1.74 (95%
CI: 1.25, 2.40), for age and sex adjusted circulatory mortality was 1.48 (95% CI: 1.10,
2.01), and for multivariate circulatory mortality was 1.39 (95% CI: 1.01, 1.91).
Stratified analyses were performed by race, sex, age, smoking, education, etc., but only for
all-cause mortality. No model diagnostics reported.
Michaels et al. 1261 males, average age (range) at the beginning
(1991) of study 49.6 years (19-83), representing 24,473
U.S.-New York person-years were followed. 498 died in the
City interval. Subjects belonged to the International
1961-1984 Typographical Union and worked at two large
city newspapers. Hot lead linotyping was
discontinued at the newspapers during 1974-
1978, providing the primary source of
occupational exposure. Last exposure for all
subjects still employed was at the end of 1976.
Standardized mortality ratios (SMR) were
calculated using the LTAS program developed
by NIOSH, calculating the expected number of
deaths of the cohort referenced to a comparison
population, in this case disease-specific mortality
rates from New York City. Cohort was stratified
based on years of employment. Causes of death
were based on ICD-8 codes.
Exposure was estimated
based on years of linotype
employment before the
end of 1976. Authors note
that, based on
measurements at other
print shops using hot lead
linotype, air lead levels
probably did not exceed 20
ug/m3.
Standardized mortality ratio was significant (SMR = 1.68 [95% CI: 1.18, 2.31]) only for
cerebrovascular disease in those working, and thus exposed, for 30 years or more. Neither
arteriosclerotic heart disease (ICD-8 410-414) nor vascular lesions of the central nervous
system (ICD-8 430-438) had significant SMR in the total cohort not stratified by years of
exposure.
No direct measurement of lead exposure. Many groupings of ICD codes were explored in
stratified and unstratified analyses, with the only significantly elevated SMR found for
cerebrovascular disease. No a priori hypotheses. General weakness of all studies relying
on a comparison population is that the cohort belongs to the comparison population and can
influence the comparison mortality rates in direct proportion to the ratio between cohort
and comparison population size.
-------
Table AX6-5.3 (cont'd). Cardiovascular Mortality Effects of Lead
to
o
o
Reference, Study
Location, and
Period
Study Description
Lead Measurement
Findings, Interpretation
Cardiovascular Mortality
United States (cont'd)
Steenland et al. The death certificates of 1028 males of the
(1992) 1990 who worked at a smelter plant at least
U.S.-Idaho one year between 1940 and 1965 were
>1941 to 1988 examined to construct standardized mortality
ratios (SMR) for various ICD-9 disease
classifications using the U.S. population as a
referent group.
In 1976 blood lead of 173
workers averaged (SD)
56.3 ug/dL (12.9). Air
lead was measured in 1975
at3.1mg/m3in208
personal 8-hour samples.
High lead departments in
the plant were defined as
those exceeding 0.2 mg/m3
in the 1975 survey.
Non-malignant respiratory disease and accidents accounted for most of the significantly
elevated SMR in the group. Neither ischemic heart disease (410-414)(SMR = 0.94; 95%
CI: 0.84, 1.05), hypertension with heart disease (402, 404)(SMR = 0.97; 95% CI: 0.53,
1.63), hypertension with no heart disease (401, 403, 405)(SMR = 1.73; 95% CI: 0.63, 3.77)
or cerebrovascular diseases (430-436)(SMR = 1.05; 95% CI: 0.82, 1.32) were significantly
different from deaths in the U.S. population. Similar results were found for the people
working in the "high lead departments".
Though there is no doubt that this group was highly exposed to lead, exposure
characterization over the working lifetime was not well defined, few blood lead data were
available, and poor demographic data for the exposed group only allowed a comparison
with total U.S. population. As is usual with occupationally exposed groups, selection bias
may influence results. No smoking data were available for the group. In industrial
conditions smoking will be confounded with other lead exposure (constant hand to mouth
behavior on the plant floor will exposure smokers to more lead via the oral route than in
non-smokers).
-------
Table AX6-5.3 (cont'd). Cardiovascular Mortality Effects of Lead
to
o
o
Reference, Study
Location, and
Period
Study Description
Lead Measurement
Findings, Interpretation
Cardiovascular Mortality
Europe
Gerhardsson et al. 664 male workers at a secondary lead smelter
(1995) had blood lead tested every 2-3 months since
Europe-southern 1969. The past blood lead level of 201
Sweden workers who had been working at the plant
1969-1989 from before 1969 was estimated from their
1969 results. Median (10th percentile, 90th
percentile) year of birth was 1943 (1918,
1960). Median (10th percentile, 90th
percentile) duration of employment was 2.8
years (0.3, 25.7) and median (10th percentile,
90th percentile) duration of follow up was 13.8
years (2.8, 20.9). A total of 8706 person-years
were represented in the study. Standardized
mortality ratios based on county mortality
tables by calendar year, cause, sex and five-
year age group were calculated.
Cardiovascular diseases were coded by ICD-8
from death certificates.
Arithmetic mean blood
lead levels dropped from
approximately 62 ug/dL in
1969 to approximately 33
Hg/dLinl985. 95%
confidence intervals were
difficult to extract from the
presented graph, but
appeared to be no more
than 5-6 ug/dL about the
All cardiovascular disease mortality (ICD-8 390-458) was significantly elevated above that
expected from the county mortality tables (SMR = 1.46 [95% CI: 1.05, 2.02]), with 39 of
the 85 deaths observed in the cohort. For just ischemic heart disease (ICD-8 410-414),
SMR = 1.72 (95% CI: 1.20, 2.42) in the plant workers with 34 of the 85 deaths observed in
the cohort. There were no deaths recorded for cerebrovascular diseases (ICD-9 430-438).
There was no apparent concentration-response relationship, using peak blood lead and
time-integrated blood lead.
Problems inherent in using standardized mortality ratios in such mortality studies have been
discussed above. The sample size was too small (85 all cause deaths among 664 workers)
to interpret non-significant results.
M011er and
Kristensen (1992)
Europe-Denmark-
Copenhagen County-
Glostrup Population
Studies
1976-1990
See M011er et al. (1992) entry in Blood
Pressure/Hypertension for results of
cardiovascular disease and coronary heart
disease mortality.
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ANNEX TABLES AX6-6
May 2006 AX6-153 DRAFT-DO NOT QUOTE OR CITE
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Table AX6-6.1. Placental Transfer of Lead from Mother to Fetus, Human Studies
to
o
o
Reference
Study Description
Lead Measurement
Findings, Interpretation
O
O
2
o
H
O
United States
Harville et al.
(2005)
Angell and Lavery
(1982)
Bogden et al
(1978)
Fahimetal. (1976)
Gershanik et al.
(1974)
Mean maternal blood lead
concentration = 1.9 ug/dL
159 mother-infant pairs
Pittsburgh, PA from 1992-1995
Louisville, KY(n = 63 5)
Placental transfer
Newark, NJ (n = 150)
Mining town in Columbia, MO
(n = 249)
March-Sept 1972
Shreveport, LA, 98 pairs
Maternal blood lead and cord blood lead
Correlation coefficient = 0.79
Maternal and cord blood lead concentration Correlation = 0.60
Maternal and cord blood lead concentration Correlation = 0.55
Maternal and cord blood lead concentrations Correlation = 0.29
Maternal and cord blood lead concentrations Correlation = 0.64
o
HH
H
W
Europe
Graziano et al.
(1990)
Hueletal. (1981)
Rods etal. (1978)
Lauwreys et al.
(1978)
Barltrop (1968)
902 births in two towns in Kosovo,
former Yugoslavia
Hair sample pairs (n = 110)
France
Placenta transfer, Belguim
(n=474)
Placenta transfer, Belguim
(n = 500)
Stillbirths and spontaneously
aborted fetuses
Maternal blood lead concentrations at mid-
pregnancy and delivery, cord blood lead
concentrations. Geometric mean BPb in
exposed town = 17.1 ug/dL; inunexposed
town= 5.1 ug/dL
Maternal and fetal hair lead concentration
Placental lead concentration and cord blood
lead concentration
Correlations between maternal BPb
measures and cord BPb measures ranged
from 0.8 to 0.9
Correlation = 0.24
Correlation = 0.28
Maternal and cord blood lead concentrations Correlation = 0.81
Concentrations in bones, livers, blood, hearts, Lead began to cross the placenta at least at
kidneys, and brains week 12 of gestation and increased to term
-------
Table AX6-6.1 (cont'd). Placental Transfer of Lead from Mother to Fetus, Human Studies
to
o
o
Reference
Study Description
Lead Measurement
Findings, Interpretation
Mexico
Chuang et al.
(2001)
615 women in Mexico City recruited in 1994-1995 these
investigators used structural equation modeling to estimate
the associations between whole blood lead, bone lead
(cortical and trabecular), and the latent variable, plasma lead
and cord blood lead. They found the strongest associations
between whole blood lead and cord blood lead, even after
accounting for plasma lead. The greatest contributors to
plasma lead were bone lead and airborne lead.
Maternal whole blood lead, bone lead
(cortical and trabecular), plasma lead
(latent variable) and cord blood lead
Using structural equation
modeling, strongest
associations found between
maternal whole blood lead
and cord blood lead
Other Locations
Clark (1977)
Casey and
Robinson (1978)
Chaube et al.
(1972)
Placenta transfer
(n = 122) Zambia
(Broken Hill Mine)
Stillbirths and spontaneously aborted fetuses (n = 44)
New Zealand
First trimester spontaneously aborted fetuses (n = 50)
Location not specified
Maternal and cord blood lead
concentrations
Concentrations in liver, kidneys and
brains
Concentrations in liver, brain and
kidneys
Correlation = 0.77
Lead accumulation
increasing with length of
gestation
Placental transfer occurs
earlier than gestational
week 12
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Table AX6-6.2. Lead Exposure and Male Reproduction: Semen Quality, Human Studies
•<
to
o
o
ON
Reference Study Description Lead Measurement
United States
Cullen et al. (1984) Lead workers, U.S. Exposure > 60 ug/dL
Findings, Interpretation
Decreased sperm counts
Europe
Bonde et al. (2002)
European study of n = 503 Blood lead concentration
men employed in lead industry
Median sperm concentration reduced by 49% in men with
BPb > 50 jig/dL
Regression analyses indicated a threshold value of 44 ug/dL
below which no adverse associations were found.
2
©
0
0
H
O
0
H
W
O
O
H
W
Assennato et al.
(1986)
Lancranjan et al.
(1975)
Mexico
Hernandez-Ochoa
et al. (2005)
South America
Lerda (1992)
Lead workers, Italy
Lead workers (n = 150)
Europe
Northern Mexico
n = 30 lead factory workers
Argentina
Exposure > 60 ug/dL
Heavy exposure: mean
BPb = 74.5 ug/dL Moderate exposure:
Mean BPb = 52.8 ug/dL
Mean BPb = 9.3 ug/dL
Seminal fluid lead concentration
-D
Decreased sperm counts
Decreased sperm counts and increased prevalence of
morphologically abnormal sperm amongst workers with
heavy and moderate exposure to lead
Decreased sperm concentration, motility, normal
morphology and viability correlated with seminal fluid lead
and lead in spermatozoa.
No associations found with BPb.
Decreased sperm count, percent motility and increased
percent with abnormal morphology among exposed workers
-------
Table AX6-6.2 (cont'd). Lead Exposure and Male Reproduction: Semen Quality, Human Studies
to
o
o
Reference
Study Description
Lead Measurement
Findings, Interpretation
Other Locations
Benoff et al.
(2003a,b)
Alexander et al.
(1996a)
Chowdhury et al.
(1986)
Couples undergoing in vitro
fertilization or artificial
insemination
n = 119 workers with both
blood and semen samples
n = 10 men with occupational
lead exposure
Seminal fluid lead concentration
Blood lead concentration - current
Body lead burden estimated from
current BPb and historical monitoring
Exposed group average
BPb = 42.5 ug/dL
Unexposed group average
BPb = 14.8
Higher concentrations of seminal fluid lead in the male
partner of couples who did not conceive, compared to those
who did conceive
Geometric mean sperm concentrations decreased with
increasing current and long-term body burden of lead
Sperm Count * 10b
exposure. Current BPb
<15 ug/dL 79.1
15-24 ug/dL 56.5
25-39 ug/dL 62.7
>40 ug/dL 44.4
Similar results for body lead burden. Similar trends for
percent motility.
No associations for sperm morphology or reproductive
hormones.
Decrease in sperm count, percent motility and increase in
number of sperm with abnormal morphology
-------
Table AX6-6.3. Lead Exposure and Male Reproduction: Time to Pregnancy, Human Studies
to
o
o
Reference
Study Description
Lead Measurement
Findings, Interpretation
Europe
Apostoli et al.
(2000)
Sallmen et al.
(2000)
Italian men included in the Asclepios project.
n = 251 exposed men with at least one completed pregnancy.
n = 45 unexposed men with at least one completed
pregnancy.
n = 502 couples identified by the Finnish Institute of
Occupational Health. Male partner occupationally exposed
to lead.
Blood lead at time closest to
conception.
Blood lead concentration
Available close to time of
conception on 62% of men; in 38%
estimated based on BPbs obtained
at other times or based on job
histories.
Time to pregnancy shorter in
couples in which male partner
exposed.
Secondary analyses:
Among men with BPb > 40
ug/dL, time to pregnancy longest.
Limiting analysis only to
exposed men, time to pregnancy
longer among men with the
highest BPbs.
Time to pregnancy reduced
among couples in which male
partner had BPb > 10 ug/dL,
compared to those in which male
partner had BPb < 10 ug/dL.
Fecundity Density Ratios (95%
Confidence Intervals)
BPb
10-20 ug/dL 0.92 (0.73, 1.16)
21-30 ug/dL 0.89 (0.66, 1.20)
31-40 jig/dL 0.58(0.33,0.96)
>40 ug/dL 0.83 (0.50, 1.32)
-------
Table AX6-6.3 (cont'd). Lead Exposure and Male Reproduction: Time to Pregnancy, Human Studies
to
o
o
Reference
Study Description
Lead Measurement
Findings, Interpretation
Europe (cont'd)
Joffee et al. (2003)
Asclepoios Project, large European collaborative cross-
sectional study.
n = 1108 men; 638 occupationally exposed to lead at the time
of pregnancy. Remainder exposed but exposure did not
coincide with pregnancy.
Live births only.
Blood lead concentration
Fecundity Density Ratios
(95% Confidence Intervals)
BPb
<20 ug/dL
20-29 jig/dL
30-39 ug/dL
>40 ug/dL
1.12(0.84, 1.49)
0.96(0.77, 1.19)
0.88(0.70, 1.10)
0.93 (0.76, 1.15)
Similar results when duration of
exposure or cumulative exposure
used
Other Locations
Shiau, et al. (2004)
n = 280 pregnancies in 133 couples in which male partner
employed in battery plant.
n = 127 conceived during exposure; remainder conceived
prior to exposure.
Blood lead concentration
Fecundity Density Ratios
(95% Confidence Intervals)
BPb
30-39 ug/dL
>39 ug/dL
0.50 (0.34, 0.74)
0.38 (0.26, 0.56)
Using BPb as a continuous
variable and restricting the
analysis to BPb between 10 and
40 ug/dL, time to pregnancy
increased by 0.15 months for
each 1 ug/dL increase in BPb.
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Table AX6-6.4. Lead Exposure and Male Reproduction: Reproductive History, Human Studies
to
o
o
Reference
United States
Lin etal. (1996)
Study Description
New York State 1981-1992
Linked records from Heavy Metal Registry to birth certificates
from New York State Office of Vital Statistics
n = 4256 men
n = unexposed 5148 men frequency matched for age
and residence
Lead Measurement
BPb
Exposure defined as at least
one BPb >25 ug/dL
Findings, Interpretation
Exposed group fewer births than
expected, especially among those
employed in lead industry over
5 years.
O
O
2
o
H
O
o
HH
H
W
Europe
Gennart et al.
(1992)
(1997)
Sallmen et al.
(2000)
Among 365 men occupationally exposed to metals,
n = 74 exposed continuously for more than 1 year
Reference group with no occupational exposure
Belgium
Bonde and Kolstad Denmark
Matched roster of male employees age 20-49 years of three
battery plants to birth registry
n=1349
Control group of 9656 men not employed in lead industry
Finland
Males monitored for occupational exposure at Finnish Institute
of Occupational Health
n=2111
n = 681 controls with BPb < 10 ug/dL
Exposure at least one year
continuously and at least
one BPb >20 ug/dL
Employment in lead
industry
Duration of employment in
lead industry
Probably exposed
Possibly exposed
Based on measured BPb in
relation to time of marriage
Compared to reference group,
probability of at least one live birth
reduced in exposed group.
Fertility decreased with increasing
exposure (although number of
exposed men small).
No associations found between
exposure measure and birth rate.
Among men in the probably
exposure group, risk ratio for failing
to achieve pregnancy compared to
unexposed:
BPb
10-20 ug/dL 1.3
>50 ug/dL 1.9
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ANNEX TABLES AX6-7
May 2006 6-161 DRAFT-DO NOT QUOTE OR CITE
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Table AX6-7.1. Recent Studies of Lead Exposure and Genotoxicity
to
o
o
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings and Interpretation
O
o
*
o
H
O
o
HH
H
W
Europe
Fracasso et al. (2002)
Italy
Palus et al. (2003)
Poland
Case-control design.
37 workers employed at a battery plant.
29 student and office worker volunteers with
no known occupational exposure to
genotoxins.
Peripheral lymphocytes isolated from whole
blood.
Reactive Oxygen Species (ROS) production,
cellular GSH level, PKC isoforms, and DNA
breaks (via comet) assayed.
ANOVA and logistic regression used to
compare workers vs. healthy volunteers.
Adjusted for age, alcohol use, and smoking.
Cross-sectional design.
Battery plant workers: 34 acid battery,
22 alkaline battery, and 52 plant personnel
from departments with no known exposure to
Pb or Cd.
Lymphocytes isolated from whole blood.
SCE, MN, DNA damage (via comet) assayed.
Means compared via ANOVA.
Battery plant workers. Blood lead
categories used for some comparisons,
with <25, 25-35, and >35 ug/lOOmL as
cutpoints.
Mean blood Pb 39.6 ug/lOOmL for
workers, 4.4 ug/100mL for volunteers.
Workers considered Pb-exposed if
from acid battery department,
Cd-exposed if from alkaline, unexposed if
from other department.
Mean blood Pb 504 ug/L for
Pb-exposed workers, 57 ug/L for
Cd-exposed, and 56 ug/L for other
workers.
OR (95% CI)
Workers vs. Volunteers:
ROS: 1.43 (0.79-2.60)
DNA Breaks (Tail Moment): 1.07 (1.02-1.12)
GSH: 0.64(0.49-0.82)
PKC a reduced in workers, atypical PKC unchanged vs. volunteers
(no statistics provided).
Means (SE) via blood lead category for ROS and GSH:
<25 ug/ug/100 mL 4.9 (0.4) and 12.8 (0.8)
25-35 ug/100 mL 5.4 (0.7) and 7.7 (1.7)
>35 ug/100 mL
5.4 (0.5) and 9.2 (1.2)
Major analyses controlled for age, smoking, and alcohol intake.
Analyses by blood lead category not controlled for age, smoking, or
alcohol intake but these factors said not to influence endpoint and/or
results "significantly." No control for potential coexposures.
Mean (SD)
Pb exposed workers (all combined):
SCEs 7.48(0.88)
MN 18.63 (5.01)
NDI 1.89 (no SD given)
Cd exposed workers (all combined):
SCEs 6.95 (0.79)
MN 15.86(4.92)
NDI 1.96 (no SD given)
Other workers (all combined):
SCEs 6.28(1.04)
MN 6.55 (3.88)
NDI 1.86 (no SD given)
Elevation of SCEs and MN vs. controls at p < 0.05 and p < 0.01,
respectively.
Both SCEs and MN elevated among Pb exposed workers as well as
Cd-exposed workers compared to controls. Differences greatest for Pb-
exposed workers.
Higher SCE and MN also occurred among Pb-exposed workers after
stratification by smoking status.
No direct control for potential coexposures, but mean blood Cd no higher
in Pb-exposed than in other worker group.
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Table AX6-7.1 (cont'd). Recent Studies of Lead Exposure and Genotoxicity
to
o
o
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings and Interpretation
Europe (cont'd)
Van Larebeke et al.
(2004)
Belgium
Cross-sectional design.
99 female nonsmokers, ages 50-65,
drawn from rural and industrial areas.
Peripheral lymphocytes isolated from
blood. HPRT variant frequency
determined.
Lead concentration measured in
blood (serum).
Women also classified as above vs.
below median for blood Pb
HPRT variant frequency
Above median serum Pb: 9.45 x 10~6
Below median serum Pb: 5.21 x 10'6
P-value for difference = 0.08 adjusted for age, education,
smoking, BMI, and serum Se. (Significant inverse association
noted between variant frequency and serum Se.)
Uncontrolled for potential exposure to other genotoxins.
O
o
*
o
H
O
o
HH
H
W
Latin America
Minozzo et al. (2004)
Brazil
Cross-sectional design.
26 workers employed at a battery
recyclery for 0.5 to 30 years.
29 healthy volunteers of similar age
range and SES.
Peripheral lymphocytes isolated from
whole blood.
Fixed blood slides stained with Giemsa
visually evaluated to determine
micronuclear frequency (MN) and
cellualr proliferation as nuclear
division index (NDI).
ANOVA and logistic regression used
to compare workers vs. healthy
volunteers. Adjusted for age, alcohol
use, and smoking.
Battery recyclery workers were
considered exposed.
Blood lead also determined.
Mean blood Pb 35.4 ng/dL for
workers, 2.0 |ig/dL for volunteers.
Mean (S.D.)
Means (SD) for workers and volunteers
MN
3.85 (2.36) and 1.45 (1.43)
NDI
1.77 (0.22) and 1.89 (0.18)
Kendal correlation coefficient
All workers {assuming recyclery workers only, not total
population, but no population number given in Table.}
Blood Pb x MN: 0.061 (p = 0.33)
Blood Pbx NDI: 0.385 (p = 0.003)
Not controlled for age or SES, although worker and volunteer
populations said to be of similar age and SES. Uncontrolled for
potential coexposures. Correlations appear uncontrolled for
smoking, age, or other factors. Differences in MN and NDI
minor for smokers vs. nonsmoker, however. Diet "type"
"similar" for workers and controls, although no definition of
similarity provided.
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Table AX6-7.2. Key Occupational Studies of Lead Exposure and Cancer
to
o
o
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings and Interpretation
O
o
2
o
H
O
o
HH
H
W
United States
Steenlandetal. (1992)
(follow-up of Selevan
etal. (1985)
U.S.
1940-1988
Wong and Harris
(2000)
(follow-up of Cooper
etal. (1985)
U.S.
1947-1995
Cohort design.
1,990 male workers employed for
at least 1 year in a lead-exposed
department at a U.S. lead smelter
in Idaho during 1940-1965.
Mortality traced through 1988 to
determine cause of death.
SMR computed for workers vs.
national rates for age-comparable
counterparts.
Cohort design.
Lead battery plant (4,518) and smelter
(2,300) workers.
Worker mortality was followed up
through 1995.
Cause of death was identified from
death certificates.
Mortality was compared with U.S.
national age-, calendar-year-, and
gender-specific rates to compute the
SMR.
(See additional entry for nested study
of stomach cancer.)
Exposure categorizations based on
airborne lead measurements from
1975 survey. High-lead-exposure
subgroup consisted of 1,436 workers
from departments with an average of
least 0.2 mg/m3 airborne lead or
>50% of jobs showing 0.40 mg/m3
or greater. Mean blood lead 56 ug/dL
in 1976.
Workers were evaluated as a whole,
and also as separate battery plant and
smelter worker populations.
Job histories were also used to
stratify workers by cumulative years
of employment (1-9,10-19, 20+),
date of hire (pre-1946 vs. 1946 on),
and lag between exposure and cancer
(<20,20-34, >34 years). Mean blood
lead 80 ug/dL during 1947-72 among
smelter workers, 63 ug/dL among
battery workers.
SMR (95% CI); no. of deaths
Total cohort:
Nonsignificantly elevated RRs: kidney, bladder, stomach, and lung
cancer.
High-lead-exposure subgroup:
Kidney 2.39 (1.03,4.71); 8
Bladder 1.33 (0.48, 2.90); 6
Stomach 1.28 (0.61,2.34); 10
Lung 1.11 (0.82, 1.47); 49.
No control for smoking or exposure to other metals.
SMR (95% CI)
Battery plant workers:
All cancer 1.05(0.97,1.13)
All respiratory 1.13 (0.98, 1.29)
Stomach 1.53 (1.12, 2.05), significant
Lung, trachea, bronchus
1.14 ( 0.99, 1.30), marginal significance
Thyroid, Hodgkin's: nonsignificant
Bladder 0.49 (0.23, 0.90), significant depression
Smelter -workers:
Digestive, respiratory, thyroid: nonsignificant
Lung
1.22 (1.00, 1.47), nonsignificant
Battery plant and smelter -workers combined:
All cancer 1.04(0.97,1.11)
All respiratory 1.15 (1.03, 1.28), significant
Stomach 1.47 (1.13, 1.90), significant
Lung, trachea, bronchus
1.16(1.04, 1.30), significant
Thyroid/endocrine 3.08 (1.33, 6.07), significant
Lung and stomach risks lower for pre-1946 hires; higher for workers
employed 10-19 years than <10, but lower for >19 years; SMRs peaked
with 20- to 34-year latency for lung, but <20 years for stomach.
No control for smoking or exposure to other agents. No assessment of
employment history after 1981.
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Table AX6-7.2 (cont'd). Key Occupational Studies of Lead Exposure and Cancer
to
o
o
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings and Interpretation
O
o
2
o
H
O
United States (cont'd)
Wong and Harris
(2000)
U.S.
1947-1995.
(Nested in Wong and
Harris 200 cohort.)
Case-control design.
Cases: the 30 stomach cancer cases
occurring in a Philadelphia lead
battery plant.
Controls'. 120 age-matched cohort
members.
Mean exposure was compared for
cases vs. controls. Odds of exposure
were also computed for increasing
quartiles of cumulative exposure.
Job titles were used to classify lead
exposure as low, intermediate, or
high; total months of any exposure,
of intermediate or high exposure
only, and of cumulative exposure,
with months weighted by 1, 2, or 3 if
spent in low-, intermediate-, or high-
exposure job.
Mean months of employment, of intermediate or high exposure, or
of weighted exposure to lead were all nonsignificantly lower
among cases.
OR for cumulative weighted exposure in the 10 years prior to
death:
First quartile
1.00
Second quartile
0.62
Third quartile
0.82
Fourth quartile
0.61
o
HH
H
W
P for trend = 0.47; ORs showed no positive association with
any index of exposure.
Analyses appear uncontrolled for smoking, other occupational
exposures, or other risk factors.
Europe
Fanning (1988)
(Cases overlap those
occurring in
Dingwall-Fordyce and
Lane, 1963; and
Malcolm and Barnett,
1982).
U.K.
1926-1985
Proportional mortality/cohort design.
Subjects: 2,073 deceased males
identified through pension records of
lead battery and other factory workers
in the U.K.
Workers dying from a specific cancer
were compared with workers dying
from all other causes
Workers were classified as High or
moderate lead exposure vs. little or
no exposure based on job titles.
OR (95% CI) [Number of deaths]
Lung cancer:
0.93(0.8, 1.1) [76 deaths]
Stomach cancer 1.34 [31 deaths]
No associations for other cancer types; elevations in stomach and
total digestive cancers limited to the period before 1966.
-------
Table AX6-7.2 (cont'd). Key Occupational Studies of Lead Exposure and Cancer
to
o
o
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings and Interpretation
Europe (cont'd)
Anttilaetal. (1995)
Finland
1973-1988
Cohort plus case-referent design.
20,700 workers with at least one blood
lead measurement between 1973 and
1983.
Workers were linked to the Finnish
Cancer Registry for follow-up through
1988. Fro deceased workers, cause of
death was identified from death
certificate.
Mortality and incidence were compared
with gender-, 5-year age, and 4-year
calendar-year matched national rates.
Blood lead concentration.
Exposure was categorized according
to the highest peak blood level
measured:
Low: 0-0.9 nmol/L
[Otol8.6ng/dL]
Moderate: 1-1.9 |imol/L
[20.7 to 39.4 ng/dL]
High: 2-7.8 (imol/L
[41.4 to 161.6 ng/dL]
Mean blood lead 26 |ig/dL.
Total cohort:
No elevation in total or site-specific cancer mortality
Moderately exposed:
Total respiratory and lung cancer:
SIR =1.4 (95% CI: 1.0, 1.9) for both
Total digestive, stomach, bladder, and nervous system:
nonsignificant elevations
Highly exposed:
No increase in risks
All cancer:
RR=1.4(95%CI: 1.1,1.8)
Lung or tracheal:
RR = 2.0(95%CI: 1.2,3.2)
No increase in high-exposure group
No RRs reported for other cancers
Case-referent substudies:
Lung cancer ORs increased with increasing cumulative
exposure to lead
Highly exposed: squamous-cell lung cancer OR = 4.1
(95% CI: 1.1, 15) after adjustment for smoking.
Short follow-up period limits statistical power, offset to a
large degree by the substantial sample size. No control for
exposure to other potential carcinogens.
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Table AX6-7.2 (cont'd). Key Occupational Studies of Lead Exposure and Cancer
to
o
o
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings and Interpretation
O
o
2
o
H
O
o
HH
H
W
Europe (cont'd)
Anttilaetal. (1996)
Finland
1973-1988
(Nested analysis based
onAntillaetal. 1995
cohort)
Gerhardsson et al.
(1995)
Sweden
1969-1989
Case-control design.
(See Anttila et al. 1995 for basic
information on the source population.)
Cases'. 26 Finnish men with CNS
cancer.
Controls: 200 Finnish men without
CNS cancer.
Nested case-control analysis.
Cohort design.
684 male Swedish secondary lead
smelter workers with lead exposure.
Cancer incidence among workers was
traced through 1989.
Incidence was compared with county
rates.
Peak blood lead levels used to
categorize exposure as 0.1-0.7,
0.8-1.3, and 1.4-4.3 ng/L.
Cumulative exposure estimated by
using mean annual blood lead level to
categorize exposure as 0, 1-6, 7-14,
or 15-49 ng/L.
Interviews were used to obtain
occupational history and other risk-
factor data from patients or next
of kin.
Blood lead level: any worker with a
detectable blood lead level was
classified as exposed.
OR (no. of cases or deaths)
CNS cancer incidence (26 cases):
Rose with increasing peak lifetime blood lead measurements; not
significant
Glioma mortality (16 deaths):
Rose consistently and significantly with peak and mean blood lead
level, duration of exposure, and cumulative exposure.
Mortality by cumulative exposure, controlled for cadmium, gasoline,
and year monitoring began:
Low (13 subjects) 2.0(2)
Medium (14 subjects) 6.2 (2)
High (16 subjects) 12.0(5)
1 death among 26 subjects with no exposure: test for trend significant
at;? = 0.02.
Controlled for smoking as well as exposure to cadmium and gasoline.
Complete follow-up with minimal disease misclassification.
SIR (95% CI); no. of cases
All malignancies:
1.27(0.91, 1.74); 40
Respiratory:
1.32 (0.49, 2.88); 6
All gastrointestinal:
cohort
1.84(0.92,3.29; 11
highest quartile 2.34 (1.07, 4.45); 9
Stomach:
1.88(0.39, 5. 50); 3
Colon:
1.46 (0.30, 4.28); 3
SIRs for all other sites except brain were nonsignificantly elevated;
too few cases.
No control for smoking. Small numbers, so meaningful dose-response
analyses not possible for most cancer sites.
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Table AX6-7.2 (cont'd). Key Occupational Studies of Lead Exposure and Cancer
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o
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings and Interpretation
O
o
2
o
H
O
o
HH
H
W
Europe (cont'd)
Lundstrom et al.
(1997)
(follow-up of
Gerhardsson et al.
(1986)
(see also subcohort
analyses of Englyst
etal.,2001).
Sweden
1928-1987
Englyst etal. (2001)
(follow-up and sub-
analysis of Lundstrom
etal., 1997).
Sweden
1928-1987
Cohort design.
3,979 copper and lead smelter workers.
Standardized mortality and incidence
ratios were computed for workers
compared with age-, year-, gender-, and
county-specific rates for the general
population.
Nested cohort analysis.
Limited to 1,093 workers in the
smelter's lead department, followed
through 1997.
Incidence was compared with county
rates; age-specific SIRs with 15-year
lag.
Fro some analyses, the entire cohort
was treated as exposed. For others,
job histories were used to single out
1,992 workers belonging to
departments thought to be exposed to
"lead only." Mean blood lead
monitoring test results across time
were used to single out a "highly
exposed" group of 1,026 workers
with blood lead levels >10 jimol/L
[>207|ig/dL].
Mean blood lead 60 ng/dL in 1959.
Workers were divided into
Subcohorts I and II for ever and
never worked in areas generally
associated with exposure to arsenic
or other known carcinogens (701 and
383 workers, respectively).
Detailed individual assessment of
arsenic exposure was made for all
lung-cancer cases.
SMR (95% CI); no. of deaths
Lung:
Total cohort 2.8 (2.0, 3.8); 39
Highly exposed 2.8 (1.8, 4.5); 19
SIR (95% CI); no. of cases
Lung with 15-year lag:
Total cohort 2.9 (2.1,4.0); 42
Highly exposed 3.4 (2.2, 5.2); 23
Lead-only 3.1 (1.7, 5.2); 14
Lead-only highly exposed 5.1 (2.0, 10.5); 7
Other highly exposed (total cohort),
•with 15-year lag:
Brain 1.6 (0.4,4.2); 4
Renal pelvis, ureter, bladder 1.8 (0.8, 3.4); 9
Kidney 0.9 (0.2, 2.5); 3
All cancer 1.1 (0.9, 1.4); 83.
No control for smoking.
SIR (95% CI); no. of cases
Subcohort I (coexposed):
Lung
2.4(1.2,4.5); 10
Subcohort II (not coexposed):
Lung
3.6(1.2, 8.3); 5
Subjects with lung cancer found to have history of "considerable"
exposure to arsenic: 9/10 among Subcohort I, 4/5 among
Subcohort II.
No control for smoking.
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Table AX6-7.2 (cont'd). Key Occupational Studies of Lead Exposure and Cancer
to
o
o
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings and Interpretation
Europe (cont'd)
Carta et al. (2003)
Sardinia
1972-2001
Cohort design.
918 lead smelter workers.
Mortality traced from 1972 through
2001.
Standardized mortality ratios
computed.
Smelter workers considered exposed.
Job histories also used to categorize
degree of exposure based on
environmental and blood lead
measurements for specific
departments and tasks during
1985-2001.
SMR; number of cases
Smelter workers as a whole
All cancer 1.01 ; 108
Gastric cancer 1.22 ; 4
Lymphoma/leukemia 1.82 ; 6
Lung cancer 1.21 ; 18
Highly exposed workers
Lung cancer 1.96 (95% C.I. 1.02, 3.68) for highest exposure
group, with statistically significant upward trend.
Analyses for worker population as a whole supported by
presence of dose-response pattern for lung cancer based on
estimated exposure. Modest population size, inability to assess
dose-response for cancers of interest other than lung. No control
for smoking or other occupational exposures.
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Table AX6-7.3. Key Studies of Lead Exposure and Cancer in the General Population
to
o
o
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings and Interpretation
O
o
2
o
H
O
o
HH
H
W
United States
Jemal et al. (2002)
(same cohort as in
Lustberg and Silbergeld,
2002 except for inclusion
criteria)
U.S.
1976-1992.
Lustberg and Silbergeld
(2002) (same cohort as
Jemal et al., 2002 except
for inclusion criteria)
U.S.
1976-1992.
Cohort design.
3,592 white participants from the 1976-
1980 NHANES II survey who had blood
lead measured at entry.
Mortality was followed through 1992 via
NDI and SSADMF.
RRs were calculated for the various
exposure groups compared to survey
participants with the lowest exposure,
adjusted for age and smoking.
Cohort design.
4,190 U.S. participants from the 1976-1980
NHANES II health and nutrition survey
who had blood lead measured at entry and
whose levels fell below 30 ug/dL.
Mortality was followed through 1992 via
NDI and SSADMF.
RRs were calculated for the various
exposure groups compared to survey
participants with the lowest exposure,
adjusted for age, smoking and other factors.
Blood lead (ug/dL) was measured by
atomic absorption and used to classify
subjects into exposure quartiles or
groups above vs. below median
exposure.
Median blood lead 12 ug/dL.
Blood lead (ug/dL) measured by
atomic absorption was used to classify
subjects into exposure groups:
Low: <10
Medium: 10-19
High: 20-19
Mean blood lead 14 ug/dL.
RR (95% CI); no. of deaths
Lung (above vs. below median):
Total cohort 1.5 (0.7, 2.9); 71
M 1.2(0.6, 2.5); 52
F 2.5 (0.7, 8.4); 19
Stomach (above vs. below median):
Total cohort 2.4 (0.3, 19.1); 5
M 3.1(0.3, 37.4); 4
F no deaths in referent group
All cancer: total cohort by quartile (age-adjusted) 1.0, 1.2, 1.3, 1.5 (P for
trend 0.16).
Smoking was controlled for. Lead levels occurring in the general population
were examined, not just those in workers with high occupational exposure
potential. Exposure to other carcinogens were not examined. Potential for
residual confounding by degree and duration of smoking exists (only
controlled for never, former, current <1, current 1+ pack/day). Limited case
numbers yield low statistical power for stomach or other cancers.
RR (95% CI)
All cancer, vs. low exposure:
Medium 1.5 (0.9, 2.5)
High 1.7(1.0,2.8)
Lung, vs. low exposure:
Medium 1.7(0.6,4.8)
High 2.2(0.8,6.1)
Non-lung, vs. low exposure:
Medium 1.5 (0.8, 2.8)
High 1.5 (0.8, 2.8).
Significant upward trends noted for all-cause and for cardiovascular mortality
with increasing lead category.
Smoking was controlled for. Lead levels occurring in the general population
were examined , with individuals showing levels consistent with intense
occupational exposure excluded, thus allowing exploration of potential effects
outside of groups experiencing intense occupational exposure. Exposure to
other carcinogens were not examined. Potential for residual confounding by
degree and duration of smoking exists (only controlled for never, former,
current <1, current 1+ pack/day). Limited case numbers yield low statistical
power for stomach or other cancers.
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Table AX6-7.4. Other Studies of Lead Exposure and Cancer
to
o
o
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings and Interpretation
O
o
2
o
H
O
o
HH
H
W
United States
Mallinetal. (1989)
Illinois
1979-1984
Coccoetal. (1998a)
U.S.
1984-1992
Coccoetal. (1998b)
U.S.
1984-1992
Case-control design.
Cases: random sample of 10,013 deaths
from 7 specific cancers, identified from
death certificates for Illinois males
between 1979 and 1984.
Controls: 3,198 randomly selected deaths
from other causes.
Odds of exposure computed for glass
workers vs. other occupations.
Case-control design.
Cases: all 27,060 brain cancer deaths
occurring among persons aged 35 or older
during 1984-1992, from U.S.
24-state death certificate registry.
Controls: 4 gender-, race-, age-, and
region-matched controls per case selected
from deaths due to nonmalignant causes.
Subjects were subdivided into 4 groups by
gender and race (white or African-
American) for all analyses.
Case-control design.
Cases: all 28,416 CNS cancer deaths
occurring among persons aged 35 or older
during 1984-1992, from U.S.
4-state death certificate registry.
Controls: 4 gender-, race-, age-, and
region-matched controls per case selected
from deaths due to nonmalignant causes.
Subjects were subdivided into 4 groups by
gender and race (white or African-
American) for all analyses.
Exposure was based on
occupations abstracted from death
certificates.
No specific measure of lead
exposure; glass workers can be
considered potentially exposed.
A job-exposure matrix was
applied to death certificate-listed
occupations to categorize persons
as having low, medium, or high
probability and intensity of
exposure.
Death certificate listed industry
and occupation was used to
categorize decedents. No
estimates of lead exposure
specifically.
Brain cancer, white male glass workers:
OR = 3.0, P < 0.05 (significant)
No significant associations for other cancer sites.
No control for smoking or other risk factors. Poor specificity for lead
exposure.
Risk of brain cancer mortality increased consistently with intensity of
exposure among African-American males, but not other race-gender
groups.
Probability of exposure alone was not consistently associated with risk.
In the high-probability group, risk increased with exposure intensity for
all groups except African-American women (only 1 death in the high-
probability group).
Exposure estimate was based solely on occupation listed on death
certificate, hence there was substantial opportunity for misclassification.
OR (95% CI)
All occupations or industries with ORs above 1.0 and P-value <0.05 in at
least one race-gender group were reported
Newspaper printing and publishing industry:
white M 1.4(1.1-1.8)
black M 3.1(0.9-10.9)
Typesetting and compositing:
white M 2.0(1.1-3.8)
white F 1.3(0.4-3.8)
black F 4.2 (0.6-30.7)
No deaths among black males.
Only two lead exposure associated occupations or industries showed a
statistically significant elevation of mortality. No specific measures of
lead exposure. Occupation based solely on death certificate, hence there
was substantial opportunity for misclassification.
-------
Table AX6-7.4 (cont'd). Other Studies of Lead Exposure and Cancer
to
o
o
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings and Interpretation
O
o
2
o
H
O
United States (cont'd)
Coccoetal. (1999)
U.S.
1984-1996
Case-control design.
Cases: all 41,957 stomach cancer deaths
occurring among persons aged 35 or
older during 1984-1996, from U.S. 24-
state death certificate registry.
Controls: 2 gender-, race-, age-, and
region-matched controls per case
selected from deaths due to
nonmalignant causes.
Subjects were subdivided into 4 groups
by gender and race (white or African-
American) for all analyses.
A job-exposure matrix was
applied to death certificate-
listed occupations to categorize
persons as having low, medium,
or high probability and intensity
of exposure.
OR (95% CI)
Adjusted for age, ethnicity, marital status, urban residence, and
socioeconomic status.
Elevated ORs:
white F, high prob.
1.53(1.10-2.12)
black M, high prob.
1.15(1.01-1.32)
black F, high prob.
1.76(0.74-4.16)
Highly exposed group included 1,503 white and 453 black men and
65 white and 10 black women; no pattern of increase across exposure
levels.
o
HH
H
W
Intensity of exposure showed no association with stomach cancer
except for black women:
Low
1.82 (1.04-3.18) (significant)
Moderate
1.39
High
1.25.
No control for other occupational exposures. Exposure estimate
based on occupation listed on death certificate and hence subject to
misclassification due to missing longest-held job.
-------
Table AX6-7.4 (cont'd). Other Studies of Lead Exposure and Cancer
to
o
o
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings and Interpretation
Canada
Rischetal. (1988)
Canada
1979-1982
Siemiatycki et al.
(1991)
Canada
Case-control design.
Cases: 826 Canadian men with
histologically confirmed bladder cancer
during 1979-1982.
Controls: 792 controls from Canadian
population, matched on age, gender, and
area.
Odds of exposure to lead for cases vs.
controls were computed, adjusted for
smoking and other risk factors.
Case-control design.
Cases: 3,730 various histologically
confirmed cancers.
Controls: specific cancer types were
compared with other cancers as a control
group, excluding lung cancer.
Separate subgroup analysis was
restricted to French Canadians.
Subjects were interviewed
regarding length of
occupational exposure to lead
compounds, as well as 17 other
substances.
Occupational exposure to
293 substances, including lead,
was estimated from interviews.
Exposure was classified as
"any"; a subgroup with
"substantial" exposure also was
identified.
OR (95% CI)
61 men ever exposed to lead (smoking-adjusted):
2.0(1.2-3.5)
Trend per 10 years' duration of exposure:
1.45 (1.09-2.02) (significant).
No other substances showed significant associations with bladder
cancer.
Controlled for smoking, marital status, socioeconomic status,
education, ethnicity, and urban vs. rural residence.
No control for other occupational exposures. Low control interview
rate (53%), which could result in biased control sample.
OR (90% CI); no. of cases
Any exposure to lead:
Lung 1.1 (0.9-1.4); 326
(French Canadians only)
Stomach 1.2 (1.0-1.6); 126
Bladder 1.3 (1.0-1.6); 155
(French Canadians only)
Kidney 1.2 (1.0-1.6); 88
ORs rose in the "substantial" exposure subgroup for stomach and
lung, but not for bladder or kidney cancer.
Controlled for smoking but not for other occupational exposures.
-------
Table AX6-7.4 (cont'd). Other Studies of Lead Exposure and Cancer
to
o
o
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings and Interpretation
O
o
2
o
H
O
o
HH
H
W
Europe
Sankilaetal. (1990)
Finland
1941-1977
Kauppinen et al.
(1992)
Finland
1976-1981
Cohort design.
1,803 male and 1,946 female glass
workers employed for at least 3 months
at one of 2 Finnish glass factories in
1953-1971 or 1941-1977.
Cancer incidence was compared with
age-, gender-, and calendar-year-specific
national rates.
Stomach, lung, and skin cancer rates also
were compared separately for 201 male
and 34 female glassblowers and non-
glassblowers.
Case-control design.
Cases: 344 primary liver cancer deaths
reported to the Finnish Cancer Registry
in 1976-1978 or 1981.
Controls: registry-reported stomach
cancer (476) or myocardial infarction
(385) deaths in the same hospitals,
frequency matched by age and gender.
No specific lead exposure
indices were computed.
Analyses did examine glass
workers as a whole and then
glassblowers specifically,
which comprised the group at
highest risk for lead exposure.
Questionnaires regarding job
history and personal habits were
sent to the closest available
relative.
U.K. based job-exposure matrix
was used to rate potential
exposure to 50 substances,
including lead compounds
Industrial hygienists also
inspected histories to identify
those with highly probable
exposure and rate it as high,
low, or moderate (<10 years
high or 10+ years low
exposure)
SIR (95% CI); no. of cases
Lung cancer, all glass workers:
male 1.3 (1.0-1.7); 62
female 1.1 (0.5-2.3); 7
Lung cancer risk showed no specificity for glassblowers.
Skin cancer, M& F combined:
All workers 1.5 (0.8-2.7); 11
little difference between genders
Glassblowers 6.2 (1.3-18.3); 3
Stomach cancer,M&Fcombined:
Glassblowers 2.3 (0.9-5.0); 6
No increase in other glass workers
No increase in cancers of other sites. No control for smoking
or occupational coexposures.
OR (95% CI)
52 workers with potential lead exposure'.
0.91(0.65-1.29)
11 -women -with potential lead exposure'.
1.84(0.83-4.06)
J men -with probable moderate exposure:
2.28 (0.68-7.67)
None had high exposure and only 1 had low exposure, whereas
4 controls had high exposure.
Female controls appeared to underreport their job history.
Most controls had stomach cancer, which if caused by lead
would bias results toward the null. Few subjects were rated as having
a high probability of exposure.
-------
Table AX6-7.4 (cont'd). Other Studies of Lead Exposure and Cancer
to
o
o
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings and Interpretation
O
o
2
o
H
O
o
HH
H
W
Europe (cont'd)
Wesseling et al. (2002)
Finland
1971-1995
Pesch et al. (2000)
Germany
1991-1995
Case-control design.
Cases: 935 renal-cell cancer patients
in five German areas.
Controls: 4,298 region, age, and
gender-matched controls from the
surrounding population.
ORs were adjusted for age, center,
and smoking.
Cohort design, but at ecologic
level.
413,877 Finnish women with
occupation reported in 1970
linked to Finnish Cancer
Registry to identify new cases
of brain or nervous system
cancer arising form 1971 to
1995.
Poisson regression was used to
calculate SIRs for exposed vs.
unexposed groups.
Job histories were used to
categorize exposure to
cadmium, lead, and other
potential as low vs. medium,
high, or substantial. Separate
exposure estimates were
obtained from British and from
German-derived job-exposure
matrices.
Reported occupation in 1970 was used to classify women into job
titles. Potential exposure for each job title was estimated using a job
matrix after excluding women in the highest social classes or in
farming. Lead and 23 other workplace agents examined. Rates for
each job title were calculated, and SIRs for low and medium/high
exposure calculated (average estimated blood lead of 0.3 jimol/L
served as cutpoint between low and medium/high exposure).
OR (95% CI); no. of cases
Substantial lead exposure based on British matrix:
M
1.5 (1.0-2.3); 29
F
2.6(1.2-5.5; 11
Substantial lead exposure based on British matrix:
M 1.3 (0.9-2.0); 30
F
Kandiloris et al. (1997)
Greece
Case-control design.
Cases: 26 patients with histologically
confirmed laryngeal carcinoma and no
history of lead exposure or toxicity.
Controls: 53 patients with similar
demographic profiles and no history of
cancer from the same hospital.
Blood lead levels and ALAD
activity were measured.
not reported.
Analyses controlled for smoking. No control for exposure to other
occupational agents.
Blood lead levels were similar, but ALAD activity was significantly
lower in cases than controls (Mean 50.79 U/L vs. 59.76 U/L,
p<0.01). No control for other risk factors. Potential distortion by
effects of disease on Pb and/or ALAD parameters.)
-------
Table AX6-7.4 (cont'd). Other Studies of Lead Exposure and Cancer
to
o
o
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings and Interpretation
O
o
2
o
H
O
o
HH
H
W
Europe (cont'd)
Cordiolietal. (1987)
Italy
1953-1967
Coccoetal. (1994a)
(expansion of Carta
etal. 1994).
Sardinia
1931-1992
Coccoetal. (1994b)
Sardinia
1951-1988
Cohort design.
1,741 male Sardinian lead and zinc
miners from two mines employed at
least one year between 1931 and 1971.
Mortality traced through 1992 to
determine cause of death.
Mortality among miners was compared
with age- and calendar-year-specific
regional rates to compute an SMR.
Cohort design.
526 female Sardinian lead and zinc
miners from the same mines as in Cocco
etal. (1994a).
Mortality traced through 1992 to
determine cause of death.
Mortality among miners was compared
with age- and calendar-year-specific
regional rates to compute an SMR.
Cohort design.
468 Italian glass workers
employed for at least one year
between 1953 and 1967.
Mortality among workers was
tracked and cause of death was
determined for deceased
workers. Standardized
mortality ratios were computed
for workers vs. national
population counterparts.
All miners were considered to
be exposed to lead.
All miners were considered to
be exposed to lead.
Workers producing low-quality glass containers were classified as
lead-exposed.
SMR (95% CI); no. of deaths
All cancer
1.21; 16
1.15; 17
1.28; 7
1.17; 8
0.61; 8
0.91; 21
0.83; 86
3.67 (1.35-7.98); 6
0.94 (0.83-1.05); 293
Prostate
Bladder
Kidney
Nervous system
Oral
Lymphohemopoietic
Digestive
Peritoneum
(significant)
No other P-values <0.05.
No control for smoking or exposure to silica, radon, or
other exposures.
SMR (95% CI)
Liver 5.02 (1.62-11.70) (significant)
Lung 2.32 (0.85-5.05) (nonsignificant)
Other cancers showed nonsignificantly reduced rates.
No control for smoking or exposure to silica, radon, or other
exposures. Low statistical power due to small population and paucity
of cancers during follow-up.
-------
Table AX6-7.4 (cont'd). Other Studies of Lead Exposure and Cancer
to Reference, Study
O Location, and Period
Study Description
Lead Measurement
Findings and Interpretation
Europe (cont'd)
Coccoetal. (1996)
Sardinia
1973-1992
Coccoetal. (1997)
Sardinia
1931-1992
Cohort design.
1,388 male production and maintenance
workers employed for at least 1 year at a
Sardinian lead and zinc smelter between
June of 1932 and July of 1971.
Mortality was followed up through 1992.
Mortality was compared with age- and
calendar-year-specific regional rates.
Since regional rates were only available
for 1965 and later, analyses were limited
to this period.
Cohort design.
1,222 male Sardinian lead and
zinc smelter workers whose
G6PD phenotypes had been
determined, employed any time
from 1973-1990.
Mortality traced through 1992
to determine cause of death.
Mortality was compared with
regional rates.
All workers were considered to
be exposed to lead.
All workers were considered to be exposed to lead.
Workers were subdivided into
6PD-normal and -deficient groups.
SMRs vs. regional rates (95% CI); no. of deaths
Lung 0.82 (95% CI 0.56-1.16); 31
Stomach 0.97 (0.53-1.62); 14
All cancers 0.93 (0.78-1.10); 132
Kidney 1.75 (0.48-4.49); 4
Bladder 1.45 (0.75-2.53); 12
Brain 2.17 (0.57-5.57); 4
Kidney cancer showed a significant trend toward increasing risk with
increasing duration of exposure
No significant trends were noted for lung or other cancers
Brain cancer excess was limited to workers employed for 10 years or
less.
No control for smoking or exposure to arsenic or other smelter-
related exposures. No data on intensity of exposure.
Strong association of smelter work with pneumoconiosis and other
respiratory disease (SMR = 4.47, 95% CI = 3.37 to 5.80); since this
outcome includes silicosis, which is thought to predispose individuals
to lung cancer, some lung cancer deaths may have been missed due to
misclassification of cause of death based on death certificates.
-------
Table AX6-7.4 (cont'd). Other Studies of Lead Exposure and Cancer
to
o
o
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings and Interpretation
O
o
2
o
H
O
o
HH
H
W
Europe (cont'd)
Wingren and Axelson (1987, 1993)
(update of Wingren and Axelson, 1985, same basic cohort as in
Wingren and Englander (1990)
Sweden
1950-1982
Wingren and
Englander (1990)
Sweden
1964-1985
(same population as in
case-control analyses
of Wingren and
Axelson 1985,1987,
1993)
Dingwall-Fordyce
and Lane (1963)
U.K.
1925-1962
Cohort design.
625 Swedish glass workers employed for
at least 1 month between 1964 and 1985.
Mortality was compared with national
rates.
Cohort design.
425 male employees drawing pensions
from U.K. battery plants.
Standardized mortality for employees vs.
national population counterparts.
Case-control design.
Source population: 5,498 men
aged 45 or older in 11 Swedish
parishes, including 887 glass
workers.
Cancer-specific nested case-
control analysis:
Cases: deaths due to stomach,
colon, and lung cancer from
1950-1982
Controls: deaths due to causes
other than cancer or
cardiovascular disease
Workers from areas with
airborne lead levels up to 0.110
mg Pb/m were classified as
exposed.
Battery plant workers were
assumed to be exposed, and
their mortality compared to that
of like age and gender in the
U.K. population as a whole.
Urinary lead excretion was also
used to categorize workers by
estimated exposure (none, light,
or heavy): 80 lightly and 187
heavily (at least 100 |ig/L)
exposed.
Glass workers were considered exposed.
Glassblowers also singled out as workers with higher exposure
potential.
Job history applied to job matrix to categorize occupations as low,
moderate, or high lead exposure.
SMR (95% CI)
Pharyngeal:
9.9 (1.2-36.1) (significant)
Lung:
1.4 (0.5-3.1) (nonsignificant)
Colon: (nonsignificant)
SMR (95% CI); no. observed deaths
All cancer:
1.2 (0.8-1.7); 267
No consistent increase in SMRs across categories of increasing lead
exposure.
Limitations: No cancer site-specific analyses. No control for
potential confounders including smoking and exposure to arsenic or
other metals.
-------
Table AX6-7.4 (cont'd). Other Studies of Lead Exposure and Cancer
to
o
o
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings and Interpretation
O
o
2
o
H
O
o
HH
H
W
Europe (cont'd)
Malcolm and Barnett (1982) (follow-up of Dingwall-Fordyce and Lane,
1963)
U.K.
1925-1976
Ades and Kazantzis
(1988)
U.K.
1943-1982
Cohort design.
4,393 male zinc, lead, and cadmium smelter
workers.
(Workers bom after 1939 or who had worked
less than one year at the facility were excluded.)
Workers followed up for mortality.
Nested case-control analysis also conducted to
quantitatively assessed cadmium and,
secondarily, arsenic, lead, and other metal
exposures among 174 cases.
Cohort design.
1,898 lead-acid battery workers.
Mortality was traced for the lead-
acid battery workers to determine
cause of death. The proportion of
deaths due to cancer (all types and
major subcategories) among the
worker population was compared to
that seen in corresponding members
of the general population, yielding a
PMR.
Job histories were used to quantify
cadmium exposure and assign
ordinal ranks for exposure to lead
and other metals.
Standardized lung cancer mortality
ratio computed for workers vs.
national rates.
Job histories were reviewed to classify workers' lead
exposure as high, medium, or none.
SMR (95% CI); no. of deaths
Cohort:
Lung 1.25 (1.07-1.44) (174)
Increased significantly with duration of employment.
Nested case-control analyses did not implicate any
department or process, nor did cadmium, zinc, sulfur dioxide,
or dust exposure account for the observed increase.
Cumulative exposure to lead and to arsenic both showed
positive associations with lung cancer, but the relative
importance of these two exposures could not be determined.
Cadmium exposure did not account for the elevated SMR,
but analyses could not control for exposure, and were not
adjusted for smoking.
Asia
Huetal. (1998)
China
1989-1996
Case-control design.
Cases: 218 patients with histologically-
confirmed primary gliomas occurring during
1989-1996 at 6 Chinese hospitals.
Controls: 436 patients with non-neurological,
nonmalignant disease, matched by age, gender,
and residence from the same hospitals
(excluding one cancer-only center).
Patients were interviewed, and those
with factory or farm occupations
were further interviewed to identify
exposure to lead (or other
potentially toxic substances).
Occupational exposure to lead
Not reported for any glioma patients, but was reported for
4 controls.
No control for exposure to other occupational or
environmental agents.
-------
Table AX6-7.4 (cont'd). Other Studies of Lead Exposure and Cancer
to
o
o
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings and Interpretation
Asia (cont'd)
Huetal. (1999)
China
1989-1996
Shuklaetal. (1998)
India
1995-1996
Case-control design.
Cases: 383 patients with histologically
confirmed primary meningiomas
occurring during 1989-1996 at 6 Chinese
hospitals.
Controls: 366 patients with non-
neurological, nonmalignant disease
matched by age, gender, and residence
from the same hospitals (excluding one
cancer-only center).
Case-control design.
Cases: 38 patients with newly
diagnosed, histologically confirmed gall
bladder cancer cases assembled from a
surgical unit.
Controls: 58 patients with gall stones
diagnosed at the same surgical unit,
matched on geographic area.
Mean bile lead content was compared
between cases and controls.
Patients were interviewed, and
those with factory or farm
occupations were further
interviewed to identify
exposure to lead (or other
potentially toxic substances).
Heavy metal content was
measured in bile drawn from
the gall bladder at time of
surgery.
OR (95% CI); no. of cases
Occupational exposure to lead:
M
7.20 (1.00-51.72); 6
F
5.69(1.39-23.39); 10
Results were adjusted for income, education, and fruit and vegetable
intake, plus cigarette pack-years for the women. No control for
exposure to additional metals or other occupational exposures.
Bile lead content: mean (SE) (mg/L):
Gall bladder cancer: 58.38 (1.76)
Gallstones: 3.99(0.43)
Cadmium and chromium levels also were elevated in cancer patients,
but less than lead. No control for smoking or any other risk factors.
-------
ANNEX TABLES AX6-8
May 2006 AX6-181 DRAFT-DO NOT QUOTE OR CITE
-------
Table AX6-8.1. Effects of Lead on Immune Function in Children
to
o
o
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
X
Oi
oo
to
H
6
o
o
H
O
o
H
W
O
O
HH
H
W
United States
Joseph et al. (2005)
S.E. Michigan
(1994-1998)
Sarasua et al. (2000)
ATSDR Multi-site
Study: Granite City,
IL, Galena, KA; Joplin,
MO; Palmerton, PA
1991
Rabinowtiz et al. (1990)
Boston, MA
1979-1987
Design: prospective (1 year following 3-year
baseline recruitment)
Subjects: children (n = 4634), age range:
0.4-3.0yr)
Outcome measures: asthma prevalence and
incidence.
Analysis: multivariate proportional hazard
model (Cox)
Design: cross-sectional
Subjects: children and adults (n = 2036)
Outcome measures: total lymphocyte count,
lymphocyte phenotype abundance, serum
IgA, IgG, and IgM.
Analysis: multivariate linear regression
Design: cross-sectional
Subjects: infants/children (n= 1768)
Outcome measures: incidence of illness in
children was solicited from parents by
questionnaire
Analysis: relative risk of illness estimated
from incidence ratios, highest: combined
lower blood lead deciles, without adjustment
for covariates or confounders.
Blood lead (ug/dL) mean
(SD, median, %>10):
5.5(4.0,4.0,8.6%)
Blood lead (ug/dL) mean
(SD, 5^-95^ %tile):
6-35 mo: 7.0(16,1.1-
16.1)
36-71 mo: 6.0(4.3,1.6-
14.1)
6-15 yr: 4.0(2.8,1.1-9.2)
16-75 yr: 4.3(2.9,1.0-
9.9)
Cord blood lead (ug/dL)
~90th%ti\e: 10
Shed tooth lead (ug/g)
~90th%ti\e: 5
Covariate-adjusted hazards ratio (HR, asthma incidence <5 ug/dL
compared to >5 or >10 ug ML): Caucasian: >5 ug/dL, 1.4 (95%CI:
0.7-2.9); >10 ug/dL, 1.1 (95% CI: 0.2-8.4). African American:
>5 ug/dL, 1.0 (95%CI: 0.8-1.3); >10 ug/dL, 0.9 (95% CI: 0.5-1.4).
HR for asthma incidence in African Americans, compared to
Caucasians (<5 ug/dL) were: <5 ug/dL: 1.6 (95% CI: 1.4-2.0);
>5 ug/dL, 1.4(95%CI: 1.2-1.6); >10 ug/dL,2.1 (95% CI: 1.2-3.6).
Covariates included: average annual income, birth weight, and
gender.
Significant association (p < 0.05) between increasing blood lead and
increasing serum IgA, IgG, IgM, and B-cell abundance (%, no.), and
decreasing T-cell abundance (%) in 6-35 mo age category; adjusted
for age, sex, and study site. Comparison of outcome means across
blood lead quartiles (1st quartile as reference, [+], higher, [-] lower):
[+] lymphocyte count (4* quartile, p = 0.02), T-cell count (4th
quartile, p = 0.09), B-cell count (4th quartile, p < 0.01), B-cell % (4th
quartile, p = 0.09).
Relative risk (unadjusted) was elevated for the following illness
categories: severe incidence of ear infection, 1.2 (95% CI: 1.0-1.4),
other respiratory illness, 1.5 (96% CI: 1.0-2.3), school absence for
illness other than cold or flu, 1.3 (95% CI: 1.0-1.5)
-------
Table AX6-8.1 (cont'd). Effects of Lead on Immune Function in Children
g Reference, Study
O Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
United States (cont'd)
X
ON
oo
Lutzetal. (1999)
Springfield-Green Co,
MO
NR
Design: cross-sectional
Subjects: children (n = 279; age range: 9
mo-6 yr)
Outcome measures: differential blood cell
counts; lymphocyte phenotype abundance
(%); and serum IL-4, soluble CD25, CD27,
IgE and IgG (Rubella).
Analysis: nonparametric comparison of
outcome measures (adjusted for age) for
blood lead categories, correlation
Blood lead (ug/dL) range:
Blood lead categories:
<10 ug/dL, 10-14 ug/dL,
15-1 9 ug/dL, 20^15 ug/dL
Significant association (p < 0.05) between increasing blood lead
(categorical) and increasing serum IgE levels, after adjusting for age.
H
6
o
o
H
O
o
H
W
O
O
HH
H
W
Europe
Annesi-Maesano et al.
(2003)
France
1985,1992
Karmaus et al. (2005)
Germany
1994-1997
Design: cross-sectional
Subjects: mother/newborn pairs (n = 374),
mean age 30 yr
Outcome measures: maternal venous and
newborn cord serum IgE levels
Analysis: multivariate linear regression,
ANOVA
Design: cross-sectional
Subjects: children (n = 331, 57% male), age
7-8 yrs (96%), 9-10 yrs (4%)
Outcome measures: differential blood cell
count; lymphocyte phenotype abundance;
and serum IgA, IgE, IgG, IgM
Analysis: multivariate linear regression
Blood lead (ug/dL) mean
(SD)
infant cord: 67.3(47.8)
maternal: 96.4(57.7)
Hair lead (ppm) mean
(SD):
infant: 1.38(1.26)
maternal: 5.16(6.08)
Blood lead (ug/dL) mean
(95% CI):
males: 2.5(1.1^.4)
females (2.8 (1.5^.8)
Blood lead quartile ranges:
<2.2 (n = 82)
2.2-2.8 (n= 81)
2.8-3.4 (n= 86)
>3.4(n=82)
Significant (p < 0.0001) association between increasing infant hair
lead and infant cord serum IgE levels.
Although medical histories were taken to identify potential IgE risk
factors (asthma, allergies) and "confounders" (e.g., smoking), these
do not appear to have been quantitatively integrated into the
regression models. Allergy status and blood levels were reportedly
unrelated to load biomarkers or serum IgE (basis for conclusion not
reported).
Significant association between blood lead (p < 0.05) and serum IgE
(not monotonic with quartile range). Comparison of adjusted mean
outcomes (p<0.05) across blood lead quartiles (1st quartile as
reference, [+], higher, [-] lower): [-] CD3+ T-cells (2nd quartile), [-]
C3+CD8+ T-cells (2nd quartile), [+] C3+CD5+CD19+B-cells (2nd
quartile).
Covariates retained: age, sex, environmental exposure to tobacco
smoke, infections (in last 12 mo), serum cholesterol, and
triglycerides.
-------
Table AX6-8.1 (cont'd). Effects of Lead on Immune Function in Children
g Reference, Study
O Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
X
ON
oo
H
6
o
o
H
O
o
H
W
O
O
HH
H
W
Europe (cont'd)
Reigart and Graber
(1976)
NR
NR
Wagnerova et al. (1986)
Czech
NR
Design: clinical
Subjects: children (n = 19), ages 4-6 years
Outcome measures: serum IgA, IgG, IgM,
total complement and C-3, before and after
immunization with tetanus toxoid
Analysis: none; presentation of prevalence
of clinically low, normal, and high values of
outcome measures
Design: longitudinal cohort (repeated
measures for 2-years)
Subjects: children (n = 92, 38 females) ages
11-13 yrs residing near a smelter; reference
group (n = 67, 36 females), ages 11-13 years
Outcome measures: serum IgA, IgE, IgG,
IgM
Analysis: comparison of outcome measures
and between exposed and reference groups,
stratified sex and season of sampling
Blood lead (|ig/dL) mean
(range):
high: >40(n= 12): 45.3
(41-51)
low: <30(n=7): 22.6
(14-30)
Blood lead (jig/dL) mean:
lead: -23^2
reference: —5-22
No apparent difference in prevalence of abnormal values for serum
immunoglobulin or complement (no statistical analysis applied).
Significant (p NR, statistic NR) lower serum IgE and IgM levels in
exposed group compared to reference group.
Latin America
Pineda-Zavaleta et al.
(2004)
Mexico
NR
Design: cross-sectional
Subjects: children (n = 30 female, 35 male)
ages 6-11 years, residing near smelter
Outcome measures: mitogen- (PHA) and
cytokine- (IFN-y) induced nitric oxide and
superoxide production in lymphocytes
Analysis: multivariate linear regression
Blood lead (ng/dL) mean
(range) for 3 schools:
l(n = 21): 7.0(3.5-25.3)
2(n = 21): 20.6
(10.8^9.2)
3(n = 23): 30.4
(10.3^7.5)
Significant (p = 0.036) association between increasing blood lead
concentration and covariate adjusted decreasing nitric oxide
production in PHA-activated lymphocytes (P = -0.00089, 95% CI:
-0.0017 to-0.00005).
Significant (p = 0.034) association between increasing blood lead
concentration and covariate adjusted increasing super oxide
production in IFN-y-activated lymphocytes (P = -0.00389, 95% CI:
0.00031 to 0.00748). Covariates considered included age, sex,
allergies, and blood arsenic (age, sex, and blood arsenic were
retained).
Significant effect of sex on associations, significant blood lead-
arsenic interaction.
Covariates considered included age, sex, allergies, urinary arsenic
(age, sex, and urinary arsenic were retained).
-------
Table AX6-8.1 (cont'd). Effects of Lead on Immune Function in Children
to
o
o
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
X
ON
oo
Asia
Sunetal. (2003); Zhao
et al. (2004)
China
NR
Design: cross-sectional
Subjects: children (n = 73) age 3-6 yrs
Outcome measures: serum IgE, IgG, IgM;
lymphocyte phenotype abundance
Analysis: Nonparametric comparisons of
outcome measures stratified by blood lead
Blood lead (ng/dL) mean
(SD, range) (n = 217):
9.5(5.6,2.6^3.7)
Females: significantly higher (p < 0.05) IgE levels in high blood
lead category (> 10 ng/dL, n = 16) compared to low category
(<10 ng/dL, n = 17), and significantly lower IgG and IgM levels.
A ultivariate analysis of association between blood lead and IgE was
noted but not described in sufficient detail to evaluate.
All children: significantly lower (p < 0.05) CD3+CD4+ (%),
CD3+CD8+ (%), CD4+CD8+ (%) in high blood lead (> 10 ng/dL,
n = 38) compared to low blood lead (10 |ig/dL, n = 35) group.
H
6
o
o
H
O
o
H
W
O
O
HH
H
W
ANOVA, analysis of variance; CI, confidence interval; Ig, immunoglobulin; IFn-y interferon- y; IgG, immunoglobulin G; IgM, immunoglobulin M; IL-4, interleukin-4; NR, not
reported; PHA, phytohemagglutinin; SD, standard deviation
-------
Table AX6-8.2. Effects of Lead on Immune Function in Adults
to
o
ON
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
United States
X
ON
oo
Pinkertonetal. (1998)
U.S.
NR
Design: cross-sectional cohort
Subjects: adult male smelter workers
(n = 145, mean age 32.9±8.6); reference
group, male hardware workers (n = 84, mean
age30.1±9.3)
Outcome measures: differential blood cell
counts; lymphocyte phenotype abundance;
serum IgA, IgG, IgM; salivary IgA;
lymphocyte proliferation (tetanus toxoid)
Analysis: multivariate logistic regression
with comparison of adjusted outcome
measures between exposed and nonexposed
groups
Blood lead (ug/dL) median
(range)
lead: 39(15-55)
reference: <2 (<2-12)
Covariate-adjusted outcomes in lead workers that were significantly
(p < 0.05) different from nonexposed ([+], higher, [-] lower): [-] %
monocytes, [-] % CD4+CD8+ cells, [-] % CD8+CD56+cells.
Significant (p < 0.05) adjusted regression coefficients in exposed
group for independent variable:
blood lead: [+] CD19+ B-cells (%, no)
time-integrated blood lead: [-] serum IgG, [+] CD4+CD45RA+ cells
(%, no.)
Covariates considered in the analysis included age, race, smoking
habits, alcohol consumption, marijuana use, work shift, and various
factors that might stimulate or suppress the immune system (e.g.,
exposure to direct sunlight, sleep hours, allergy, flu or cold
symptoms). Covariates retained in the final model were age, age,
race, work shift, smoking habits.
H
6
o
o
H
O
o
H
W
O
O
HH
H
W
Sarasua et al. (2000)
ATSDR Multi-site
Study: Granite City,
IL, Galena, KA; Joplin,
MO; Palmerton, PA
1991
Design: cross-sectional cohort
Subjects: children and adults (n = 2036)
Outcome measures: total lymphocyte count,
lymphocyte phenotype abundance, serum
IgA, IgG, and IgM
Analysis: multivariate linear regression
Blood lead (ug/dL) mean
(SD, 5^-95^ %tile):
6-35 mo: 7.0(16,1.1-
16.1)
36-71mo: 6.0(4.3,1.6-
14.1)
6-15 yr: 4.0(2.8,1.1-9.2)
16-75 yr: 4.3(2.9,1.0-
9.9)
No significant association (<0.05) between blood lead and outcomes
in adults (age > 16 yr).
Covariates retained: age, sex, cigarette smoking, and study site.
-------
Table AX6-8.2 (cont'd). Effects of Lead on Immune Function in Adults
g Reference, Study
O Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
United States (cont'd)
X
ON
oo
Fischbein et al. (1993)
New York
NR
Design: cross-sectional cohort
Subjects: adult firearms instructors (n = 51),
mean age 48 yr; age-matched reference
subjects (n = 36).
Outcome measures: lymphocyte phenotype
abundance, lymphocyte proliferation (PHA,
PWM, Staph. aureus)
Analysis: comparison of outcome measures
between reference and blood lead categories;
multivariate linear regression
Blood lead (|ig/dL) mean
(SD)
lead high (>25): 31.4(4.3)
lead low (<25): 14.6(4.6)
reference: <10
Outcomes in lead workers that were significantly (p < 0.05) different
from reference group ([+], higher, [-] lower):
[-] CD+3 cells (%, no.), [-] CD4+ cells (%, no.), [-] CD4+CD8+ cells
(no.), [-] HLA-DR cells (no.), [+] CD20+ cells (%, no.), [-] mitogen
(PHA)-induced lymphocyte proliferation, [-] mitogen (PWM)-
induced lymphocyte proliferation; [-] lymphocyte response in
mixed-lymphocyte culture. No effect on antigen (Staph. aureus)-
induced lymphocyte proliferation.
Significant (p < 0.05) association between increasing blood lead and
decreasing abundance of CD4+ phenotypes (%), and decreasing
lymphocyte proliferative response in mixed lymphocyte cultures.
Covariates retained: age, sex, smoking habits, and duration of
exposure.
H
6
o
o
H
O
o
H
W
O
O
HH
H
W
Europe
Bergeret et al. (1990)
France
NR
Design: cross-sectional cohort
Subjects: adult battery smelting workers
(n = 34), mean age 40 yr; reference subjects
(n = 34), matched for age, sex, ethnic origin,
smoking and alcohol consumption habits,
intake of antibiotics, andNSAIDs
Outcome measures: PMN chemotaxis
(FMLP); PMN phagocytosis (opsonized
zymosan)
Analysis: comparison of outcome measures
between worker and reference groups
Blood lead (|ig/dL) mean
(SD):
lead: 70.6(18.)
reference: 9.0(4.3)
Significantly (p < 0.05) lower PMN chemotactic response (index)
and phagocytic response in lead workers.
-------
Table AX6-8.2 (cont'd). Effects of Lead on Immune Function in Adults
g Reference, Study
O Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
X
Oi
oo
oo
H
6
o
o
H
O
o
H
W
O
O
HH
H
W
Europe (cont'd)
Ewers etal. (1982)
Germany
NR
Cosciaetal. (1987)
Italy
NR
Governaetal. (1987)
Italy
NR
Design: cross-sectional cohort
Subjects: adult male battery manufacture or
smelter workers (n = 72), mean age 36.4 yr
(16-58); reference workers (n = 53), mean
age 34.8 yr (21-54)
Outcome measures: serum IgA, IgG, IgM,
C3; saliva IgA
Analysis: parametric and nonparametric
comparison of outcome measures between
lead workers and reference subjects; linear
regression
Design: cross-sectional cohort
Subjects: adult lead workers (n = 32, 2
female), mean age 42.8 yr (SD 11.5);
reference subjects (n = 25), mean age 38.6 yr
(SD 13.3)
Outcome measures: serum IgA, IgG, IgM,
C3-C4; lymphocyte phenotype abundance
Analysis: parametric comparison of
outcome measures between worker and
reference groups
Design: cross-sectional cohort
Subjects: adult male battery manufacture
workers (n = 9), mean age 38.4 yr (SD 13.7);
age-matched reference subjects (n = 18)
Outcome measures: PMN chemotaxis
(zymosan-activated serum)
Analysis: parametric comparison of
outcome measures between worker and
reference groups, correlation
Blood lead (|ig/dL) mean
(range):
lead: 55.4.0(18.6-85.2)
reference: 12.0 (6.6-20.8)
Blood lead (ng/dL) mean
(SD):
lead: 62.3(21.6)
reference: NR
Blood lead (|ig/dL) mean
(SD):
lead: 63.2(8.2)
reference: 19.2(6.4)
Significantly (p < 0.05) lower serum IgM, lower salivary IgA in lead
workers compared to reference group.
Outcomes in lead workers that were significantly (p < 0.05) different
from reference group ([+], higher, [-] lower): [-] serum IgM, [+]
serum C4, [+] lymphocyte abundance (%), [-] T-cell abundance (%,
no., E-rosette forming cells), [+] B-cell abundance (%,no.,
immunoglobulin-bearing cells), [+] CD8+ cell abundance (no.).
Significantly (p < 0.05) lower PMN chemotactic response to
zymosan activated serum. Effect magnitude was not correlated with
blood lead.
-------
Table AX6-8.2 (cont'd). Effects of Lead on Immune Function in Adults
to
o
o
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
X
Oi
oo
VO
H
6
o
o
H
O
o
H
W
O
O
HH
H
W
Europe (cont'd)
Valentino etal. (1991)
Italy
NR
Kimberetal. (1986)
UK
NR
Design: cross-sectional cohort
Subjects: adult male lead scrap refining
workers (n = 10), mean age 41.1 yr (SD 7.3,
range: 28-54); age-matched reference
subjects (n= 10)
Outcome measures: PMN chemotaxis (C5 or
FMLP) and phagocytosis (FMLP)
Analysis: comparison of outcome measures
between worker and reference groups,
correlation
Design: cross-sectional cohort
Subjects: adult male TEL manufacture
workers (n = 39) mean age: 45.1 yr; and
age-matched reference subjects (n = 21);
mean age 32.2 yr
Outcome measures: serum IgA, IgG, IgM;
mitogen (PHA)-induced
lymphoblastogenesis; and NK cell
cytotoxicity
Analysis: comparison of outcome measures
for exposed and reference groups
Blood lead (ug/dL) mean
(SD, range):
lead: 33.2(5.6,25^12)
reference: 12.6 (2.5, 8.9-
18)
Significantly (p < 0.002) lower PMN chemotactic response to C5 or
FMLP and higher stimulated production of LT (leukotriene)B4 in
lead workers compared to reference group. Effect magnitude
correlated with blood lead. No effect on phagocytic activity.
Blood lead (ug/dL) mean
(SD, range):
lead: 38.4(5.6,25-53)
reference: 11.8(2.2,8-17)
No significant (p < 0.05) differences in outcomes between exposed
and reference groups.
Latin America
Queiroz etal. (1993)
Brazil
NR
Design: cross-sectional cohort
Subjects: adult male battery manufacture
workers (n = 39), mean age 33.9 yr (SD 12.1,
range: 18-56); reference subjects (n = 39)
matched by age and race
Outcome measures: PMN chemotaxis
(endotoxin LPS); phagocytic (endotoxin
LPS) respiratory burst activity (NBT
reduction)
Analysis: nonparametric comparison of
outcome measures between worker and
reference groups
Blood lead (ug/dL) range:
lead: 14.8-91.4 (>30,
n=52)
reference: <10
Significantly (p < 0.001) lower chemotactic activity of PMNs, and
lower phagocytic respiratory burst, in lead workers relative to
reference group.
-------
Table AX6-8.2 (cont'd). Effects of Lead on Immune Function in Adults
to
o
o
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
X
Oi
H
6
o
o
H
O
o
H
W
O
O
HH
H
W
Latin America (cont'd)
Queirozetal. (1994a)
Brazil
NR
Queirozetal. (1994b)
Brazil
NR
Design: cross-sectional cohort
Subjects: adult male battery manufacture
workers (n = 60), mean age 33.9 yr (range:
18-56); reference subjects (n = 49) matched
by age and race
Outcome measures: PMN phagocytic/lytic
activity (opsonized yeast)
Analysis: nonparametric comparison of
outcome measures between worker and
reference groups
Design: cross-sectional cohort
Subjects: adult male battery manufacture
workers (n = 33), mean age 32.4 yr (range:
18-56); reference subjects (n = 20) matched
by age and race
Outcome measures: serum IgA, IgG, IgM;
mitogen (PHA)-induced lymphocyte
proliferation
Analysis: parametric comparison of
outcome measures between worker and
reference groups
Blood lead (ug/dL) range:
lead: 14.8-91.4 (>30,
n = 27)
reference: <10
Significantly (p < 0.001) lower lytic activity of PMNs in lead
workers relative to reference group.
Blood lead (ug/dL) range:
lead: 12.0-80.0 (>30,
n = 27)
reference: <10
No significant difference in outcomes (p < NR; SD of lead worker
and reference groups overlap) between lead workers and reference
group.
Asia
Kuoetal. (2001)
China
NR
Design: cross-sectional cohort
Subjects: adult battery manufacture workers
(n = 64, 21 female), ages: <40 yr 14, >50yr,
14); nonexposed reference subjects (n = 34,
17 female).
Outcome measures: differential blood cell
counts, lymphocyte phenotype abundance
Analysis: comparison of outcome measures
in exposed and reference groups,
multivariate linear regression
Blood lead (ug/dL) mean:
lead workers: 30
Significantly (p < 0.05) adjusted mean higher monocytes (%, no.),
lower B cells (%), lower lymphocytes (no.), and lower granulocytes
(no.) in lead workers compared to controls.
Covariates retained: age, gender, and disease status (definition not
reported).
-------
Table AX6-8.2 (cont'd). Effects of Lead on Immune Function in Adults
to
o
o
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
X
Oi
H
6
o
o
H
O
o
H
W
O
O
HH
H
W
Asia (cont'd)
Mishra et al. (2003)
India
NR
Alomran and
Shleamoon(1988)
Iraq
NR
Cohen etal. (1989)
Israel
NR
Design: cross-sectional cohort
Subjects: adult males occupationally
exposed to lead (n = 84), mean age 30 yr;
reference subjects (n = 30), mean age 29 yr
Outcome measures: serum IFN-y level,
mitogen (PHA)-induced lymphocyte
proliferation, NK cell cytotoxicity
Analysis: comparison of outcome measures
between lead-exposed and reference groups,
correlation
Design: cross-sectional cohort
Subjects: adult lead (oxide) workers
(n = 39), mean age 35.6 yr (9,2, SD); age-
matched reference subjects (n = 19)
Outcome measures: serum IgA, IgG;
mitogen (PHA, Con-A)-induced lymphocyte
proliferation
Analysis: comparison of outcome measures
between lead workers and reference group
Design: cross-sectional cohort
Subjects: adult male occupationally lead
exposed (n = 10), age range 22-70; age-
matched reference subjects (n = 10)
Outcome measures: mitogen (Con A, PHA)-
induced-lymphocyte proliferation and T-
suppressor cell proliferation; lymphocyte
phenotype abundance
Analysis: parametric comparison of
outcome means between lead-exposed and
reference groups
Blood lead (ng/dL) mean
(SD, range):
3-wheel drivers (n = 30):
6.5(4.7,0.0-17.5)
battery workers (n = 34):
128.1 (13.2^100.8)
jewelry makers: 17.8
(18.5,3.1-76.8)
reference: 4.5 (NR, 1.6-
9.8)
Blood lead (jig/dL) mean:
lead: 54-64
reference: NR
Blood lead (ng/dL) range:
exposed: 40-51
reference: <19
Significantly (p < 0.001) lower lymphocyte proliferative response to
PHA in lead-exposed groups compared to reference groups, higher
IFN-y production by blood monocytes.
Significantly (p < 0.05) lower lymphocyte proliferative response to
PHA or Con A in lead workers, compared to reference group.
Significantly (p < 0.02) higher mitogen (Con-A)-induced suppressor
cell activity. No significant (p not reported) effects on abundance of
T-cells (E-rosette-forming cells), OKT+4, OKT+8, or OKT4+/T8+
ratio; mitogen (Con A or PHA)-induced lymphocyte proliferation.
-------
Table AX6-8.2 (cont'd). Effects of Lead on Immune Function in Adults
to
o
o
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
X
Oi
to
H
6
o
o
H
O
o
H
W
O
O
HH
H
W
Asia (cont'd)
Sataetal. (1998)
Japan
NR
Design: cross-sectional cohort
Subjects: adult male lead stearate
manufacture workers (n = 71), mean age 48
yr (range: 24-74); reference subjects
(n = 28), mean age 55 yr (range: 33-67).
Outcome measures: lymphocyte phenotype
abundance
Analysis: comparison of outcome measures
in exposed and reference groups
(ANCOVA), multivariate linear regression
Sataetal. (1997) Design: clinical
Japan Subjects: adult male lead smelter workers
NR (n = 2) who underwent CaEDTA therapy
Outcome measures: serum IgA, IgG, IgD,
IgM; lymphocyte phenotype abundance
Analysis: Parametric comparison of
outcome measures before and after
treatment, correlation of outcome means with
blood lead
Heo et al. (2004) Design: cross-sectional cohort
Korea Subjects: adults, battery manufacture
NR workers (n = 606; 52 females); ages: <30 yr,
n=184;>40yr,n=123.
Outcome measures: serum IgE, IL-4, IFNy
Analysis: comparison of outcomes measures
(ANOVA), stratified by age and blood lead
Blood lead (ug/dL) mean
(range):
lead: 19(7-50)
reference: NR
Blood lead (ug/dL):
subject 1: 81 ug/dL at
referral; mean before
EDTA: 45.1(SD16.0);
after chelation: 31.0(9.8)
subject 2: 68 ug/dL at
referral; mean before
EDTA: 43.3 (SD 14.1);
after chelation: 33.7(7.2)
Blood lead (ug/dL) mean
(SD):
<30yr: 22.0(10.4)
30-39 yr: 23.0(11.3)
>40yr: 24.1(9.3)
Lead workers vs. reference: significantly (p < 0.05) covariate-
adjusted lower CD3+CD45RO+ (no.) and higher CD8+ cells (%).
Significant (p < 0.05) association between exposure (categorical:
yes/no) and lower CD3+CD45RO+cells (no.).
Covariates retained: age and cigarette smoking habits.
Blood lead and outcome measures were sampled prior to and 24
hours after 3 CaEDTA treatments (on consecutive days) per week
for 10 weeks. Comparison of mean outcome measures assessed
before and after treatments showed significantly (p < 0.05) higher
IgA, IgG, and IgM; and significantly higher CD8+ T-cells and
CD57+ NK cells after treatment in subject 1. Serum IgG levels in
subject 1 were significantly correlated (r-0.72) with blood lead
concentration.
Significantly higher (p < 0.05) serum IgE levels in blood lead
category (>30 ug/dL) compared to low categories (<10 or 10-
29 ug/dL).
-------
Table AX6-8.2 (cont'd). Effects of Lead on Immune Function in Adults
to
o
o
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
X
Oi
H
6
o
o
H
O
o
H
W
O
O
HH
H
W
Asia (cont'd)
Undegeretal. (1996);
Ba^aran and Undeger
(2000)
Turkey
NR
Yucesoy etal. (1997a)
Turkey
NR
Yucesoy etal. (1997b)
Turkey
NR
Design: cross-sectional cohort
Subjects: adult male battery manufacture
workers (n = 25), mean age, 33 yr (22-55);
reference subjects (n = 25) mean age 33 yr
(22-56).
Outcome measures: differential blood cell
counts; lymphocyte phenotype abundance;
serum IgA, IgG, IgM, C3, and C4; neutrophil
chemotaxis (zymosan-activated serum); latex
particle-induced neutrophil phagocytic (latex
particles) respiratory burst (NET reduction)
Analysis: nonparametric and parametric
comparisons of outcome measures for
exposed and reference groups
Design: cross-sectional cohort
Subjects: adult male battery manufacture
workers (n = 20), ages 39^8 yr; age-
matched reference subjects (n = 12)
Outcome measures: serum cytokines IL-1 p,
IL-2, TNFa, IFN-y
Analysis: parametric and nonparametric
comparison of outcome measures in exposed
and reference groups
Design: cross-sectional cohort
Subjects: adult male battery manufacture
workers (n = 50), ages 39^8 yr; age-
matched reference subjects (n = 10)
Outcome measures: lymphocyte phenotype
abundance, NK cell cytotoxicity
Analysis: comparison of outcome measures
in exposed and reference groups
Blood lead (|ig/dL) mean
(SD):
lead: 74.8(17.8)
reference: 16.7(5.0)
Blood lead (|ig/dL) mean
(SE, range):
lead: 59.4(3.2,42-94)
reference: 4.8(1.0,2-15)
Blood lead (|ig/dL) mean
(SE, range):
lead 1 (n = 20): 59.4 (3.2,
42-94)
lead 2 (n= 30): 58.4
(2.5,26-81)
reference: 4.0 (0.4, 2-6)
Workers relative to reference: significantly (p < 0.05) lower serum
IgG, IgM, C3, and C4 levels; lower CD4+ ("T-helper") abundance,
lower neutrophil chemotactic response; no significant difference in
CD20+ (B-cell), CD8+ ("T-suppressor") cell, CD56+ (NK) cell
abundance, or particle-induced NK cell respiratory burst.
Significantly (p < 0.05) lower serum IL-1 p and IFN-y levels in lead
workers compared to controls.
Significantly (p < 0.05) lower CD20+ B-cell (%) abundance in lead
workers compared to controls, no difference in % CD4+ T-cell
abundance.
-------
Table AX6-8.2 (cont'd). Effects of Lead on Immune Function in Adults
g Reference, Study
O Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
Africa
X
Oi
Anetor and Adeniyi
(1998)
Nigeria
NR
Design: cross-sectional cohort
Subjects: adult male "lead workers"
(n = 80), mean age, 36 yr (21-66) and
reference subjects (n = 50), mean age 37 yr
(22-58).
Outcome measures: serum IgA, IgG, and
IgM; lymphocyte count
Analysis: comparison of outcomes measures
in workers and reference group, linear
regression, principal component analysis
Blood lead (ug/dL) mean
(SE):
lead: 53.6(0.95)
reference: 30.4(1.4)
Significantly lower (p < 0.05) serum IgA, IgG, and total blood
lymphocyte levels; significant associations and interactions between
blood lead and serum total globulins (note high blood lead levels in
reference).
H
6
o
o
H
O
o
H
W
O
O
HH
H
W
ANOVA, analysis of variance; EDTA, ethylenediaminetetraacetic acid; FMLP, N-formyl-L-methionyl-L-leucyl-L-phenyl-alanine; IFn-y interferon-y; Ig, immunoglobulin A;
LPS, lipopolysaccharide; LT, leukotriene; NET, nitroblue tetrazolium; NK, natural killer; NR, not reported; NSAIDS, non-steroidal anti-inflammatory agents; PHA,
phytohemagglutinin; PWM pokeweed mitogen; SD, standard deviation; SE, standard error; TEL, tetraethyl lead
-------
ANNEX TABLES AX6-9
May 2006 AX6-195 DRAFT-DO NOT QUOTE OR CITE
-------
Table AX6-9.1. Effects of Lead on Biochemical Effects in Children
to
o
ON
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
United States
Marcus and Schwartz
(1987)
U.S.
1976-1980
Design: cross-sectional national survey
(NHANES II)
Subjects: ages 2-6 yr(n= 1677)
Outcome measures: EP, red blood cell count,
mean corpuscular volume, iron status variables
Analysis: nonlinear least squares regression
Blood lead (ug/dL)
range: 6-65
Non-linear regression used to fit kinetic model relating blood lead
to EP, in strata having low (<14%), medium (14-31%), or high
(>31%) percent transferrin saturation (PST). Parameters in model
included: parameters for total red cell surface area, maximum red
cell lead concentration, equilibrium concentration ratio for plasma
and whole blood. Blood lead increase (from 10 ug/dL) predicted
to double EP: 22 (PST < 14%), 24 (PST = 14-31%), 37
(PST > 31%).
X
ON
Piomellietal. (1982)
New York
1976
Design: cross-sectional
Subjects: children (n = 2002), ages 2-12 yr
Outcome measures: EP
Analysis: linear regression
Blood lead (ug/dL)
range: 2-98
Regression equation relating blood lead concentration to EP
(log-transformed):
a = 1.099, p = 0.016, r: = 0.509, p < 0.001
Threshold for increase in EP estimated to be: 1 5 .4 ug/dL (95%
CI: 12.9-18.2)
H
6
o
o
H
O
O
H
W
O
O
HH
H
W
Soldin et al. (2003)
Washington DC
2001-2002
Design: cross-sectional
Subjects: children (n = 4908, 1812 females),
age range 0-17 yr
Outcome measures: EP
Analysis: locally weighted scatter plot
smoother (LOWESS)
Blood lead (ug/dL):
mean (range 1-17 yr):
2.2-3.3
median (1-17 yr): 3
range: < 1-103
EP increases as blood lead concentration increased above 1 5
mg/dL. A doubling of EP occurred with an increase in blood lead
concentration of approximately 20 ug/dL (a polynomial
expression for EP as a function of blood lead (PbB) is:
EP = -O.OOlS(PbB)3 + 0.1854(PbB)2 - 2.7554(PbB) + 30.911
(r2 = 0.9986)
(derived from data in Table 2 of Soldin et al. (2003)
Europe
Roels and Lauwerys
(1987)
Belgium
1974-1980
Design: cross-sectional
Subjects: children (n = 143), age range 10-13
yr
Outcome measures: ALAD, urinary ALA, EP
Analysis: linear regression, correlation
Blood lead (ug/dL) range:
Linear regression for EP (log-transformed) and blood lead
concentration:
a = 1.321, p = 0.025, r = 0.73 (n = 51)
Linear regression for ALA (log-transformed) and blood lead
concentration:
a = 0.94, p = 0.11, r= 0.54 (n = 37)
Linear regression for ALAD (log-transformed) and blood lead
concentration:
a = 1.864, p = -0.015, r = -0.87 (n = 143)
-------
Table AX6-9.1 (cont'd). Effects of Lead on Biochemical Effects in Children
g Reference, Study
O Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
Latin America
H
6
o
o
H
O
o
H
W
O
O
HH
H
W
Perez-Bravo et al. Design: cross-sectional;
(2004) Subjects: children (n = 93, 43 males), age
Chile range: 5-12yrs who attended school near a
NR powdered lead storage facility
Outcome measures: blood Hgb and Hct, ALAD
genotype
Analysis: comparison of outcome measures
between ALAD genotype strata
Blood lead (|ig/dL) mean
(SE):
ALADl(n = 84): 13.5
(8.7)
ALAD 2 (n = 9): 19.2(9.5)
Mean blood lead, blood Hgb, and Hct not different between
ALAD genotypes (p = 0.13)
X
Oi
ALA, 5-aminolevulinic acid; ALAD, 5-aminolevulinic acid dehydratase; EP, erythrocyte protoporphyrin
-------
Table AX6-9.2. Effects of Lead on Biochemical Effects in Adults
to
o
o
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
X
Oi
oo
H
6
o
o
H
O
O
H
W
O
O
HH
H
W
Europe
Gennartetal. (1992)
Belgium
NR
Mohammed-Brahim
etal. (1985)
Belgium
NR
Roels and Lauwerys
(1987)
Belgium
1974-1980
Design: cross-sectional cohort
Subjects: adult battery manufacture workers
(n = 98), mean age, 37.7 yr (range 22-55);
reference group (n = 85), mean age 38.8 yr
(24-55)
Outcome measures: blood Hct, blood EP,
urine ALA
Analysis: linear regression
Design: cross-sectional cohort
Subjects: adult smelter and ceramics
manufacture workers (n = 38, 13 females);
reference subjects (n = 100) matched with
worker group by age, sex, and socioeconomic
status.
Outcome measures: blood P5N, EP, ALAD,
R/ALAD (ratio of ALAD before and after
reactivation).
Analysis: comparison of outcome measures
(ANOVA) between lead workers and reference
group; correlation
Design: cross-sectional
Subjects: adults (n = 75, 36 females)
Outcome measures: ALAD, urinary ALA, EP
Analysis: linear regression, correlation
Blood lead (ug/dL) mean
(SD, range):
lead: 51.0(8.0,40-70)
reference: 20.9
(11.1,4.4-30.0
Blood lead (ug/dL) mean
(SD, range):
lead: 48.5(9.1,27.8-66.6)
reference: 14.3 (6.7, 5.6-
33.6)
Urine lead (ug/g creatinine)
mean (SD, range):
lead: 84.0(95.9,21.8-587)
reference: 10.5(8.2,1.7-
36.9)
Blood lead (ug/dL) range:
adult males: 10-60
adult females: 7-53
Significant association between increasing blood lead
concentration and increasing (log) blood EP (a = 0.06, p = 0.019,
r = 0.87, p = 0.0001) or (log) urine ALA (a = 0.37, (3 = 0.008,
r = 0.64, p< 0.0001)
(No apparent analysis of covariables)
Significantly lower (p = NR) P5N in lead workers (males or
females, or combined) compared to corresponding reference
groups.
Correlations with blood lead:
log P5N r = -0.79 (p < 0.001)
log ALAD r = -0.97 (p = NR)
R/ALAD r = -0.94 (p < 0.001)
logEPr=0.86(p = NR)
Correlations with urine lead:
log P5Nr=-0.74 (p = NR)
log ALAD r = -0.79 (p = NR)
R/ALAD r = -0.84 (p < 0.001)
logEPr=0.80(p = NR)
Linear regression for EP (log-transformed) and blood lead
concentration:
adult male (n = 39): a= 1.41, p = 0.014, r = 0.74, p< 0.001
adult female (n = 36): a = 1.23, p = 0.027, r = 0.81, p < 0.001
Linear regression for ALA (log-transformed) and blood lead
concentration:
adult male (n = 39): a = 0.37, p = 0.006, r = 0.41, p < 0.01
adult female (n = 36): a = 0.15, p = 0.015, r = 0.72, p < 0.001
-------
Table 6-9.2 (cont'd). Effects of Lead on Biochemical Effects in Adults
g Reference, Study
O Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
X
Oi
H
6
o
o
H
O
o
H
W
O
O
HH
H
W
Europe (cont'd)
Grandjean(1979) Design: longitudinal
Denmark Subjects: male battery manufacture workers
NR (n = 19), mean age 32 yr (range 22^9)
Outcome measures: EP
Analysis: EP and blood lead for serial
measurements displayed graphically
Alessioetal. (1976)
Italy
NR
Coccoetal. (1995)
Italy
1990
Design: cross-sectional
Subjects: adult male lead worker (n = 316),
age range NR
Outcome measures: blood ALAD, EP, urine
ALA, CP
Analysis: linear regression, correlation
Design: longitudinal
Subjects: adult male foundry workers
(n = 40), mean age 25.1 yr (SD 2.1, range 21-
28)
Outcome measures: serum total-, HDL- and
LDL-cholesterol, blood Hgb, urine ALA,
erythrocyte G6PD
Analysis: comparison of outcomes between
pre-exposure (at start of employment, sample
1) and after 172 (range 138-217, sample 2)
days
Blood lead (ug/dL) median
(range):
Groupl(n=5): 47.7
(22.8-53.9)
group 2 (n = 5): 37.3
(35.2-53.9)
Blood lead (ug/dL) range:
10-150
Blood lead (ug/dL) mean
(range):
sample 1: 10.0(7-15)
sample 2: 32.7(20-51)
Five subjects (group 1) showed declines in EP with declining
blood lead (33-58 ug/dL) over a 10-month period; 5 subjects
(group 2) showed no change in EP with a change in blood lead
concentration (25-54 ug/dL) over the same period.
Regression relating outcomes to blood lead concentration:
ALAD (In-transformed) (n = 169): a = 3.73, (3 = -0.031, r = 0.871
ALAU (In-transformed) (n = 316): a = 1.25, (3 = 0.014, r = 0.622
UCP (In-transformed) (n = 252): a = 2.18, (3 = 0.34, r = 0.670
EP (log-transformed (males, n = 95): a = 0.94, p = 0.0117
EP (log-transformed (females, n = 93): a = 1.60, p = 0.0143
G6PD levels were unrelated to starting blood lead; however, they
increased in subjects whose blood lead concentration increased
from <30 ug/dL to >30 ug/dL or decreased from >30 ug/dL to
<30 ug/dL. Increasing exposure duration was significantly
associated with decreasing magnitude of change in G6PD (sample
1 <30 ug/dL: p = -0.3980, SE 0.1761, p < 0.05; sample
1 >30 ug/dL:
P = -1.3148, SE 0.3472, p < 0.05) and, in the >30 ug/dL subgroup,
increasing blood lead was associated with decreasing magnitude of
change of G6PD (P = -2.0797, SE 0.7173, p < 0.05).
Serum cholesterol levels were unrelated to blood lead
concentration.
-------
Table 6-9.2 (cont'd). Effects of Lead on Biochemical Effects in Adults
g Reference, Study
O Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
X
Oi
to
o
o
Europe (cont'd)
Fracaso et al. (2002)
Italy
NR
Hemberg et al. (1970)
Poland
NR
Design: cross-sectional cohort
Subjects: adult battery manufacture workers
(n = 37, 6 females), mean age 41 yr (SD 7);
reference office workers (n = 29, 8 females),
meanage38yr(SD21)
Outcome measures: lymphocyte DNA strand
breaks, ROS, GSH
Analysis: comparison of outcome measures
between lead workers and reference group
(ANOVA), logistic regression
Design: cross-sectional
Subjects: adult lead workers (n = 166);
reference group (n = 16)
Outcome measures: blood ALAD
Analysis: regression, correlation
Blood lead (ug/dL) mean
(SD):
lead: 39.6(7.6)
4.4 (8.6)
Blood lead (ug/dL) range:
5-95
Covariate-adjusted DNA strand breaks were significantly higher in
lead workers compared to the reference group and significantly
associated with increased blood lead (p = 0.011).
Covariate-adjusted lymphocyte ROS was significantly higher and
GSH significantly lower in the lead workers compared to the
reference group. Lower GSH levels were significantly associated
with increasing blood lead concentration (p = 0.006).
Odds ratios (OR) for DNA strand breaks and lower GSH levels
were significant (lead workers vs. reference):
DNA strand breaks: OR = 1.069 (95% CI: 1.020-1.120,
p = 0.005)
GSH: OR = 0.634 (95% CI: 0.488-0.824, p = 0.001)
ROS: OR = 1.430 (95% CI: 0.787-2.596, p = 0.855)
Covariates retained: age, alcohol consumption and tobacco
smoking.
Linear regression for blood ALAD (log-transformed) and blood
lead concentration (n = 158):
a = 2.274, p = -0.018, r = -0.90, p < 0.001
H
6
o
o
H
O
o
H
W
O
O
HH
H
W
Bergdahl et al. (1997)
Sweden
NR
Selander and Cramer
(1970)
Sweden
NR
Design: cross-sectional
Subjects: adult smelter worker (n = 89);
reference groups (n = 24)
Outcome measures: blood lead, erythrocyte
ALAD-bound lead, ALAD genotype
Analysis: comparison of outcome measures
Design: cross-sectional
Subjects: adult battery manufacture workers
(n=177)
Outcome measures: urine ALA
Analysis: regression, correlation
Blood lead (ug/dL):
range 0.8-93
Urine lead (mg/L):
range 1-112
Bone lead (ug/g)
range-19-101
Blood lead (ug/dL) range:
6-90
No association between ALAD genotype and lead measures.
Linear regression for urine ALA (log-transformed) and blood lead
concentration (n = 150):
a = -1.0985, p = 0.0157, r = 0.74
-------
Table 6-9.2 (cont'd). Effects of Lead on Biochemical Effects in Adults
g Reference, Study
O Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
Europe (cont'd)
Wildtetal. (1987)
Sweden
NR
Design: longitudinal
Subjects: adult battery manufacture workers
(n = 234, 37 females) mean age 35 y (range
17-70); reference group (n = 951, 471
females), mean age 39 yr (range 19-67)
Outcome measures: EP
Analysis: analysis of variability over time,
linear regression, correlation
Blood lead (ug/dL) mean
(range):
lead: 10-80
reference:
male: 11.3(8-27)
female: 8.5(5-21)
Linear regression for EP (log-transformed) and blood lead
concentration:
males (n= 851): a= 1.21, (3 = 0.0148, r = 0.72
females (n= 139): a= 1.48, (3 = 0.0113, r = 0.56
Asia
X
Oi
to
o
Hsieh et al. (2000)
China
NR
H
6
o
o
H
O
o
H
W
O
O
HH
H
W
Jiun and Hsien( 1994)
China
1992
Froometal. (1999)
Israel
1980-1993
Design: cross-sectional
Subjects: Adults in general population
(n = 630, 255 females)
Outcome measures: blood Hgb, Hct, RBC
count, ALAD genotype
Analysis: comparison of outcome measures
between ALAD genotype strata
Design: longitudinal
Subjects: adult male lead workers (n = 62),
ages NR; reference group (n = 62, 40 females),
agesNR
Outcome measures: plasma MDA
Analysis: comparison of outcome measures
between lead workers and reference group,
linear regression
Design: longitudinal survey
Subjects: adult male battery manufacturing
workers (n = 94), mean age, 38 yr (SD 9,
range 26-60)
Outcome measures: blood Hgb, blood EP
Analysis: multivariate linear regression
Blood lead (ug/dL) mean
(SD):
ALAD l,l(n= 630):
6.5(5.0)
ALAD 1,1/2,2 (n = 30)
7.8 (6.0)
Blood lead (ug/dL) mean
(SD, range):
lead: 37.2
(12.5, 18.2-76.0)
reference: 13.4
(7.5,4.8^3.9)
Blood lead (ug/dL) range of
13-yr individual subject
means
20-61 ug/dL
Mean blood lead not different between ALAD genotype strata
(p = 0.17). RBC count, Hgb, Hct not different between ALAD
genotype strata (p = 0.7)
Plasma MDA levels significantly (p < 0.0001) higher
(approximately 2x) in lead workers whose blood lead
concentration 35 ug/dL compared to <30 ug/dL. In subjects with
blood lead >35 ug/dL, blood lead and plasma MDA were
significantly correlated:
blood lead = 9.584(MDA)+24.412 (r = 0.85)
Weak (and probably not significant) covariate-adjusted association
between blood Hgb and individual sample blood lead
(P = -0.0039, SE 0.0002), subject average blood lead
(P = -0.0027, SE 0.0036), or blood EP (P = "0.001, SE 0.0007)
Covariates retained in model were age and smoking habits.
-------
Table 6-9.2 (cont'd). Effects of Lead on Biochemical Effects in Adults
g Reference, Study
O Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
X
Oi
to
o
to
H
6
o
o
H
O
o
H
W
O
O
HH
H
W
Asia (cont'd)
Kristal-Boneh et al.
(1999)
Israel
1994-1995
Solliway et al. (1996)
Israel
NR
Itoetal. (1985)
Japan
NR
Design: cross-sectional cohort
Subjects: adult male battery manufacture
workers (n = 56), mean age 43.1 yr (SD 10.6);
reference group (n = 87), mean age 43.2 yr
(SD 8.3)
Outcome measures: serum total-, HDL-, LDL-
cholesterol, HDL:total ratio, triglycerides
Analysis: comparison of outcome measures
between lead workers and reference group
(ANOVA), multivariate linear regression
Design: cross-sectional cohort
Subjects: adult male battery manufacture
workers (n = 34), mean age: 44 yr (SD 13);
reference subjects (n = 56), mean age 43 yr
(SD 12); cohorts constructed to have similar
age, ethnic characteristics, socioeconomic
status, education level, and occupation
Outcome measures: urinary ALA, erythrocyte
GSH-peroxidase
Analysis: parametric comparison of outcome
measures between lead and reference groups,
correlation
Design: cross-sectional cohort
Subjects: adult male steel (smelting, casting)
workers (n = 712), age range 18-59 yr;
reference (office workers) group (n = 155,
total), age range 40-59 yr
Outcome measures: serum LPO and SOD,
total and HDL-cholesterol, phospholipid
Analysis: comparison of outcome measures
between lead workers and reference group,
correlation
Blood lead (jig/dL) mean
(SD):
lead: 42.3(14.9)
reference: 2.7(3.6)
Blood lead (ng/dL) mean
(SD, range):
lead: 40.7(9.8,23-63)
reference: 6.7(2.4,1-13)
Blood lead (|ig/dL) range:
lead: 5-62
reference: NR
Covariate-adjusted serum total-cholesterol (p = 0.016) and HDL-
cholesterol (p = 0.001) levels were significantly higher in lead
workers compared to reference group. Covariates retained in
ANOVA: age, body mass index, season of sampling, nutritional
variables (dietary fat, cholesterol, calcium intakes), sport activities,
alcohol consumption, cigarette smoking, education, job seniority.
Increasing blood lead concentration was significantly associated
with covariate-adjusted total cholesterol (P = 0.130, SE 0.054,
p = 0.017) and HDL-cholesterol (P = 0.543, SE 0.173, p = 0.002).
Covariates retained: age, body mass index. Stepwise inclusion of
other potential confounders had no effect.
Significantly lower mean erythrocyte GSH-peroxidase activity
(p < 0.005) in and higher urinary ALA (p < 0.001) in lead workers
compared to reference group.
When stratified by age, significantly (p < 0.05) higher serum HDL-
cholesterol and LPO in lead workers, age range 40^9 yr,
compared to corresponding strata of reference group. Serum
lipoperoxide levels increased as blood lead increased above 30
Hg/dL (p = NR), SOD appeared to decrease with increasing blood
lead concentration (p = NR)
-------
Table 6-9.2 (cont'd). Effects of Lead on Biochemical Effects in Adults
g Reference, Study
O Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
Asia (cont'd)
Makinoetal. (1997)
Japan
1990-1994
Design: longitudinal survey
Subjects: adult male pigment or vinyl chloride
stabilizer manufacture workers (n = 1573)
mean age 45 yr
Outcome measures: blood Hgb, Hct, RBC
count
Analysis: parametric comparison of outcome
measures, stratified by blood lead, linear
regression
Blood lead (ug/dL) mean
(SD, range):
12.6 (2.0, 1-39)
Urine lead (ug/L) mean
(SD, range):
10.2 (2.7, 1-239)
Significantly higher (p < 0.001) Hct, blood Hgb and RBC count in
blood lead category 16-39 ug/dL, compared to 1-15 ug/dL
category.
Significant positive correlation between blood lead concentration
and Hct: a = 42.95, p = 0.0586 (r = 0.1553, p < 0.001), blood
Hgb: a= 14.65, p = 0.0265 (r = 0.1835, p< 0.001) andRBC
count a = 457, p = 0.7120 (r = 0.1408, p < 0.001).
X
Oi
to
o
Moritaetal. (1997)
Japan
NR
Design: cross-sectional cohort
Subjects: male lead workers (n = 76), mean
age 42 yr (range 21-62); reference subjects
(n = 13, 6 females), mean age, males 41 yr
(range 26-52), females 45 yr (range 16-61)
Outcome measures: blood NADS, ALAD
Analysis: comparison of outcome measures
(ANOVA) between blood lead categories,
linear regression
Blood lead (ug/dL) mean
(SD, range)
lead: 34.6(20.7,2.2-81.6)
Significantly lower (p < 0.01) blood NADS and ALAD in blood
lead categories >20 ug/dL compared to <20 ug/dL, with dose trend
in magnitude of difference.
Significant associations between increasing blood lead and
decreasing blood NADS and ALAD in lead workers:
NADS: a = 0.843, p = -0.00971, r = -0.867, p < 0.001, n = 76
logALAD: a = 1.8535, p = -0.015, r = -0.916, p < 0.001, n = 58
H
6
o
o
H
O
o
H
W
O
O
HH
H
W
Oishietal. (1996)
Japan
NR
Design: cross-sectional
Subjects: adult glass and pigment manufacture
workers (n = 418, 165 females), mean age 33
yr (range 18-58); reference workers (n = 227,
89 females), mean age 30 yr (range 17-59)
Outcome measures: plasma ALA, urinary
ALA
Analysis: linear regression, correlation
Blood lead (ug/dL) mean
(SD, range):
lead: 48.5 (17.0,10.3-99.4
reference: 9.6 (3.3, 3.8-
20.4)
Significant correlation between blood lead concentration and
plasma and urinary ALA (both log-transformed):
plasma ALA: a = 0.327, p = 0.022, r= 0.742
urinary ALA: a = -0.387, p = 0.022, r = 0.711
Significant correlation between plasma and urinary ALA:
a = 6.038, P = 4.962, r= 0.897
-------
Table 6-9.2 (cont'd). Effects of Lead on Biochemical Effects in Adults
to
o
o
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
X
Oi
to
o
H
6
o
o
H
O
O
H
W
O
O
HH
H
W
Asia (cont'd)
Sugawaraetal. (1991)
Japan
NR
Kim et al. (2002)
Korea
1996
Lee et al. (2000)
Korea
NR
Design: cross-sectional cohort
Subjects: adult lead workers and reference
group (n = 32, total), ages NR
Outcome measures: plasma and erythrocyte
lipoperoxide and SOD; erythrocyte CAT,
GSH, and methemoglobin
Analysis: comparisons of outcome measures
between lead workers and reference group,
linear regression and correlation
Design: cross-sectional cohort
Subjects: adult male secondary lead smelter
workers (n = 83), mean age: 38.7 yr (SD
10.8); reference subjects (n = 24), mean age:
32.0 (SD 10.8)
Outcome measures: blood Hgb, blood ALAD,
blood EP, blood P5N
Analysis: parametric comparison (ANOVA)
of outcome measures between lead workers
and reference group, correlation, multivariate
linear regression
Design: cross-sectional cohort
Subjects: adult male lead workers (n = 95;
secondary smelter, PVC-stabilizer
manufacture, battery manufacture); mean age
42.8 yr (SD 9.3, range 19-64); reference group
(n = 13), mean age 35.1 yr (SD 9.9, range 22-
54)
Outcome measures: urinary ALA, EP
Analysis: correlation
Blood lead (ug/dL) mean
(SD, range):
lead: 57.1(17.6,20-96)
reference: NR
Blood lead (ug/dL) mean
(SD)
lead: 52.4(17.7)
reference: 6.2 (2.8)
Blood lead (ug/dL) mean
(SD, range):
lead: 44.6(12.6,21.4-
78.4)
reference: 5.9 (1.2, 4.0-
7.2)
Significantly (p < 0.01) higher erythrocyte LPO and lower SOD,
CAT and GSH levels in workers compared to reference group.
Erythrocyte lipoperoxide (r = 0.656) and GSH (r = -0.631) were
significantly correlated with blood lead.
Significantly (p < 0.05) lower blood P5N, ALAD, and Hgb; and
higher blood EP in lead workers compared to controls.
Significant (p < 0.001) correlations (in lead worker group) with
blood lead: P5N (r = -0.704), log EP (r = 0.678), log ALAD
(r = -0.622).
Significant association between increasing EP and decreasing
blood Hgb:
blood lead >60 ug/dL: p = -1.546 (95% CI: -2.387 to -0.704,
r2 = 0.513, p = 0.001)
blood lead <60 ug/dL:
r2 = 0. 177, p = 0.003)
Significant association between increasing P5N and increasing
blood Hgb (high blood lead group only):
blood lead > 60 ug/dL: p = 0.222 (95% CI: 0.015 to 0.419,
r2 = 0.513, p = 0.036)
Covariates included in model: P5N, log serum ferritin, log EP
Significant correlation between increasing DMSA-provoked
urinary lead and urinary ALA (r = 0.3 1 , p < 0.002) and EP
(r = 0. 35, p< 0.001).
= -1.036 (95% CI: -1.712 to -0.361,
-------
Table 6-9.2 (cont'd). Effects of Lead on Biochemical Effects in Adults
to Reference, Study
§ Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
X
Oi
to
o
Asia (cont'd)
Schwartz etal. (1997)
Korea
1994-1995
Gurer-Orhan et al.
(2004)
Turkey
NR
Design: cross-sectional
Subjects: adult male battery manufacture
workers (n = 57), mean age 32 yrs (SD 6).
Outcome measures: blood Hgb, HgbA1, and
HgbA2, ALAD genotype
Analysis: comparison of outcome measures
between ALAD genotype strata
Design: cross-sectional cohort
Subjects: adult male battery manufacture
workers (n = 20), mean age 35 yr (SD 8);
reference workers (n = 16), mean age 32 yr
(SD9)
Outcome measures: blood ALAD, EP,
erythrocyte MDA, CAT, G6PD, blood
GSH: GSSG
Analysis: comparison of outcome measures
between lead workers and reference group,
correlation
Blood lead (ug/dL) mean
(SD):
ALADl,l(n=38): 26.1
(9.8)
ALAD12(n= 19): 24.0
(11.3)
Blood lead (ug/dL) mean
(SD):
lead: 54.6(17)
reference: 11.8(3.2)
Mean blood lead (p = 0.48) and blood Hgb levels (p = 0.34) were
not different between ALAD genotype strata.
Significant correlation between blood lead concentration and blood
ALAD (r = -0.85, p < 0.0001) and EP (r = 0.83, p < 0.001).
Significant correlation between blood lead concentration and
erythrocyte MDA (r = 0.80, p = <0.0001), erythrocyte G6PD
(r = 0.70, p < 0.0001, erythrocyte CAT (r = 0.62, p < 0.001), blood
GSH (r = 0.64, p < 0.0005), blood GSSG (r = 0.67, p < 0.0001).
GSH: GSSG ratio lower (p = NR) in lead workers (3.2), compared
to controls (8.0).
H
6
o
o
H
O
o
H
W
O
O
HH
H
W
Suzenetal. (2003)
Turkey
NR
Design: cross-sectional
Subjects: Male lead battery manufacture
workers (n = 72), age range 24-45 yrs.
Outcome measures: blood ALAD, urine ALA,
ALAD genotype
Analysis: comparison of outcome measures
between ALAD genotype strata
Blood lead (ug/dL) mean
(SD, range):
All: 34.5(12.8,13.4-71.8
ALADl,l(n=51)
34.4(13.1,13.4-71.8)
ALAD2(n = 21)
34.9(12.6,19.2-69.6)
Mean blood lead concentration (p = 0.88) and blood ALAD
activity (p = 0.33) were not different between ALAD genotype
strata. Mean urinary ALA was significantly higher (p < 0.05) in
the ALAD 1-1 stratum.
ALA, 5-aminolevulinic acid; ALAD, 5-aminolevulinic acid dehydratase; ANOVA, analysis of variance; CAT, catalase; CP, coproporphryn; DMSA, dimercaptosuccinic acid;
EP, erythrocyte protoporphyrin; G6PD, glucose-6-phosphate dehydrogenase; GSH, reduced glutathione; GSSG, glutathione disulfide; Hgb, blood hemoglobin; Hct, hematocrit;
HDL, high-density lipoprotein; LDH, lactate dehydrogenase; LPO, lipoperoxide; MDA, malondialdehyde; NADS, adenine dinucleotide synthetase; OR, odds ratio; P5N,
erythrocyte pyrymidine-5'nucleotidase; R/ALAD, ratio of ALAD activity, before and after reactivation; RBC, red blood cells; ROS reactive oxygen species; SD, standard
deviation; SE, standard estimation; SOD, superoxide dismutase; UCP, urinary coproporphyrin
-------
Table AX6-9.3. Effects of Lead on Hematopoietic System in Children
to
o
ON
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
United States
Liebeltetal. (1999)
Connecticut
NR
Design: cross-sectional
Subjects: children (n = 86, 31 female), ages
1-6 yr
Outcome measures: serum EPO, blood Hgb
Analysis: ANOVA of outcome measures
stratified by blood lead, linear regression
Blood lead (ng/dL) median
(range):
18(2-84)
84% <35
Significant association between increasing blood lead concentration
and decreasing serum EPO concentration (P = -0.03, p = 0.02).
Covariates included in model were blood Hgb (P = -1.36, p < 0.01)
(age was not included), R2 = 0.224. Predicted decrease in serum
EPO per 10 |ig/dL was 0.03 mlU/mL. No significant association
between blood lead and blood Hgb.
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O
O
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Schwartz etal. (1990)
Idaho
1974
Design: cross-sectional
Subjects: children (n = 579), ages 1-5 yr,
residing near an active smelter (with
uncontrolled emissions)
Outcome measures: Hct
Analysis: logistic regression
Blood lead (ng/dL) range:
11-164
Significant association between increasing blood lead concentration
and probability of anemia (Hct< 35%) (fa: 0.3083, SE 0.0061) and
age(p2: -0.3831, SE 0.1134). A 10% probability of anemia was
predicted to be associated with blood lead concentration of
approximately 20 ng/dL at age 1 yr, 50 ng/dL at age 3 yr, and
75 ng/dL at age 5 yrs (from Fig. 2 Schwartz et al. (1990).
Regression model relating Hct to blood lead (BL jig/dL) and age
(AGE, yr): Hct = A/(l+exp(p0+PiBL+p2AGE)):
A = 39.42 (SE 0.79, p = 0.0001)
Po = -3.112 (SE 0.446, p = 0.0001)
P! = 0.0133 (SE 0.0041, p = 0.0005)
P2 = -0.2016 (SE 0.0905, p = 0.0129)
Based on above model, a 10% decrease in hematocrit (from 39.5 to
35.5%) is predicted in association with blood lead concentrations of
85, 115, and 145 ng/dL, at ages 1, 3, and 5 yrs, respectively.
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Table AX6-9.3 (cont'd.). Effects of Lead on Hematopoietic System in Children
to
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Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
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Europe
Graziano et al. (2004)
(Factor Litvak et al.
(1999,1998)
Yugoslavia
1985-1998
Design: prospective
Subjects: children (n = 311; age range:
4.5-12 yr) from high-lead (smelter/refinery)
and low-lead areas
Outcome measures: blood Hgb, serum EPO.
Analysis: multivariate linear regression
(GEE for repeated measures)
Blood lead (|ig/dL) range:
4.5 yr: 4.6-73.1
6.Syr: 3.1-71.7
9.0 yr: 2.3-58.1
Blood lead (jig/dL) means
for ages 4.5 - 12 yrs:
high lead: 30.6-39.3
low lead: 6.1-9.0
Significant association between increasing blood lead concentration
and increasing serum EPO concentration at ages 4.5 (p < 0.0001)
and 6.5 yr (p < 0.0007), with decreasing regression slope with age:
4.5 yr: p = 0.21 (SE 0.043, p = 0.0001); 6.5 yr: p = 0.11 (SE 0.41,
p = 0.0103); 9.5 yr: p = 0.029 (SE 0.033, p = 0.39); 12 yr: p = 0.016
(SE 0.031, p = 0.60).
Covariates retained in regression model were age (a), blood lead (P),
and blood Hgb (y). GEE for repeated measures yielded (Factor-
Litvak et al. 1998, updated from personal communication from
Graziano 07/2005):
y: 0.6097 (95% CL-0.0915, -0.0479; p < 0.0001)
4.5 yr: a = 1.3421 (95%CI: 1.0348-1.6194, p< 0.0001), p = 0.2142
(0.1282-0.3003, p< 0.0001)
6.5 yr: a= 1.66201.3737-1.9503, p< 0.0001), p = 0.1167 (0.0326-
0.2008, p< 0.001)
9.5 yr: a = 1.7639 (1.4586-2.0691, p< 0.0001), p = 0.0326
(-0.0346-0.0998, p = 0.1645).
12 yr: a = 1.8223 (1.524-2.1121, p < 0.0001), p = 0.0112 (-0.0359-
0.0584, p = 0.1645).
Based on the GEE, the predicted increase in serum EPO per 10
Hg/dL increase in blood lead concentration (at Hgb =13 g/dL) \ was:
1.25 mlU/mL (36%) at age 4.5 yr and 1.18 (18%) at age 6.5 y.
Blood Hgb levels were not significantly different in children from
high-lead area (mean 25-38 ng/dL) compared to low-lead area
(mean: 5-9 ng/dL).
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Table AX6-9.3 (cont'd). Effects of Lead on Hematopoietic System in Children
g Reference, Study
O Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
Latin America
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Perez-Bravo et al. Design: cross-sectional;
(2004) Subjects: children (n = 93, 43 males), age
Chile range: 5-12 yrs who attended school near a
NR powdered lead storage facility
Outcome measures: blood Hgb and Hct,
ALAD genotype
Analysis: comparison of outcome measures
between ALAD genotype strata
Blood lead (|ig/dL) mean
(SE):
ALAD1 (n=84): 13.5
(8.7)
ALAD 2 (n= 9): 19.2(9.5)
Mean blood lead, blood Hgb, and Hct not different between ALAD
genotypes
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EPO, serum erythropoietin; GEE, generalized estimating equation; Hct, hematocrit; Hgb, blood hemoglobin; SE, standard estimation
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Table AX6-9.4. Effects of Lead on Hematopoietic System in Adults
to
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Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
United States
Huetal. (1994) Design: survey
U.S. Subjects: adult male carpentry workers
1991 (n= 119), mean age: 48.6 yr (range: 23-67)
Outcome measures: blood Hct, blood Hgb
Analysis: multivariate linear regression
Blood lead (ug/dL) mean
(SD, range):
8.3 (4.0, 2-25)
Bone lead (ug/g) mean
(SD, range)
tibia: 9.8 (9.5,-15-39)
patella: 13.9 (16.6,-11-78)
Significant association between increasing patella bone lead and
decreasing covariate adjusted blood Hgb ((3 = -0.019, SE 0.0069,
p = 0.008, R2 = 0.078) and blood Hct ((3 = -0.052, SE 0.019,
p = 0.009, R2 = 0.061). After adjustment for bone lead measurement
error, a 37 ug/dL increase in patella bone lead level (from the lowest
to highest quintile) was associated with a decrease in blood Hgb and
Hct of 11 g/L (95% CI: 2.7-19.3 g/L) and 0.03 (95% CI, 0.01 -
0.05), respectively.
Covariates considered: age, body mass index, tibia lead, patella
lead, blood lead, current smoking status, alcohol consumption
Covariates retained: patella bone lead, alcohol consumption, body
mass index.
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Europe
Osterode et al. (1999)
Austria
NR
Design: cross-sectional cohort
Subjects: adult male lead workers (n = 20),
ages 46 yr (SD, 7); age-matched reference
group (n = 20)
Outcome measures: blood PCV, blood Hgb,
serum EPO, blood erythroid progenitor
(BFU-E) cell count, blood pluripotent
progenitor (CFU-GEMM) cell count, blood
granulocyte/macrophage progenitor (CFU-
GM) cell count.
Analysis: parametric and nonparametric
comparison of outcomes between lead
workers and reference group; correlation
Blood lead (ug/dL) mean
(range):
lead: 45.5(16-91)
reference: 4.1(3-14)
Urine lead (ug/L) mean
(range):
lead: 46.6(7-108)
reference: 3.7(2-16)
Significantly lower (p < 0.001) BFU-E counts in lead workers who
had blood lead concentrations >60 ug/dL, compared to reference
group. Significant negative correlation between blood lead or urine
lead and CFU-GM and CFU-E. Serum EPO was not correlated with
Hct in lead workers, however, serum EPO increased exponentially
with decrease in Hct in reference group.
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Table AX6-9.4 (cont'd). Effects of Lead on Hematopoietic System in Adults
to
o
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Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
X
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O
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O
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Europe (cont'd)
Gennartetal. (1992)
Belgium
NR
Mohammed-Brahim
etal. (1985)
Belgium
NR
Hajemetal. (1990)
France
NR
Design: cross-sectional cohort
Subjects: adult battery manufacture workers
(n = 98), mean age, 37.7 yr (range: 22-55);
reference group (n = 85), mean age 38.8 yr
(24-55)
Outcome measures: blood Hgb, RBC count,
Hct, blood EP
Analysis: linear regression
Design: cross-sectional cohort
Subjects: adult smelter and ceramics
manufacture workers (n = 38, 13 females);
reference subjects (n = 100) matched with
worker group by age, sex, and
socioeconomic status
Outcome measures: blood P5N, EP, ALAD,
R/ALAD (ratio of ALAD before and after
reactivation).
Analysis: comparison of outcome measures
(ANOVA) between lead workers and
reference group; correlation
Design: cross-sectional
Subjects: adult males (n = 129), mean age
36 yr (SD 7.8, range: 24-55), with no
environmental exposure to lead
Outcome measures: erythrocyte membrane
activities of Na+-K+-ATPase, Na+-K+-co-
transport, Na+-Li+-antiport, and passive Na+
and K+ permeability
Analysis: linear regression, correlation
Blood lead (ug/dL) mean
(SD, range):
lead: 51.0(8.0,40-70)
reference: 20.9(11.1,4.4-
30.0)
Blood lead (ug/dL) mean
(SD, range):
lead: 48.5(9.1,27.8-66.6)
reference: 14.3 (6.7, 5.6-
33.6)
Urine lead (ug/g creatinine)
mean (SD, range):
lead: 84.0(95.9,21.8-587)
reference: 10.5(8.2,1.7-
36.9)
Blood lead (ug/dL)
geometric mean (95% CI
range):
16.0(15.2-16.8,8.0-33.0)
Hair lead (ug/g) geometric
mean (95% CI range):
5.3 (4.44-6.23, 0.9-60)
Significant association between increasing blood lead concentration
and decreasing blood Hgb (P = -0.011, r = 0.22, p = 0.003) or Hct
(P =-0.035, r= 0.24, p< 0.01)
Significant association between increasing blood lead concentration
and increasing blood EP (P = 0.0191, r = 0.87, p = 0.0001)
(No apparent analysis of covariables)
Significantly lower (p = NR) P5N in lead workers (males or females,
or combined) compared to corresponding reference groups.
Correlations with blood lead:
log P5Nr=-0.79 (p< 0.001)
log ALAD r = -0.97 (p = NR)
R/ALAD r = -0.94 (p < 0.001)
logEPr=0.86(p = NR)
Correlations with urine lead:
log P5Nr= -0.74 (p = NR)
log ALAD r = -0.79 (p = NR)
R/ALAD r = -0.84 (p < 0.001)
logEPr=0.80(p = NR)
Na+-K+-co-transport activity negatively correlated with blood lead
concentration (r = -0.23, p = 0.02); linear regression:
a = 583.19, p =-170.70.
Na+-K+-ATPase activity negatively correlated with hair lead
(r = -0.18, p = 0.04); simple linear regression:
a = 3.34, p = -0.02.
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Table AX6-9.4 (cont'd). Effects of Lead on Hematopoietic System in Adults
g Reference, Study
O Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
X
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Europe (cont'd)
Poulosetal. (1986)
Greece
NR
Romeo etal. (1996)
Italy
NR
Graziano etal. (1990)
Yugoslavia
1986
Design: cross-sectional cohort
Subjects: adult male cable production
workers who were exposed to lead (worker
1; n = 50, mean age: 37 yr); male cable
workers who had not direct contact with lead
(worker 2, n = 75, mean age: 36. Syr);
reference group (n = 35, mean age: 39 yr)
Outcome measures: blood Hgb, Hct
Analysis: simple linear regression in the
form: mean Hct = a+p(individual Hct -
group mean Hct)
Design: cross-sectional cohort
Subjects: adult male lead workers (n = 28),
age range, 17-73; reference group (n = 113),
age range, 21-75 yr
Outcome measures: serum EPO, blood Hgb
Analysis: nonparametric comparison of
outcome measures between lead workers and
reference group; correlation
Design: prospective
Subjects: pregnant women (n = 1502) from
high-lead (smelter/refinery) and low-lead
areas
Outcome measures: Hgb
Analysis: comparison of outcome measures
between high-and low-lead groups
Blood lead (|ig/dL) mean
(SE):
worker 1: 27.0(0.7)
worker 2: 18.3(0.6)
reference: 21.5(1.5)
Blood lead (ng/dL) mean
(SD, range):
leadl: 32.3(5.6,30^9)
lead 2: 65.1(16,50-92)
reference: 10.4 (4.3, 3-20)
Blood lead (ng/dL) mean
(95% CI):
high lead: 17.1(6.9^2.6)
low lead: 5.1 (2.5-10.6)
Significant association between increasing blood lead and decreasing
Hct:
worker 1: a = 46.50, (3 = -0.170, SE 0.079, p < 0.05
worker 2: a = 44.57, p = -0.180, SE 0.083, p < 0.05
reference: a = 44.69, p = -0.255, SE 0.044, p < 0.001
Significant association between increasing blood lead and decreasing
blood Hgb:
worker 1: a = 15.23, p = -0.058, SE 0.028, p< 0.05
worker 2: a = 14.58, p = -0.071, SE 0.034, p < 0.05
reference: a = 14.64, p = -0.087, SE 0.015, p < 0.001
Significantly (p = 0.021) lower serum EPO in lead workers
compared to reference group. No significant (p < 0.05) lead effect
on blood Hgb.
Mean blood Hgb levels (g/dL) in high-lead group (12.4; 95% CI:
10.3-14.5) not different from low-lead group (12.3; 95% CI: 10.0-
14.7).
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Table AX6-9.4 (cont'd). Effects of Lead on Hematopoietic System in Adults
g Reference, Study
O Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
Europe (cont'd)
Graziano et al. (1990)
Yugoslavia
1986
Design: prospective
Subjects: pregnant women (n = 48) from
high-lead (smelter/refinery) and low-lead
areas (6 highest and lowest mid-pregnancy
blood lead concentrations), within each of 4
Hgb strata (g/dL): 9.0-9.9, 10.0-10.9,11.0-
11.9,12.0-12.9
Outcome measures: Hgb, EPO
Analysis: ANOVA of outcome measures in
subjects stratified by blood lead and blood
Hgb
Blood lead (ug/dL) mean
range for Hgb strata
high lead: 16.9-38.6
low lead: 2.4-3.6
Significant effect of blood lead (p = 0.049) and blood Hgb
(p = 0.001) on mid-term and term serum EPO (blood lead p = 0.055,
Hgb p = 0.009), with significantly lower serum EPO associated with
higher blood lead.
X
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H
O
o
H
W
O
O
HH
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Asia
Hsiao etal. (2001)
China
1989-1999
Hsieh et al. (2000)
China
NR
Design: longitudinal
Subjects: adult battery manufacture workers
(n = 30, 13 females), mean age 38.3 yr
Outcome measures: blood Hgb, Hct, RBC
count
Analysis: GEE for repeated measures
(models: linear correlation, threshold
change, synchronous change, lag change);
logistic regression
Design: cross-sectional
Subjects: Adults in general population
(n = 630, 255 females)
Outcome measures: blood Hgb, Hct, RBC
count, ALAD genotype
Analysis: comparison of outcome measures
between ALAD genotype strata
Blood lead (ug/dL) mean:
1989: 60
1999: 30
Blood lead ( ug/dL) mean
(SD):
ALAD l,l(n= 630):
6.5(5.0)
ALAD: 1,1/2,2 (n= 30)
7.8 (6.0)
Significant association between increasing blood lead and increasing
RBC count and Hct:
Odds ratios (95% CI):
synchronous change model:
blood Hgb (0.95, 0.52-1.78)
RBC count (3.33, 1.78-6.19)
Hct (2.19, 1.31-3.66)
lag change:
blood Hgb (1.70, 0.99-2.92)
RBC count (2.26, 1.16-4.41)
Hct (2.08, 1.16-4.41)
Mean blood lead not different between ALAD genotype strata
(p = 0.17). RBC count, Hgb, Hct not different between ALAD
genotype strata (p = 0.7)
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Table AX6-9.4 (cont'd). Effects of Lead on Hematopoietic System in Adults
to
o
o
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
X
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o
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H
O
O
H
W
O
O
HH
H
W
Asia (cont'd)
Froometal. (1999)
Israel
1980-1993
Solliwayetal. (1996)
Israel
NR
Horiguchi et al. (1991)
Japan
NR
Design: longitudinal survey
Subjects: adult male battery manufacturing
workers (n = 94), mean age, 38 yr (SD 9,
range: 26-60)
Outcome measures: blood Hgb, blood EP
Analysis: multivariate linear regression
Design: cross-sectional cohort
Subjects: adult male battery manufacture
workers (n = 34), mean age: 44 yr (SD 13);
reference subjects (n = 56), mean age 43 yr
(SD 12); cohorts constructed to have similar
age, ethnic characteristics, socioeconomic
status, education level, and occupation
Outcome measures: blood Hgb, RBC count
Analysis: parametric comparison of
outcome measures between lead and
reference groups, correlation
Design: cross-sectional cohort
Subjects: adult male secondary lead refinery
workers (n = 17), mean age: 44.9 yr (range:
24-58); reference male subjects (n = 13),
mean age: 33.5 yr (range: 22^4)
Outcome measures: RBC deformability
(microfiltration at -20 cm H2O pressure),
RBC count, Hct, blood Hgb
Analysis: comparisons of outcome measures
between lead workers and reference group
Blood lead (ug/dL) range
of 13-yr individual subject
means
20-61 ug/dL
Blood lead (ug/dL) mean
(SD, range):
lead: 40.7(9.8,23-63)
reference: 6.7(2.4,1-13)
Blood mead (ug/dL) mean
(SD):
lead: 53.5(16.1)
reference: NR
Urine lead (ug/L) mean
(SD):
lead: 141.4(38.1)
reference: NR
Week (and probably not significant) covariate-adjusted association
between blood Hgb and individual sample blood lead (P = -0.0039,
SE 0.0002), subject average blood lead (P = -0.0027, SE 0.0036) or
blood EP (P = -0.001, SE 0.0007).
Covariates retained in model were age and smoking habits.
Significantly lower (p < 0.05) mean RBC count in lead workers
compared to reference group. Significant negative correlation
between blood lead concentration and RBC count (r = -0.29,
p < 0.05). Mean comparison for blood Hgb (p = 0.4); correlation
with blood lead concentration (r = -0.05, p = 0.7).
Significantly lower RBC deformability (p < 0.01), RBC count
(p < 0.01) Hct (p < 0.01), and blood Hgb (p > 0.001) in lead workers
compared to reference group.
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Table AX6-9.4 (cont'd). Effects of Lead on Hematopoietic System in Adults
to Reference, Study
§ Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
Asia (cont'd)
Makinoetal. (1997)
Japan
1990-1994
Design: longitudinal survey
Subjects: adult male pigment or vinyl
chloride stabilizer manufacture workers
(n = 1573) mean age 45 yr
Outcome measures: blood Hgb, Hct, RBC
count
Analysis: parametric comparison of
outcome measures, stratified by blood lead,
linear regression
Blood lead (ug/dL) mean
(SD, range):
12.6 (2.0, 1-39)
Urine lead (ug/L) mean
(SD, range):
10.2 (2.7, 1-239)
Significantly higher (p < 0.001) Hct, blood Hgb, and RBC count in
blood lead category 16-39 ug/dL, compared to 1-15 ug/dL
category.
Significant positive correlation between blood lead concentration
and Hct: a = 42.95, p = 0.0586 (r = 0.1553, p < 0.001), blood Hgb:
a = 14.65, p = 0.0265 (r = 0.1835, p < 0.001), and RBC count
a = 457, p = 0.7120 (r = 0.1408, p < 0.001).
Moritaetal. (1997)
Japan
NR
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O
HH
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Kim et al. (2002)
Korea
1996
Design: cross-sectional cohort
Subjects: male lead workers (n = 76), mean
age 42 yr (range: 21-62); reference subjects
(n = 13, 6 females), mean age, males 41 yr
(range: 26-52), females 45 yr (range: 16-
61)
Outcome measures: blood NADS, ALAD
Analysis: comparison of outcome measures
(ANOVA) between blood lead categories,
linear regression
Design: cross-sectional cohort
Subjects: adult male secondary lead smelter
workers (n = 83), mean age: 38.7 yr (SD
10.8); reference subjects (n = 24), mean age:
32.0 (SD 10.8)
Outcome measures: blood Hgb, blood
ALAD, blood EP, blood P5N
Analysis: parametric comparison (ANOVA)
of outcome measures between lead workers
and reference group, correlation, multivariate
linear regression
Blood lead (ug/dL) mean
(SD, range)
lead: 34.6(20.7,2.2-81.6)
Blood lead (ug/dL) mean
(SD)
lead: 52.4(17.7)
reference: 6.2 (2.8)
Significantly lower (p < 0.01) blood NADS and ALAD in blood lead
categories >20 ug/dL compared to <20 ug/dL, with dose trend in
magnitude of difference.
Significant associations between increasing blood lead and
decreasing blood NADS and ALAD in lead workers:
NADS: a = 0.843, p = -0.00971, r = -0.867, p < 0.001, n = 76
logALAD: a = 1.8535, p = -0.015, r = -0.916, p < 0.001, n = 58
Significantly (p < 0.05) lower blood P5N, ALAD, and Hgb; and
higher blood EP in lead workers compared to controls.
Significant (p < 0.001) correlations (in lead worker group) with
blood lead: P5N (r = -0.704), log EP (r = 0.678), log ALAD
(r = -0.622).
Significant association between increasing EP and decreasing blood
Hgb:
blood lead >60 ug/dL: p =-1.546 (96% CI: -2.387 to-0.704,
r2 = 0.513, p = 0.001)
blood lead <60 ug/dL: p =-1.036 (96% CI: -1.712 to-0.361,
r2 = 0.177, p = 0.003)
Significant association between increasing P5N and increasing blood
Hgb (high blood lead group only):
blood lead >60 ug/dL: p = 0.222 (96% CI: 0.015 to 0.419,
r2 = 0.513, p = 0.036)
Covariates included in model: P5N, log serum ferritin, log EP
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Table AX6-9.4 (cont'd). Effects of Lead on Hematopoietic System in Adults
to Reference, Study
§ Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
Asia (cont'd)
Schwartz etal. (1997)
Korea
1994-1995
Design: cross-sectional
Subjects: adult male battery manufacture
workers (n = 57), mean age 32 yrs (SD 6).
Outcome measures: blood Hgb, HgbA1, and
HgbA2, ALAD genotype
Analysis: comparison of outcome measures
between ALAD genotype strata
Blood lead (|ig/dL) mean
(SD):
ALADl,l(n=38): 26.1
(9.8)
ALADl,2(n= 19): 24.0
(11.3)
Mean blood lead (p = 0.48) and blood Hgb levels (p = 0.34) were not
different between ALAD genotype strata.
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ALAD, 8-aminolevulinic acid dehydratase; BFU-E, blood erythroid progenitor; CFU-GM, colony forming unit-granulocyte/macrophage progenitor; CFU-E, colony forming
unit blood-erythroid progenitor; CFU-GEMM, colony forming unit blood-pluripotent progenitor; EP, erythrocyte protoporphyrin; EPO, serum erythropoietin; GEE, generalized
estimation equation; Hgb, blood hemoglobin; Hct, blood hematocrit; NADS, nicotinamide adenine dinucleotide; PCV, packed cell volume; P5N, pyrimidine 5'-nucleotidase;
R/ALAD, ratio of ALAD activity, before and after reactivation; RBC, red blood cells.
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Table AX6-9.5. Effects of Lead on the Endocrine System in Children
to
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Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
United States
Mahaffeyetal. (1982)
Wisconsin, New York
NR
Design: cross-sectional
Subjects: children/adolescents (n = 177),
ages l-16yr
Outcome measures: serum 1,25-OH-D
Analysis: comparison of outcome measures
between age, location and blood lead strata,
linear regression
Blood lead (ng/dL) range:
12-120
Serum 1,25-OH-D levels were significantly (p = 0.05) higher in the
age group 11-16 yr compared to age groups 1-5 or 6-10 yr.
Increasing blood lead (log-transformed) significantly associated with
decreasing serum 1,25-OH-D levels in children 1-5 yr of age
(a = 74.5, p = -34.5, r = -0.884, n = 50)
Dietary calcium: NR
X
ON
to
Rosen etal. (1980)
New York
NR
Design: cross-sectional
Subjects: children (n = 45), ages 1-5 yr
Outcome measures: serum calcium, PTH,
25-OH-D, 1,25-OH-D
Analysis: comparison of outcome measures
between blood lead strata, and before and
after chelation, correlation
Blood lead (ng/dL) mean
(SE, range):
<29(n=15): 18(1,10-
26)
30-59 (n= 18): 47(2,33-
55)
>60 (n = 12): 74 (98, 62-
120)
Significantly higher serum PTH levels and lower 25-OH-D in high-
lead group compared to low-lead group; significantly lower 1,25-
OH-D levels in moderate- and high-lead group compared to low-lead
group. Serum levels of 1,25-OH-D were negatively correlated with
blood lead (high lead: r = -0.71, moderate: r = -0.63, p < 0.01).
After chelation therapy, blood lead decreased and serum 1,25-OH-D
levels increased to levels not significantly different (p > 0.1) from
low-lead group, 25-OH-D levels were unchanged.
Dietary calcium intake (mg/day) mean (SE):
low lead: 800(30)
moderate lead: 780 (25)
high lead: 580(15)
H
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o
H
O
O
H
W
O
O
HH
H
W
Sorrelletal. (1977)
New York
1971-1975
Design: cross-sectional
Subjects: children (124), ages 1-6 yr
Outcome measures: serum calcium,
phosphate, 25-OH-D
Analysis: comparison of outcome measures
between blood lead strata, correlation
Blood lead (ng/dL) mean
(SE):
<29(n = 40): 23(1)
30-59 (n = 35): 48(1)
>60(n = 49): 84(5.0)
Serum calcium and 25-OH-D were significantly lower in high lead
group (p < 0.001). Significant negative correlation between blood
lead and serum calcium (high lead, r = -0.78, p < 0.001) or calcium
intake high lead, (r = -0.82, p < 0.001) in all three lead strata. Serum
25-OH-D was significantly positively correlated with vitamin D
intake, but not with blood lead.
Dietary calcium intake (mg/day) mean (SE):
low lead: 770(20)
moderate lead: 760 (28)
high lead: 610(20)
-------
Table AX6-9.5 (cont'd). Effects of Lead on the Endocrine System in Children
to
o
o
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
United States (cont'd)
X
Oi
to
Siegeletal. (1989)
Connecticut
1987
Design: cross-sectional
Subjects: children (n = 68, 32 female), ages
11 mo to 7 yr
Outcome measures: serum FT4, TT4
Analysis: linear regression
Koo et al. (1991) Design: longitudinal (subset of prospective)
Ohio Subjects: children (n = 105, 56 females), age
NR 21,27, 33 mo
Outcome measures: serum calcium
magnesium, phosphorus, PTH, CAL, 25-OH-
D, 1,25-OH-D, and bone mineral content
Analysis: structural equation modeling
Blood lead (iig/dL) mean
(range):
25 (2-77)
Blood lead (ng/dL)
geometric mean (GSD,
range):
lifetime mean, based on
quarterly measurements:
9.74(1.44,4.8-23.6)
concurrent:
15.01 (1.52,6^4)
maximum observed:
18.53(1.53,6-63)
No significant association between blood lead concentration and
thyroid hormone outcomes. Linear regression parameters:
FT4: a = 1.55 (SE 0.05), (3 = 0.0024 (SE 0.0016), r2 = 0.03, p = 0.13
TT4: a = 8.960 (SE 0.39), p = 0.0210 (SE 0.0127), r2 = 0.04,
p = 0.10
Significant association between increasing blood lead (In-
transformed) and covariate-adjusted decreasing serum phosphorus
(a = 1.83, p = -0.091). No other covariate-adjusted outcomes were
significantly associated with blood lead.
Covariates retained: age, sex, race, and sampling season.
Dietary calcium intake (mg/day)
<600: n = 4(4%)
600-1200: n=58(55%)
>1200: n = 43(41%)
H
6
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H
O
O
H
W
O
O
HH
H
W
CAL, calcitonin; FT4, free thyroxine; GSD, geometric standard deviation; 25-OH-D, 25-hydroxyvitamin D; 1,25-OH-D, 1,25-dihydroxyvitamin D; PTH, parathyroid hormone;
RBP, retinal binding protein; SE, standard estimation; TRH, thyroid releasing hormone; TSH, thyroid stimulating hormone; TT3, total triiodothyronine; TT4, total thyroxine;
TTR, transthyretin
-------
Table AX6-9.6. Effects of Lead on the Endocrine System in Adults
to
o
o
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
United States
Cullenetal. (1984)
Connecticut
1979
NR
Design: clinical case study
Subjects: adult males with neurological
symptoms of lead poisoning
Outcome measures: serum, FSH, LH, PRL,
TES
Analysis: clinical outcomes in terms of
abnormal values
Blood lead (ng/dL) (range):
66-139
Five subjects with defects in spermatogenesis (including
azospermia), with no change in basal serum FSH, LH, PRL, and
TES.
X
Oi
to
oo
H
6
o
o
H
O
O
H
W
O
O
HH
H
W
Robins etal. (1983 Design: cross-sectional
Connecticut Subjects: adult male brass foundry workers
NR (n = 47), age range 20-64 yr
Outcome measures: FT4
Analysis: simple linear regression with
stratification by age and race.
Braunstein et al. Design: clinical
(1978) Subjects: adult male secondary lead smelter
California (n = 12), mean age 38 yr, reference group,
NR (n = 9), mean age 29 yr
Outcome measures: serum EST, FSH, LH,
TES, HCG-stimulated EST and TES, GnRH-
stimulated serum FSH and LH
Analysis: comparisons of outcome measures
between patients symptomatic for lead
poisoning, lead-exposed patients not
symptomatic, reference group
Refowitz (1984) Design: cross-sectional survey
NR Subjects: secondary copper smelter workers
(n=58)
Outcome measures: FT4, TT4
Analysis: linear regression
Blood lead (jig/dL) range:
16-127
Blood lead (jig/dL) mean
(SD):
symptomatic (n = 9):
time of test: 38.7(3.0)
highest: 88.2(4.0)
asymptomatic (n = 4):
time of test: 29.0(5.0)
highest: 80.0(0.0)
reference: 16.1(1.7)
Blood lead (jig/dL) range:
5-60
Significant association between increasing blood lead concentration
and decreasing FT4 (a = 1.22, (3 = -0.0042; 95% CI: -0.0002, -
0.0082; r2 = 0.085, p = 0.048). Significant interaction between race
(black, white) and blood lead. When stratified by race:
black: a = 1.13, p =-0.0051, 95% CI: 0.0007,-0.0095, r2 = 0.21,
p = 0.03)
white: r2 = 0.05, p = 0.27
Strength of association not changed by including age in the
regression model.
Statistically significant (p < 0.05) lower basal serum TES, higher
TES response to HCG, and significantly reduced LH response to
GnRH in workers symptomatic for lead poisoning (including
EDTA-provoked urinary lead >500 ng/24 hr).
No significant association between blood lead and hormone levels:
FT4: a = 2.32, p =-0.0067 (95% CI: -0.18 -+0.0043)
TT4: a = NR, p =-0.28 (95% CI: -0.059 -+0.0002)
No significant association when ratified by race (black, white)
-------
Table AX6-9.6 (cont'd). Effects of Lead on the Endocrine System in Adults
g Reference, Study
O Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
Canada
X
Oi
to
VO
Alexander et al. (1998,
1996a)
British Columbia
1993
Design: cross-sectional
Subjects: adult male primary smelter workers
(n = 152), mean age 40 yr
Outcome measures: serum FSH, LH, TES
Analysis: multivariate linear regression
Schumacher et al. Design: cross-sectional
(1998) Subjects: adult male smelter workers
British Columbia (n = 151) mean age 40 yr (SD 7.2)
1993 Outcome measures: serum FT4, TT4, TSH
Analysis: linear regression, ANOVA
Blood lead (jig/dL) range
(n=81):
5 (DL)-58 (75th %tile: 29)
Semen lead (jig/dL) range:
0.3 (DL)-17.6
Blood lead (ng/dL) mean:
24.1 (n= 151)
<15(n=36)
15-24(n=52)
25-39 (n = 41)
>40 (n = 22)
No significant association between covariate-adjusted blood lead
and hormone levels (p>0.5) or prevalence of abnormal levels.
Significant association between covariate-adjusted increasing
semen lead concentration and decreasing serum TES ((3 = -1.57,
p = 0.004).
Covariates considered: age, smoking, alcohol, other metals in blood
(As, Cd, Cu, Zn), abstinence days prior to sample collection, and
sperm count.
No significant effect of blood lead (categorical) on covariate-
adjusted or unadjusted FT4 (p = 0.68), TT4 (p = 0.13), TSH
(p = 0.54). No significant association of blood lead with prevalence
of abnormal values of hormones. No significant association
between 10-yr average blood lead and hormone levels or prevalence
of abnormal values.
Covariates considered: age and alcohol consumption.
H
6
o
o
H
O
o
H
W
O
O
HH
H
W
Europe
Gennartetal. (1992)
Belgium
NR
Design: cross sectional cohort
Subjects: adult battery manufacture workers
(n = 98), mean age 37.7 yr (SD 8.3, range:
22-55); reference worker group (n = 85),
mean age 38.8 yr (SD 8.7, range: 22-55)
Outcome measures: serum TT3, FT4, TT4,
TSH, FSH, LH
Analysis: comparison of outcome measures
between lead workers and reference group
Blood lead (|ig/dL) mean
(SD, range):
lead: 51.0(8.0,40.0-75.0)
reference: 20.9(11.1,4.4-
39.0)
Urine lead (|ig/g cr) mean
(range):
lead: 57.8(1.95,4.3-399)
reference: 9.75 (2.73,1.45-
77.7)
Mean hormone levels in lead workers and reference group not
different (p = NR); no association between hormone levels and
blood lead or exposure duration quartile.
-------
Table AX6-9.6 (cont'd). Effects of Lead on the Endocrine System in Adults
g Reference, Study
O Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
Europe (cont'd)
Assennato et al. (1987)
Italy
NR
Design: cross-sectional
Subjects: adult male battery manufacture
workers (n = 39), mean age 41 yr (SD 10);
reference cement plant workers (n = 18), mean
age 40 yr (SD 10)
Outcome measures: serum FSH, LH, PRL,
TES; urinary 17-ketosteroids
Analysis: parametric comparison of outcome
measures between lead and reference groups
Blood lead (ug/dL) mean
(SD):
lead: 61 (20)
reference: 18(5)
Urinary lead (ug/L) mean
(SD):
lead: 79(37)
reference: 18(8)
No significant association (p > 0.05) between blood lead and
hormone levels.
X
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to
to
o
Govonietal. (1987)
Italy
NR
Design: cross-sectional
Subjects: adult male pewter manufacture
workers (n = 78), mean age 35 yr (SD 19,
range: 19-52)
Outcome measures: serum PRL
Analysis: parametric comparison of outcome
measures between blood lead and ZPP strata
Blood lead (ug/dL) mean
(SD)/bloodZPP(ug/dL)
mean (SD):
A(n = 22): 28.2(7.1)724.4
(8.7)
B(n=33):
60/3(19.3)7131(107)
C(n= 13):
33.1(6.7)777.0(42.2)
D (n = 8):
49.1(4.2)734.0(4.8)
Significantly (p < 0.02) higher serum PRL in high ZPP strata (B and
C, compared to low ZPP strata A).
H
6
o
o
H
O
o
H
W
O
O
HH
H
W
Rodamilans et al. Design: cross-sectional cohort
(1988) Subjects: adult male lead smelter workers
Spain (n = 23), age range 21^4 yr; reference group
NR (n = 20), age range 20-60 yr.
Outcome measures: serum: FSH, LH, TES,
FTES, SHBG
Analysis: comparison of outcome measures
between exposure duration strata
Blood lead (ug/dL) mean
(SD)
lead5yr(n= 10): 76(11)
reference (n = 20): 17.2
(13)
Serum TES (p = 0.01) and FTES (p = 0.001) significantly lower and
SHBG significantly higher (p < 0.025) in >5-yr exposure group
compared to reference group; serum LH was significantly (p < 0.01)
higher in all exposure groups compared to reference group.
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Table AX6-9.6 (cont'd). Effects of Lead on the Endocrine System in Adults
g Reference, Study
O Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
X
Oi
to
to
Europe (cont'd)
Erfurthetal. (2001)
Sweden
NR
Design: cross-sectional cohort
Subjects: adult male active secondary smelter
workers (n = 62), mean age 43 yr (range: 21-
78) reference worker group (: 26), mean age
43 yr (range: 23-66)
Outcome measures: serum FT3, FT4, TSH,
TES, SHBG; TRH-stimulated serum TSH;
GnRH-stimulated serum FSH, LH, and PRL
Analysis: nonparametric comparison of
outcome measures between lead workers and
reference group; multivariate linear regression
Blood lead (ng/dL) median
(range):
lead: 31.1(8.3-93.2)
reference: 4.1 (0.8-6.2)
Plasma lead (ng/dL) median
(range:
lead: 31.1(8.3-93.2)
reference: 4.1 (0.8-6.2)
Urine lead (ng/g cr) median
(range):
lead: 19.6(3.1-80.6)
reference: 4.1 (2.4-7.3)
Bone (finger) lead (ng/g)
median (range):
lead: 25 (-13-99)
reference: 2 (-21-14)
Basal hormone levels in workers not different from reference group
(p>0.05); age-adjusted basal hormone levels not associated with
plasma lead, blood lead, urine lead, or bone lead. In an age-
matched subset of the cohorts (n = 9 lead workers, n = 11
reference), median GnRH-stimulated serum FSH was significantly
(p = 0.014) lower (77IU/L x hr) in lead workers than in reference
group (162 IU/L x hr). No association between stimulated TSH,
LH, FSH or PRL and lead measures.
H
6
o
o
H
O
o
H
W
O
O
HH
H
W
Gustafsonetal. (1989)
Sweden
NR
Campbell etal. (1985)
UK
NR
Design: cross-sectional cohort
Subjects: adult male secondary smelter
workers (n = 21) mean age 36.0 yr (SD 10.4);
individually matched for age, sex, and work
shift (n = 21),
Outcome measures: serum FTES, TTES;
FSH, LH, PRL, COR, TSH, TT3, TT4
Analysis: nonparametric comparison of
outcome measures between lead workers and
reference group, correlation
Design: cross-sectional cohort
Subjects: adult male welders (n = 25);
reference subjects (n = 8) (ages NR)
Outcome measures: plasma ACE, AI, PRA,
plasma ALD
Analysis: linear regression, nonlinear least
squares
Blood lead (ng/dL) mean
(SE):
lead: 39.4(2.1)
reference: 5.0 (0.2)
Blood lead (ng/dL) mean
(SD, range):
35.6(15.3,8-62)
Significantly higher TT4 (p < 0.02) and lower serum FSH
(p = 0.009) in lead workers compared to reference group. When
restricted to the age range <40 yr, lead workers had significantly
higher TT4 (p = 0.01) and lower FSH (p = 0.03), LH (p = 0.04), and
COR (p = 0.04), compared to the reference group.
Significant positive correlation between blood lead concentration
and plasma ALD level (r = 0.53, p < 0.002), PRA (r = -0.76,
p < 0.001), AI (r = 0.68, p < 0.002), and ACE (r = 0.74, p < 0.001).
-------
Table AX6-9.6 (cont'd). Effects of Lead on the Endocrine System in Adults
g Reference, Study
O Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
X
Oi
to
to
to
H
6
o
o
H
O
o
H
W
O
O
HH
H
W
Europe (cont'd)
Chalkleyetal. (1998)
UK
1979-1984
Mason etal. (1990)
UK
NR
McGregor and Mason,
(1990)
UK
NR
Design: cross-sectional
Subjects: adult male primary metal (Cd, Pb,
Zn) workers (n = 19), ages NR
Outcome measures: blood calcium, serum 25-
OH-D, 1,25-OH-D, 24,25-OH-D
Analysis: comparison of outcome measures
(ANOVA) in group stratified by blood lead
and urinary cadmium
Design: cross-sectional
Subjects: adult male lead workers (n = 63),
age range 21-63 yr; reference male subjects
(n = 75), age range 22-64 yr
Outcome measures: serum calcium
phosphate, PTH, 1,25-OH-D
Analysis: comparison of al outcome measures
between lead workers and reference group,
multivariate regression
Design: cross-sectional cohort
Subjects: adult male lead workers (n = 90),
mean age (31.5 yr (SD 11.9); reference
workers (n = 86), mean age 40.6 yr (SD 11.8
Outcome measures: serum FSH, LH, TES,
SHBG
Analysis: comparison of outcome means
between lead workers and reference groups,
multivariate regression, correlation
Blood lead (ug/dL) mean
(SD, range):
47 (21-76)
Blood lead (ug/dL) range:
lead (15-94)
reference: NR
Tibia lead (ug/g)
lead: 0-93
reference: NR
Blood lead (ug/dL) range:
lead: 17-77
reference: <12
After stratification by blood lead and urinary cadmium, serum 1,25-
OH-D levels in strata were significantly different (p = 0.006), with
higher mean values in high blood lead (>40ug/dL)/high blood
cadmium (>0.9 ug/L)/high urine cadmium >3.1 ug/L) stratum
compared to low blood lead (<40ug/dL)/high blood cadmium (>0.9
ug/L)/high urine cadmium >3.1 ug/L) stratum. Serum 24,25-OH-D
levels decreased with increasing urinary cadmium (p = NR)
Significantly higher (p < 0.025) prevalence of elevated 1,25-OH-D
(>2 SD of reference mean) in lead workers (8/63, 13%) compared to
reference group (1/75, 1.3%). Serum levels of 1,25-OH-D
significantly (p < 0.05) higher in lead workers compared to
reference group.
After stratification of lead workers into exposure categories (high:
blood lead >40 ug/dL and bone lead >40 ug/g, low: blood lead <40
ug/dL and bone lead <40 ug/g), serum 1,25-OH-D levels were
significantly (p < 0.01) higher in the high lead group.
Increasing blood lead was significantly (p = NR) associated with
increasing 1,25-OH-D levels (r2 = 0.206; with age and bone lead
included, r2 = 0.218). After excluding 12 subjects whose blood lead
concentrations >60 ug/dL, r2 = 0.162 (p = 0.26).
Age-adjusted serum FSH was significantly (p = 0.004) higher in
lead workers compared to reference group.
Increasing serum FSH significantly (p = NR) associated with blood
lead and age. Increasing serum LH significantly associated with
increasing exposure duration (not blood lead or age).
No significant association between serum TES or SHBG and blood
lead or exposure duration.
No significant difference in prevalence of abnormal hormone levels
between groups.
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Table AX6-9.6 (cont'd). Effects of Lead on the Endocrine System in Adults
g Reference, Study
O Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
Latin America
Lopez et al. (2000)
Argentina
NR
Design: cross sectional
Subjects: adult male battery manufacture
workers (n = 75), age range 21-56 yr;
reference group (n = 62), age NR
Outcome measures: serum TT3, FT4, TT4,
TSH
Analysis: comparison of outcome measures
between lead workers and reference group,
correlation
Blood lead (ug/dL) mean
(range):
lead: 50.9(23.3,8-98)
reference: 19.1(7.1,4-39)
Significantly higher serum FT4 (p < 0.01) and TT4 (p < 0.05) in
lead workers compared to reference group. Significant positive
correlation between blood lead and serum TT3 (p < 0.05), FT4
(p < 0.01), TT4 (p < 0.05), and TSH (p < .05), for blood lead range
8-50 ug/dL; and for TSH (p < 0.05) for blood lead range 8-
26 ug/dL.
X
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to
to
Roses etal. (1989)
Brazil
NR
Design: adult male lead workers (n = 70), age
range 20-53 yr; reference group (n = 58), age
range 25-37 yr.
Outcome measures: serum PRL
Analysis: comparison of outcome measure
between lead workers and reference group,
linear regression
Blood lead (ug/dL) (range):
lead: 9-86
reference: 8-28
Serum RL levels in lead workers and reference group not
significantly different (p = NR). Correlation between serum PRL
and blood lead (r = 0.57, p = NR).
Asia
H
6
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o
H
O
o
H
W
O
O
HH
H
W
Dursun and Tutus Design: cross-sectional
(1999) Subjects: adult metal powder manufacture
Turkey workers (n = 27) mean age 41.1 yr (SD 5.45,
NR range: 25-50); reference group (n = 30),
mean age 42 yr(SD 3.42, range: 28^19)
Outcome measures: serum FT4, TT4, FT3,
TT3, TSH
Analysis: parametric comparison of outcome
measures between lead and reference groups,
simple and multivariate linear regression
Blood lead (ug/dL) mean
(range):
lead: 17.1(9.0,6-36)
reference: 2.4(0.1,1^1)
Significantly (p < 0.0001) higher mean TT4, FT4, and FT3 in lead
workers compared to reference group.
Significant association between TT4, age (P = 0.23, p < 0.006), and
exposure duration (P = -0.20, p > 0.01), but not blood lead
(P = 0.00, NR) in linear regression model that included age, blood
lead, and exposure duration (a = 2.76, r2 = 0.3, p = 0.03).
-------
Table AX6-9.6 (cont'd). Effects of Lead on the Endocrine System in Adults
to
o
o
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
X
Oi
to
to
H
6
o
o
H
O
O
H
W
O
O
HH
H
W
Asia (cont'd)
Kristal-Boneh et al.
(1998)
Israel
NR
Horiguchi et al. (1987)
Japan
NR
Ng et al. (1991)
China
NR
Design: cross-sectional cohort
Subjects: adult male battery
manufacture/recycling workers (n = 56), mean
age 43.4 yr (SD 11.2); reference workers
(n = 90), mean age 41.5 yr (SD 9.3)
Outcome measures: serum calcium,
magnesium, phosphorus, PTH, 25-OH-D,
1,25-OH-D
Analysis: parametric comparison of outcome
measures between lead workers and reference
group, multivariate linear regression
Design: cross-sectional
Subjects: adult secondary lead refinery
(n = 60, 8 females), mean age 49 yr (range:
15-69)
Outcome measures: serum TT3, TT4, TSH
Analysis: comparison of outcome measures
(method NR), between job categories,
correlation
Design: cross-sectional cohort
Subjects: adult male battery manufacture
workers (n = 122), mean age 32.6 (SD 8,2,
range: 17-54); reference group (n = 49),
mean age 43.4 yr (SD 13.4, range: 18-74)
Outcome measures: serum FSH, LH, PRL,
TES
Analysis: multivariate linear regression
ANCOVA
Blood lead (ug/dL) mean
(SD, range):
lead: 42.6(14.5,20-77)
reference: 4.5(2.6,1.4-19)
Blood lead (ug/dL) mean
(SD):
male: 31.9(20.4)
female: 13.5(9.5)
Urine lead (ug/L) mean
(SD):
male: 59.3(76.3)
female: 26.0(19.7)
Blood lead (ug/dL) mean
(SD, range):
lead: 35.2(13.2,9.6-77.4)
reference: 8.3 (2.8, 2.6-
14.8)
Serum 1,25-OH-D (p = 0.0001) and PTH (p = 0.042) were
significantly higher in lead workers compared to reference group
Increasing blood lead concentration (In-transformed) was
significantly associated with covariate-adjusted increasing serum
PTH and 1,25-OH-D levels:
PTH: P = 4.8(95%CI: 0.8-8.8, r2 = 0.12)
1,25-OH-D: (3 = 4.8(95%CI: 2.7-6.9, r2 = 0.10)
Occupational lead exposure (yes) significantly associated with
increasing PTH and 1,25-OH-D levels.
Covariates retained: age, alcohol consumption, smoking; calcium,
magnesium, and calorie intake:
PTH: (3 = 7.81 (95%CI: 3.7-11.5)
1,25-OH-D: p = 12.3 (95% CI: 3.84-20.8)
No significant differences (p = NR) between hormone levels in job
lead categories: mean blood lead (ug/dL, SD): 17.9 (10.7), 25.6
(15.4), 49.9 (18.7). No significant correlations (p = NR) between
hormone levels and blood or urine lead levels.
When cohorts were stratified by age serum FSH and LH were
significantly (p < 0.02) higher in lead workers <40 yrs of age
compared to corresponding age strata of the reference group; serum
TES was significantly (p < 0.01) lower in lead workers >40 yr of
age. Covariate-adjusted serum TES were significantly lower
(p < 0.01) in lead workers in the > 10-yr exposure duration category,
compared to the reference group. Covariate-adjusted serum FSH
and LH were significantly higher (p < 0.01) in lead workers in the
<10-yr exposure duration category, compared to the reference
group.
Covariates: age and tobacco smoking.
-------
Table AX6-9.6 (cont'd). Effects of Lead on the Endocrine System in Adults
g Reference, Study
O Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
X
Oi
to
to
Asia (cont'd)
Zheng etal. (2001)
China
NR
Singh et al. (2000)
India
NR
Design: retrospective cross-sectional
Subjects: adult hospital patients (n = 82, 32
females) mean age 49.6 yr (SD 18.7)
Outcome measures: serum and CSF TTR,
TT4
Analysis: simple and multivariate linear
regression
Design: cross-sectional cohort
Subjects: adult male petrol pump attendants
(n=58), mean age 31.7 yr(SD 10.6);
reference group (n = 35), mean age 28.9 yr
(SD 4.20)
Outcome measures: serum TT3, TT4, TSH
Analysis: parametric comparison of outcome
measures between lead workers and reference
group, stratified by blood lead or exposure
duration
Blood lead (ug/dL) mean
(SD):
all: 14.9(8.3)
female: 14.2(8.76)
male: 15.4(8.07)
Blood lead (ug/dL) mean
(SD):
lead: 51.6(9.3)
reference: 9.5(8.7)
No significant association between blood lead and serum TTR
(r = -0.114, p = 0.307), TT4 (r = -0.160, p = 0.152).
Significant association between age-adjusted CSF lead and CSF
TTR (r = -030, p = 0.023).
No significant association between CSF lead and CSF TT4
(r = -0.22, p = 0.090).
Serum TSH significantly higher (p < 0.01) in lead workers
compared to reference group, significantly higher in high blood lead
category (< 70 ug/dL, mean 54.5 ug/dL) compared to low worker
group <41 ug/dL, mean 31.3 ug/dL). Serum TSH significantly
higher in lead workers who were exposed for <60 mo, compared to
workers exposed for >60 mo.
Africa
H
6
o
o
H
O
o
H
W
O
O
HH
H
W
Tuppurainen et al.
(1988)
Kenya
1984
Design: cross-sectional
Subjects: adult male battery manufacture
workers (n = 176), mean age 34.1 yr
(SD 8.1, range: 21-54)
Outcome measures: serum TT3, FT4,
TT4, TSH
Analysis: multivariate linear regression
and correlation
Blood lead (ug/dL) mean
(SD, range):
55.9(23.8,14.5-133.6)
Increasing exposure duration significantly associated with
decreasing FT4 (r2 = 0.071, p = 0.001) and TT4 (r2 = 0.059,
p = 0.021); regression not improved by including age or blood lead.
Strength of association greater when restricted to workers who had
an exposure duration >7.6 yrs: FT4: r2 = 0.33, p< 0.002; TT4:
r2 = 0.21, p< 0.001.
No significant association between blood lead and hormone levels.
1,25-OH-D, 1,25-dihydroxyvitamin D; 25-OH-D, 25-hydroxyvitamin D; ACE, angiotensin converting enzyme; Al, angiotensin I; ALD, aldosterone; ANOVA, analysis of
variance; CAL, calcitonin; COR, cortisol; cr, creatinine; CSF, cerebral spinal fluid; EDTA, ethylenediaminetetraacetic acid; EST, estradiol; FSH, follicle stimulating hormone;
FT4, free thyroxine; FTES, free testosterone; GnRH, gonadotropin releasing hormone; HCG, human chorionic gonadotropin; LH, luteinizing hormone; NR, not reported; PRL,
prolactin; PTH, parathyroid hormone; RBP, retinal binding protein; SD, standard deviation; SE, standard estimation; SHBG, sex hormone binding globulin; TES, testosterone;
TRH, thyroid releasing hormone; TSH, thyroid stimulating hormone; TT3, total triiodothyronine; TT4, total thyroxine; TTES, total testosterone; TTES, total testosterone;
TTR, transthyretin; ZPP, zinc protoporphyrin
-------
Table AX6-9.7. Effects of Lead on the Hepatic System in Children and Adults
to
o
ON
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
Children
United States
X
ON
to
to
ON
Saengeretal. (1984)
New York
NR
Design: clinical cases
Subjects: children (n = 26) ages 2-9 yr; age-
matched reference group (n = NR)
Outcome measures: urinary cortisol and 6P-
OH-cortisol (CYP3A metabolite of cortisol)
Analysis: comparison of outcome measure
between children who qualified for EDTA
treatment (EDTA provocation >500 jig/24
hr)
Blood lead (|ig/dL) mean
(SE, range):
chelated: 46(2,33-60)
not chelated: 42 (3, 32-60)
Urinary lead (ng/24 hr)
mean (SE, range), EDTA-
provocation:
chelated: 991(132,602-
2247)
not chelated: 298 (32, 169-
476)
Significantly lower (~45% lower) urinary excretion of 6P-OH-
cortisol (p = 0.001) and urinary 6p-OH-cortisol: cortisol ratio
(p < 0.001) in children who qualified for chelation than in children
who did not qualify and significantly lower than age-matched
reference group. Urinary 6p-OH-cortisol: cortisol ratio was
significantly correlated with blood lead (r = -0.514, p < 0.001),
urinary lead, and EDTA provocation urinary lead (r = -0.593,
p< 0.001).
Adults
Asia
H
6
o
o
H
O
O
H
W
O
O
HH
H
W
Al-Neamyetal. (2001)
United Arab Emirates
1999
Design: cross-sectional cohort
Subjects: adult male (n = 100) workers (e.g.,
gas pump attendants, garage workers,
printing workers, construction workers),
mean age 34.6 yr (SD 8.0); reference group
(n = 100) matched with lead workers for age,
sex, nationality.
Outcome measures: serum protein, albumin,
ALT, AP, AST, BUN, yGT, LDH
Analysis: comparison of outcome measures
between lead workers and reference group
Blood lead (ng/dL) mean
(SD):
lead: 77.5(42.8)
reference: 19.8(12.3)
Significantly higher serum AP (p = 0.012) and LDH (p = 0.029) in
lead workers compared to reference group (values within normal
range).
-------
to
o
o
Reference, Study
Location, and Period
Table AX6-9.7 (cont'd). Effects of Lead on the Hepatic System in Children and Adults
Study Description
Lead Measurement
Findings, Interpretation
Adults, Asia (cont'd)
X
Oi
to
to
Hsiao etal. (2001)
China
1989-1999
Satarug et al. (2004)
Thailand
NR
Design: longitudinal
Subjects: adult battery manufacture workers
(n = 30, 13 females), mean age 38.3 yr
Outcome measures: serum ALT
Analysis: GEE for repeated measures
(models: linear correlation, threshold
change, synchronous change, lag change);
logistic regression
Design: cross-sectional
Subjects: adults from general population
(n = 118, 65 female), age range, 21-57 yr
Outcome measures: coumarin-induced
urinary 7-OH-coumarin (marker for
CYP2A6 activity)
Analysis: multivariate linear regression
Blood lead (ug/dL) mean:
1989: 60 (-25-100)
1999: 30 (-10-60)
Urinary lead (ug/g cr)
mean (SD, range):
males: 1.3(1.8,0.1-12)
females: 2.4(1.1,0.6-6.5
Serum lead (ug/L) mean
(SD, range):
males: 4.2 (5.4, 1-28)
females: 3.0 (2.2, 1-12)
No association between blood lead and ALT.
Odds ratios (95% CI):
synchronous change model: 1.25 (0.69-2.25)
lag change: 1.76(0.76^.07)
Significant association between increasing urinary lead and
decreasing covariate-adjusted urinary 7-OH-coumarin (P = -0.29,
p = 0.003) in males, but not in females. Covariates retained: age
and zinc excretion. Significant association in opposite direction
between urinary cadmium and urinary 7-OH-coumarin (P = 0.38,
p = 0.006).
H
6
o
o
H
O
O
H
W
O
O
HH
H
W
yGT, y-glutamyl transferase; 6p-OH-cortisol, 6-p-hydroxycortisol; ALT, alanine aminotransferase; AP, alkaline phosphatase; AST, aspartate aminotransferase; BUN, blood
urea nitrogen; CI, confidence interval; cr, creatinine; EDTA, ethylenediaminetetraacetic acid; GEE, generalized estimating equations; LDH, lactate dehydrogenase; SD,
standard deviation; UAE, United Arab Emirates
-------
Table AX6-9.8. Effects of Lead on the Gastrointestinal System
to
o
o
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
Canada
Holness and Nethercott
(1988)
Ontario
1982-1984
Design: longitudinal
Subjects: adult male demolition workers
(n= 119), age NR
Outcome measures: prevalence of symptoms
Analysis: comparison of prevalence of
symptoms (questionnaire) stratified by job
phase or blood lead
Blood lead (ng/dL) mean
(range):
phase 1: 59(15-99)
phase 2: 30
phase 3: 19
phase 4: 17
Prevalence of reporting of symptoms of abdominal cramps or
constipation increased with increasing blood lead concentration
(p<0.05):
<50 ng/dL: 8%, 6%
50-70 ng/dL: 37%, 42%
>70 ng/dL: 77%, 62%
Caribbean
X
Oi
to
to
oo
Matte etal. (1989)
Jamaica
1987
Design: survey
Subjects: battery manufacture/repair
workers (n = 63), mean age -30 yr (range:
Outcome measures: prevalence of symptoms
Analysis: comparison of GI symptoms
(questionnaire) between blood lead strata
Blood lead (ng/dL)
geometric mean site range:
40-64
Blood lead distribution:
>60: 60%
<60: 40%
When stratified by blood lead, <60 ng/dL (low) or >60 ng/dL (high),
prevalence ratio (high/low) was not significant for abdominal pain
(1.5, 95% CI: 0.5-4.6), or for any other lead symptom (e.g. muscle
weakness).
Asia
H
6
o
o
H
O
O
H
W
O
O
HH
H
W
Bercovitz and Laufer
(1991)
Israel
NR
Design: cross-sectional
Subjects: health individuals (n = 12), peptic
ulcer patients (n = 11), and individuals with
heart disease (n = 11) with environmental
exposure
Analysis: one-way ANOVA used to
compare tooth lead concentrations in the
three groups
Tooth lead (|-ig/g dry
dentine) mean (SE):
Healthy: 25.62(10.15)
Peptic ulcer =75.02 (8.15)
Heart disease: 20.30 (2.70)
Tooth lead levels in patients with gastrointestinal ulcers (n = 11),
were significantly higher than that in healthy subjects (p = 0.001)).
Ten of the 11 peptic ulcer patients had a higher lead level than the
health subjects. In these 10 patients, increased severity of the ulcer
and longevity of suffering was associated with increased tooth lead
levels.
There was no significant difference between the tooth lead levels in
the healthy subjects and in the heart disease patients.
-------
Table AX6-9.8 (cont'd). Effects of Lead on the Gastrointestinal System
to
o
o
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
Asia (cont'd)
X
Oi
to
to
VO
Lee et al. (2000) Design: cross-sectional cohort
Korea Subjects: adult male lead workers (n= 95;
NR secondary smelter, PVC-stabilizer
manufacture, battery manufacture); mean age
42.8 yr (SD 9.3, range: 19-64); reference
group (n = 13), mean age 35.1 yr (SD 9.9,
range: 22-54)
Outcome measures: prevalence of GI
symptoms (self-administered questionnaire)
Analysis: multivariate logistic regression
Blood lead (ug/dL) mean
(SD, range):
lead: 44.6(12.6,21.4-
78.4)
reference: 5.9(1.2,4.0-
7.2)
Covariate-adjusted OR for GI symptoms (loss of appetite,
constipation or diarrhea, abdominal pain) in workers (referents not
included in model) were not significant:
blood lead: 45.7 ug/dL vs. <45.7 ug/dL: OR 1.8 (95% CI: 0.7-4.5)
DMSA-provokedurinary lead: >260.5 vs. <260.5 ug: OR= 1.1
(95% CI: 0.4-2.5)
OR for neuromuscular symptoms were significantly associated with
DMSA-provoked lead (OR = 7.8 (95% CI: 2.8-24.5), but not with
blood lead.
Covariates retained: age, tobacco smoking, and alcohol
consumption.
H
6
o
o
H
O
O
H
W
O
O
HH
H
W
Africa
Awad el Karim et al.
(1986)
Sudan
NR
Design: cross-sectional cohort
Subjects: adult male battery manufacture
workers (n = 92), mean age 31.1 yr (SD 8.2);
reference group (n = 40), mean age 33.7 yr
(SD 9.7)
Outcome measures: clinical evaluation
Analysis: comparison of prevalence of
symptoms of lead poisoning between lead
workers and reference group
Blood lead (ug/dL) mean
(SD, range):
lead: 55-81 (mean range
for various jobs), range:
39-107
Blood lead distribution
>80: 23%
40-80: 72%
<40: 5%
reference: 21 (8.5,7.4-
33.1)
Prevalences of abdominal colic (pain) and constipation were 41.3%
and 41.4 % in lead workers and 7.5% and 10%, respectively, in the
reference group.
DMSA, dimercaptosuccinic acid; GI, gastrointestinal; NR, not reported; OR, odds ratio; PAR, population attributable risk; PVC, polyvinyl chloride; SD, standard deviation; SE,
standard error
-------
Table AX6-9.9. Effects of Lead on the Respiratory Tract in Adults
to
o
o
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
Asia
H
6
o
o
H
O
O
H
W
O
O
HH
H
W
Bagci et al. (2004)
Turkey
NR
Design: cross-sectional cohort
Subjects: adult male battery manufacture and
automobile exhaust repair workers (n = 62),
mean age 32.6 yr; reference hospital workers
(n = 24), mean age 28.8 yr
Outcome measures: VC, FVC, FEVj, PEF,
FEF, MVV
Analysis: comparison of mean outcomes
(ANOVA) between lead workers and
reference group, multivariate (Pearson
partial) correlation
Blood lead (ug/dL) mean
(SD, 95% CI):
battery (n = 22): 36.8(8.1,
33.2-40.3)
exhaust (n = 40): 26.9 (9.2,
24.0-29.9)
reference (n = 24): 14.8
(3.0,13.5-16.1)
Battery manufacture workers had significantly lower FEV
(p < 0.05), FEV: VC ratio (p < 0.05), FEV: FVC ratio (p< 0.01),
FEF (p < 0.01), and MVV (p < 0.01) compared to the hospital
workers. Significant negative (partial) correlation between blood
lead and FEV/FVC (r = -0.31, p = 0.006) and FEF (r = -0.30,
r = 0.009), adjusted for age, cigarette smoking, and exposure
duration.
X
Oi
to
ANOVA, analysis of variance; CI, confidence interval; FEF, forced expiratory flow; FEV, forced expiratory volume; FVC, forced vital capacity; MVV, maximum voluntary
ventilation; PEF, expiratory peak flow; SD, standard deviation; VC, vital capacity
-------
Table AX6-9.10. Effects of Lead on Bone and Teeth in Children and Adults
to
o
o
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
X
Oi
to
H
6
o
o
H
O
O
H
W
O
O
HH
H
W
Children
United States
Moss etal. (1999)
U.S.
1988-1994
Design: cross-sectional national survey
(NHANES III)
Subjects: general population (n = 24,901),
ages 2-5 yr (n = 3,547), 6-11 yr (n = 2,894),
> 12 yr (18,460)
Outcome measures: number of caries (dfs,
DPS, DMFS)
Analysis: multivariate linear regression and
logistic regression
Blood lead (|ig/dL)
geometric mean (SE):
2-5 yr: 2.90 (0.12)
6-1 lyr: 2.07 (0.08)
> 12 yr: 2.49 (0.06)
Increasing blood lead concentration (log-transformed) significantly
associated with covariate adjusted increases in dfs:
2-5 yr: 0 = 1.78 (SE 0.59, p = 0.004)
6-1 lyr: (3= 1.42 (SE 0.51, p = 0.007)
and increases in DFS:
6-11 yr: p = 0.48 (SE 0.22, p = 0.03)
>12yr: p = 2.50 (SE 0.69, p< 0.001)
and increases in DMFS:
> 12 yr: p = 5.48 (SE 1.44, p = 0.01)
Odds ratio (OR) for caries (> 1 DMFS, ages 5-17 yr) and population
attributable risk (PAR) in association with 2nd or 3rd blood lead
tertiles, compared to 1st tertile were:
1st tertile (<1.66 |ig/dL)
2nd tertile (1.66-3.52 |ig/dL): OR 1.36 (95% CI: 1.01-2.83); PAR
9.6%
3rd tertile (>3.52 |ig/dL): OR 1.66 (95% CI: 1.12-2.48); PAR 13.5%
For an increase of blood lead of 5 |ig/dL, OR 1.8 (95% CI: 1.3-2.5)
Covariates retained were age, gender, race/ethnicity, poverty income
ratio, exposure to cigarette smoke, geographic region, educational
level of head of household, carbohydrate and calcium intakes, and
dental visits.
-------
Table AX6-9.10 (cont'd). Effects of Lead on the Gastrointestinal System in Children and Adults
g Reference, Study
O Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
Children, United States (cont'd)
X
Oi
to
OJ
to
H
6
o
o
H
O
o
H
W
O
O
HH
H
W
Schwartz etal. (1986)
U.S.
1976-1980
Gemmel et al. (2002)
Boston/Cambridge, MA
NR
Campbell et al. (2000)
New York
1995-1997
Design: cross-sectional national survey
(NHANES II)
Subjects: ages <7 yr (n = 2,695)
Outcome measures: variables of stature,
including height, weight, and chest
circumference
Analysis: multivariate weighted linear
regression
Blood lead (|ig/dL) range:
5-35
Design: cross-sectional
Subjects: children (n = 543), ages 6-10 yr
Outcome measures: number of caries (dfs,
DPS)
Analysis: multivariate linear regression
Design: retrospective cohort
Subjects: children (n = 154), ages 6.9-12 yr
Outcome measures: prevalence of caries
(dfs, DMFS)
Analysis: multivariate logistic regression
Blood lead (|_ig/dL) mean
(SD, max):
urban (n = 290): 2.9 (2.0,
13)
rural (n= 253): 1.7(1.0,7)
Blood lead (|ig/dL) mean
(range):
10.7(18.0-36.8)
(measured at ages 18 and
37 mo)
Blood lead levels were a statistically significant predictor of
children's height (p < 0.0001), weight (p < 0.001), and chest
circumference (p < 0.026), after controlling for age in months, race,
sex, and nutrition.
Height: p = -0.119 (SE 0.0005)
Weight: p = -1.0217 (SE 0.08) for log-transformed blood lead
Chest circumference: p = -0.6476 (SE 0.077) for log-transformed
blood lead
There are several explanations for the inverse correlation between
blood lead and growth in children. First, blood lead level may be a
composite factor for genetic, ethnic, nutritional, environmental, and
sociocultural factors. Second, nutritional deficits that retard growth
also enhance lead absorption. Finally, there may be a direct effect of
low level lead on growth in children.
Increasing blood lead (In-transformed) was significantly associated
with covariate-adjusted number of caries (dfs + DFS) (In-
transformed) in the urban (P = 0.22, SE 0.08, p = 0.005) group, but
not in the rural group (P = -0.15, SE 0.09, p = 0.09). When dfs
numbers were stratified by permanent or deciduous teeth, the blood
lead association in the urban group was significant for deciduous
teeth
(P = 0.28, SE 0.09, p = 0.002), but not for permanent teeth (P = 0.02,
SE 0.07, p = 0.8).
Covariates retained: age, sex, ethnicity, family income, education of
female guardian, maternal smoking, frequency of tooth brushing,
firmness of toothbrush bristles, and frequency of chewing gum.
Covariate-adjusted odds ratios for caries in association with blood
lead <10 or > 10 |-ig/dL, were:
permanent teeth (DMFS): OR 0.95 (95% CI: 0.43-2.09)
deciduous teeth (dfs): OR 1.77 (95% CI: 0.97-3.24)
Covariates retained: age, grade in school, number of tooth surfaces at
risk. Other covariates explored, that had no effect on strength of
association with blood lead were: sex, ethnicity, and oral hygiene
score.
-------
Table AX6-9.10 (cont'd). Effects of Lead on the Gastrointestinal System in Children and Adults
g Reference, Study
O Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
Adults
United States
X
Oi
to
Dye et al. (2002) Design: cross-sectional national survey
U.S. (NHANES III)
1988-1994 Subjects: adults in general population
(n = 10,033; 5,255 females), ages 20-69 yr
Outcome measures: symptoms of
periodontal bone loss (attachment loss,
periodontal pocket depth)
Analysis: multivariate linear regression
Blood lead (|ig/dL)
geometric mean (SE,
range):
2.5 (0.08) (2.36% > 10)
Increasing blood lead (log-transformed) was significantly associated
with increasing prevalence of covariate-adjusted dental furcation
(P = 0.13, SE 0.05, p = 0.005). Covariates retained: age, sex,
race/ethnicity, education, smoking, and age of home. Smoking
status interaction was significant when included in the model as an
interaction term (P = 0.10, SE 0.05, p = 0.034). When stratified by
smoking status, association between dental furcation and blood lead
was significant for current smokers (P = 0.21, SE 0.07, p = 0.004)
and former smokers (P = 0.17, SE 0.07, p = 0.015), but not for
nonsmokers (P = -0.02, SE 0.07, p = 0.747).
H
6
o
o
H
O
o
H
W
O
O
HH
H
W
Europe
Tvinnereim et al. Design: cross-sectional
(2000) Subjects: 1,271 teeth samples collected by
Norway dentists in all 19 counties in Norway
1990-1994 Analysis: Student's t-test comparing metal
concentrations in teeth with caries, roots, and
in different tooth groups
Tooth lead (|-ig/g tooth)
geometric mean (SD,
range):
1.16(1.72,0.12-18.76)
Also examined mercury, cadmium, and zinc. All tooth groups had
higher lead concentrations in carious than in non-carious teeth. The
geometric mean lead concentration in carious teeth was 1.36 |ig/g
compared to 1.10 |_ig/g (p = 0.001).
-------
Table AX6-9.11. Effects of Lead on Ocular Health in Children and Adults
to
o
o
Reference, Study
Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
X
Oi
to
Children
Latin America
Rothenberg et al.
(2002b)
Mexico
1987-1997
Design = longitudinal (subset of prospective)
Subjects: children (n = 45, 24 female), ages
7-1 Oyr
Outcome measures: ERG
Analysis: comparison of outcome measures
between blood lead tertiles (ANOVA for
repeated measures)
Blood lead (|_ig/dL) median
(range) at 85-124 mo:
1st tortile: 4.0 (2.CM.5)
2nd tortile: 6.0(5.0-6.5)
3rd tortile: 7.5 (7.0-16.0)
Blood lead (|_ig/dL) median
(range), maternal at 12 wk
of gestation =
1st tortile: 4.0 (2.0-5.5)
2ndtertile: 8.5(6.0-10.0)
3rdtertile: 14.0(10.5-32.5)
Significant association between increasing maternal blood lead at 12
wk of gestation and increasing ERG a-wave (p = 0.025) and b-wave
amplitude (p = 0.007), with significant increases in a-wave in the
2nd blood lead tortile (6.0-10.0 |ig/dL), and a-wave and b-wave in
the 3rd blood lead tortile (10.5-32.5 |ig/dL), compared to the
1st blood lead tortile.
H
6
o
o
H
O
O
H
W
O
O
HH
H
W
Adults
United States
Schaumberg et al.
(2004)
Massachusetts
1991-2002
Design = longitudinal (subset of Normative Blood lead (|ig/dL) median Significant covariate adjusted odds ratio (OR) for cataracts in 5
Aging Study)
Subjects: adult male (n = 642), mean age 69
yr (range: 60-93)
Outcome measures: cataract diagnosis
Analysis: multivariate logistic regression,
odds ratio (vs. 1st quintile)
(range):
5(0-35)
Bone lead (|ig/g) median
(range):
patella: 29 (0-165)
tibia: 20 (0-126)
tibia bone lead quintile (31.0-125 |ig/g): OR 3.19 (95% CI: 1.48-
6.90, p = 0.01). OR for cataracts were not significantly associated
with patella bone lead (5th quintile: 43.0-165 |ig/g): OR 1.88 (95%
CI: 0.88^.02) or blood lead (5th quintile: 8.17-35.0 |ig/dL): OR
0.89 (95% CI: 0.46-1.72, p = 0.73).
Covariates retained: age, smoking, history of diabetes, daily intake
of vitamin C, vitamin E, and carotenoids.
-------
Table AX6-9.11 (cont'd). Effects of Lead on Ocular Health in Children and Adults
g Reference, Study
O Location, and Period
Study Description
Lead Measurement
Findings, Interpretation
Adults
Europe
Cavalleri et al. (1982)
Italy
NR
Design = cross-sectional cohort
Subjects: adult male vinyl chloride pipe
manufacture workers, exposed to lead
stearate (n = 35), mean age 45 yr (SD 14,
range: 21-59); reference group (n = 35)
matched for age, smoking, and alcohol
consumption.
Outcome measures: visual field
Analysis: comparison of outcome measures
between lead workers and reference group
Blood lead (|_ig/dL) mean
(SD, range):
lead: 46 (14, 21-82)
reference: 23 (4, 21-37)
Urine lead (|-ig/L) mean
(SD, range):
lead: 71 (18, 44-118)
reference: 30(5,21^12)
Visual sensitivity was significantly (p = 0.003) lower in lead workers
compared to the reference group; however, visual sensitivity index
was not significantly associated with blood or urine lead. Mesopic
field scotoma prevalence was 10 of 35 (28%) in lead workers and
0% in the reference group.
X
Oi
to
ERG, electroretinogram
H
6
o
o
H
O
o
H
W
O
O
HH
H
W
-------
ANNEX SECTION AX6-10
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1 The analyses fitting both the linear and log-linear models assumed that the error in the
2 response variable was constant across the range of values of the independent variable.
3 Violations of this assumption (heteroscedasticity) could potentially bias the estimated slope of
4 the model. Two models were considered:
5
6 Linear model: IQ = 90.0 - 0.4 x (blood lead level - 10.0), and
7 Log-linear model: IQ = 90.0 - 4.0 x [ln(blood lead level) - ln(10.0)].
8
9 The standard deviation of IQ was assumed to be equal to 15 x (blood lead level / I0)h
10 where h is the heteroscedasticity factor. When h = 0 there is no heteroscedasticity, and when
11 h = 1 the standard deviation is proportional to the value of the blood lead. The value of h = 1
12 would be comparable to the situation where there is a lognormal error.
13 The linear regression models described above were simulated for a sample size of
14 200 subjects and a lognormal distribution of blood lead levels with a geometric mean of 10.0
15 and a geometric standard deviation of 1.5. For each set of models and values of h, 100,000
16 simulations were performed.
Table AX6-10.1. Average Estimated Slopes for Linear and Log-linear Models in the
Presence of Heteroscedasticity
Heteroscedasticity
(h)
0.0
0.5
1.0
Linear Model
(True slope = -0.4)
-0.400
-0.400
-0.399
Log-Linear Model
(True slope = -4.0)
-4.00
-4.00
-4.01
17 The simulations indicated that any presence of heteroscedasticity would have no
18 noticeable bias on the estimation of the slopes of the models.
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i AX8. CHAPTER 8 ANNEX - ENVIRONMENTAL
2 EFFECTS OF LEAD
3
4
5 AX8.1 TERRESTRIAL ECOSYSTEMS
6 AX8.1.1 Methodologies Used in Terrestrial Ecosystems Research
7 The distribution of Pb throughout the terrestrial ecosystem, via aerial deposition, has been
8 discussed throughout this document. Its further impacts on soil, sediment, and water provide
9 numerous pathways that may promote unacceptable risk to all levels of biota. Stable isotopes of
10 Pb have been found useful in identifying sources and apportionment to various sources. One of
11 the key factors affecting assessment of risk is an understanding, and perhaps quantification, of
12 bioavailability. Therefore, the bioavailability of Pb is a key issue to the development of
13 NAAQSs. However, the discussion of all methods used in characterizing bioavailability is
14 beyond the scope of this chapter. The following topics are discussed in this chapter.
15 • Lead Isotopes and Apportionment
16 • Methodologies to determine Pb speciation
17 • Lead and the Biotic Ligand Model (BLM)
18 • In situ methods to reduce Pb bioavailability
19
20 AX8.1.1.1 Lead Isotopes and Apportionment
21 Determination of the extent of Pb contamination from an individual source(s) and its
22 impact are of primary importance in risk assessment. The identification of exposure pathway(s)
23 is fundamental to the risk analysis and critical in the planning of remediation scenarios.
24 Although societies have been consuming Pb for nearly 9,000 years, production of Pb in
25 the United States peaked in 1910 and 1972, at approximately 750 and 620 kt/year, respectively
26 (Rabinowitz, 2005). The diversity of potential Pb sources (fossil fuel burning, paint pigments,
27 gasoline additives, solders, ceramics, batteries) and associated production facilities (mining,
28 milling, smelting-refining) make fingerprinting of sources difficult. (See Chapter 2 and its
29 Annex for additional information on sources.) Therefore, dealing with multiple sources (point
30 and non-point), a reliable and specific fingerprinting technique is required. It has been well
31 established (Sturges and Barrie, 1987; Rabinowitz, 1995) that the stable isotope composition of
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1 Pb is ideally suited for this task. Lead isotopic ratio differences often allow multiple sources to
2 be distinguished, with an apportionment of the bulk Pb concentration made to those sources.
3 Lead has four stable isotopes: 204Pb, 206Pb, 207Pb, and 208Pb in natural abundances of 1.4,
4 24.1, 22.1, and 52.4%, respectively. The radiogenic 206Pb, 207Pb, and 208Pb are produced by
5 radioactive decay of 238U, 235U, and 232Th, respectively. Thus, the isotopic composition of Pb
6 varies based on the U:Pb and Th:Pb ratios of the original ore's source and age (Faure, 1977).
7 Because of the small fractional mass differences of the Pb isotopes, ordinary chemical and
8 pyrometallurgical reactions will not alter their original composition. Therefore, anthropogenic
9 sources reflect the isotopic composition of the ores from which the Pb originated.
10 To acquire the Pb isotopes, a sample, generally in aqueous form, is analyzed on an
11 ICP/MS (quadrapole, magnetic sector, or time-of-flight). Studies reviewing the most common
12 analytical and sample preparation procedures include Ghazi and Millette (2004), Townsend et al.
13 (1998), and Encinar et al. (2001a,b). The correction factors for mass discrimination biases are
14 generally made by analyzing the National Institute for Standards and Technology (NIST),
15 Standard Reference Material (SRM) 981 and/or spiked 203T1 and 205T1 isotopes (Ketterer et al.,
16 1991; Begley and Sharp, 1997). The overall success of Pb isotope fingerprinting is generally
17 dependent on analysis precision, which in turn depends on the type of mass analyzer used
18 (Table AX8-1.1.1).
19
20
Table AX8-1.1.1. Relative Standard Deviation (RSD) for Lead Isotope
Ratios on Selected Mass Spectrometers
RSD
204pb.206pb
207pb.206pb
208pb:206pb
Quadrapole
0.0031
0.0032
0.0026
Double-Focusing
0.0032
0.0027
0.0024
Single-Focusing
Magnetic Sector
0.00053
0.00053
0.00053
High-Resolution
Magnetic Sector
ICP/MS
0.0011
0.00048
0.00046
21 An extensive database comprising primarily North American Pb sources can be assembled
22 from Doe and Rohrbough (1977), Doe and Stacey (1974), Doe et al. (1968), Heyl et al. (1974),
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1 Leach et al. (1998), Stacey et al. (1968), Zartman (1974), Cannon and Pierce (1963), Graney
2 et al. (1996), Unruh et al. (2000), James and Henry (1993), Rabinowitz (2005), and
3 Small (1973).
4 The use of Pb isotopes to quantitatively apportion source contributions follows the simple
5 mixing rule when only two sources are possible (Faure, 1977). Once multiple sources need to be
6 considered, a unique solution can no longer be calculated (Fry and Sherr, 1984). Phillips and
7 Gregg (2003) have designed a model to give feasible source contributions when multiple sources
8 are likely. Many studies have demonstrated the usefulness of this apportionment technique.
9 Media of all types have been studied: water (Flegal et al., 1989a,b; Erel et al., 1991; Monna
10 et al., 1995), ice (Planchon et al., 2002), dust (Adgate et al., 1998; Sturges et al., 1993), and
11 soil/sediments (Hamelin et al., 1990; Farmer et al., 1996; Bindler et al., 1999; Haack et al., 2004;
12 Rabinowitz and Wetherill, 1972; Rabinowitz, 2005; Ketterer et al., 2001).
13
14 AX8.1.1.2 Speciation in Assessing Lead Unavailability in the Terrestrial Environment
15 One of the three processes defined by the National Research Council in its review on
16 bioavailability (NRC, 2002) is "contaminant interactions between phases", more commonly
17 referred to as "speciation."
18 A wide variety of analytical (XRD, EPMA, EXAFS, PIXE, XPS, XAS, SIMS) and
19 chemical speciation modeling (SOILCHEM, MINTEQL, REDEQL2, ECOSAT, MINTEQA2,
20 HYDRAQL, PHREEQE, WATEQ4F) tools have been used to characterize a metal's speciation
21 as it is found in various media. Currently, for risk assessment purposes (not considering
22 phytotoxicity), where large sites with numerous media, pathways, and metals must often be
23 characterized in a reasonable time frame, electron microprobe analysis (EMPA) techniques
24 provide the greatest information on metal speciation. Other techniques such as extended X-ray
25 absorption fine structure (EXAFS) and extended X-ray absorption near edge spectroscopy
26 (EXANES) show great promise and will be important in solving key mechanistic questions.
27 In the case of phytotoxicity, the speciation of metals by direct measurement or chemical models
28 of pore-water chemistry is most valuable. Further work needs to be done in developing
29 analytical tools for the speciation of the methyl-forming metals (Hg, As, Sb, Se, and Sn) in soils
30 and sediments.
31
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1 Concept
2 For a given metal or metalloid (hereafter referred to as metal), the term speciation refers
3 to its chemical form or species, including its physicochemical characteristics that are relevant to
4 bioavailability. As a result of the direct impact these factors often have on a metal's
5 bioavailability, the term "bioaccessibility" has been adopted to define those factors.
6
7 Speciation Role
8 The accumulation of metals in the lithosphere is of great concern. Unlike organic
9 compounds, metals do not degrade and, thus, have a greater tendency to bioaccumulate. It is
10 now well accepted that knowledge of the bulk, toxic characteristic leaching procedure (TCLP),
11 or synthetic leaching procedure (SLP) concentrations for any metal is not a controlling factor in
12 understanding a metal's environmental behavior or in developing remedies for its safe
13 management. Although these tests are essential to site characterization and management, they
14 offer no insight into risk assessment. Rather, it is the metal's bioavailability (the proportion of a
15 toxin that passes a physiological membrane [the plasma membrane in plants or the gut wall in
16 animals] and reaches a target receptor [cytosol or blood]), which plays a significant role in the
17 risk assessment of contaminated media.
18 The NRC review (NRC, 2002) on bioavailability defined bioavailability processes in
19 terms of three key processes:
20 • contaminant interactions between phases (association-dissociation/bound-released),
21 • transport of contaminants to organism, and
22 • passage across a physiological membrane.
23
24 As mentioned previously, the first process is more commonly referred to as speciation.
25 The speciation of a toxic metal in the environment is a critical component of any ecosystem
26 health risk assessment. Four important toxicologic and toxicokinetic determinants relating
27 speciation to bioavailability are the (1) chemical form or species, (2) particle size of the metal
28 form, (3) lability of the chemical form, and (4) source.
29
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4
5
6
7
8
9
10
Chemical Form of Species
The solid phase in a medium controls the activity of a metal in solution, whether the
solution is surface, ground, or pore water or GI fluids, and plays a profound role in metal
bioavailability. This is perhaps best illustrated by in vivo and in vitro results for many of the
common Pb-bearing minerals (Drexler, 1997) (Figure AX8-1.1.1). The metal species found in
media are often diverse, and data suggest that their bioavailability may be significantly
influenced by site-specific variations within these identified metal species (Davis et al., 1993;
Ruby et al., 1992; Drexler and Mushak, 1995).
LOW (1-20)
PbS
Slag
Pb-FeOOH
Native Pb
MEDIUM (20-60)
Pb-MnOOH
PbSO4
PbPO4
15 8
-------
1 (1979) observed that "the smaller the lead particle, the higher blood lead level." Similar
2 observations were made by Healy et al. (1992) using galena (PbS) and an in vitro dissolution
3 technique. Drexler (1997) presented in vitro results on numerous Pb-bearing phases ranging in
4 particle size from 35 to 250 jim. While all phases studied showed increased bioavailability with
5 decreasing particle size, more significantly, not all forms showed the same degree or magnitude
6 of change (Figure AX8-1.1.2). Atmospheric particles are generally found to occur in bimodal
7 populations: fine; 0.1 to 2.5 jim and coarse; 2.5 to 15 jim. This distribution is both a function of
8 there transport mechanism and emission source. Although the upper size limit for particles that
9 can be suspended in air is about 75 jim (Cowherd et al., 1974), other means of mechanical
10 entrainment (saltation, and creep) can transport particles as large as 1000 jim, supporting the
11 importance of fugative emissions on media contamination. In addition, particle size can change
12 post depositonal, as soluble forms re-precipitate or sorb onto other surfaces. Limited data are
13 available on the particle-size of discrete Pb phases from multi-media environments. One
14 example is the study by Drexler, 2004 at Herculaneum, Missouri. At this site, galena (PbS) was
15 the dominant Pb species with mean particle-size distributions of 4, 6 and 14 jim in PMio filters,
16 house dust, and soils, respectively. These findings support the conclusion that aerial transport
17 was the primary mechanism for Pb deposition in residential yards. Finally, such laboratory data
18 have been supported by extensive epidemiologic evidence, enforcing the importance of particle
19 size (Bornschein et al., 1987; Brunekreef et al., 1983; Angle et al., 1984).
20
21 Particle Lability
22 The impact on bioavailability of a metal particle's lability (its associations within the
23 medium matrix) is not well documented, but it follows the premise put forth by many of the
24 developing treatment technologies regarding its being bound or isolated from its environment.
25 Data from several EPA Superfund sites and the Region VIII swine study (U.S. Environmental
26 Protection Agency, 2004a) suggest that matrix associations, such as liberated versus enclosed,
27 can play an important part in bioavailability. As illustrated in Figure AX8-1.1.3, two different
28 media with similar total Pb concentrations and Pb forms (slag, Pb-oxide, and Pb-arsenate)
29 exhibit significantly different bioavailabilities. In the Murray, UT sample (bioaccumulation
30 factor [BAF] = 53%), a greater fraction of the more bioavailable Pb-oxides are liberated and
31 not enclosed in the less-soluble glass-like slag as observed in the Leadville, CO sample
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Anglesite
20 40
Minutes
60
38|jm
b.
150r
Slag
c.
20 40
Minutes
60
100,-
PbO
Figure AX8-1.1.2. Variation of bioavailability with particle size.
4
5
6
7
(BAF = 17%). Other evidence is more empirical, as illustrated in Figure AX8-1.1.4, where a
large particle of native Pb is shown to have developed a weathering ring of highly bioavailable
Pb-chloride and Pb-oxide. Such observations can be useful in understanding the mechanistic
phenomena controlling bioavailability. In addition, they will aid in developing and validating
models to predict metal-environment interactions.
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Murry
BAF 53%
11500 mg/kg Pb
20% liberated
Leadville AV
BAF 17%
10600 mg/kg Pb
5% liberated
Figure AX8-1.1.3.
Illustration of particle lability and bioavailability at two different sites
with similar total Pb concentrations and Pb forms.
1500|jm
BEI Baseline
Figure AX8-1.1.4. Scanning electron micrograph of a large native Pb particle showing
outer ring of highly bioavailable Pb-chloride and Pb-oxide.
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1 Source
2 Although the source of a metal is not directly related to bioavailability, it plays an
3 important role in risk assessment with the evaluation of metal (1) pathways, (2) background,
4 and (3) apportionment. It is important to understand a metal's pathway before any remedial
5 action can be taken; otherwise, recontamination of the primary pathway and reexposure can
6 occur. Knowledge of background is important, as an action level cannot be established below
7 natural background levels.
8
9 Plants
10 When considering the bioavailability of a metal to plants from soils and sediments, it is
11 generally assumed that both the kinetic rate of supply and the speciation of the metal to either the
12 root or shoot are highly important. In soils and sediments generally, only a small volume of
13 water is in contact with the chemical form, and although the proportion of a metal's
14 concentration in this pore water to the bulk soil/sediment concentration is small, it is this phase
15 that is directly available to plants. Therefore, pore water chemistry (i.e., metal concentration as
16 simple inorganic species, organic complexes, or colloid complexes) is most important.
17 Tools currently used for metal speciation for plants include (1) in-situ measurements
18 using selective electrodes (Gundersen et al., 1992; Archer et al., 1989; Wehrli et al., 1994);
19 (2) in-situ collection techniques using diffusive equilibrium thin films (DET) and diffusive
20 gradient thin films (DOT) followed by laboratory analyses (Davison et al., 1991, 1994; Davison
21 and Zhang, 1994; Zhang et al., 1995); and (3) equilibrium models ( SOILCHEM) (Sposito and
22 Coves, 1988).
23
24 AX8.1.1.3 Tools for Bulk Lead Quantification and Speciation
25 Bulk Quantification
26 The major analytical methods most commonly used for bulk analyses outlined in the 1986
27 Pb ACQD included:
28 • Atomic Absorption Spectrometry (AAS)
29 • Emission Spectrometry (Inductively coupled plasma/atomic emission spectrometry)
30 • X-ray Fluorescence (XRF)
31 • Isotope Dilution Mass Spectrometry (ID/MS)
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1 • Colorimetric
2 • Electrochemical (anodic stripping voltametry and differential pulse polarography).
3 The choice of analytical method today for bulk quantification is generally ICP/AES or
4 ICP/MS (U.S. Environmental Protection Agency, 2001). Since 1986, numerous SRMs have
5 been developed for Pb (Table AX8-1.1.2), and several significant technological improvements
6 have been developed.
7
8
Table AX8-1.1.2.
NIST SRM
2710
2711
2709
2587
2586
2783
1648
1649a
2584
2583
1515
1575
National Institute of Standards
Medium
Soil
Soil
Soil
Soil (paint)
Soil (paint)
Filter (PM2 5)
Urban particulate
Urban dust
Indoor dust
Indoor dust
Apple leaves
Pine needles
and Technology Lead SRMs
Mean Pb
mg/kg
5532
1162
18.9
3242
432
317
6550
12,400
9761
85.9
0.47
0.167
9 Modern spectrometry systems have replaced photomultiplier tubes with a charge-coupled
10 device (CCD). The CCD is a camera that can detect the entire light spectrum (>70,000 lines)
11 from 160 to 785 nm. This allows for the simultaneous measurement of all elements, as well as
12 any interfering lines (a productivity increase), and increases precision. The detection limit for Pb
13 in clean samples can now be as low as 40 ppb.
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1 Modern ICP/AES systems offer a choice of either axial viewed plasma (horizontal),
2 which provides greater sensitivity (DL= 0.8 jig Pb/L), or radial (vertical) viewed plasma, which
3 performs best with high total dissolved samples (DL = 5.0 jig Pb/L).
4 The development of reaction or collision cells have expanded the capabilities of ICP/MS
5 and lowered detection limits for many elements that were difficult to analyze because of
6 interferences such as Se, As, Ti, Zn, Ca, Fe, and Cr. The cells provide efficient interference
7 (isobaric, polyatomic, and argide) removal independent of the analyte and sample matrix by
8 using various reaction gases (H2, He, NH3), eliminating the need for interference correction
9 equations.
10
11 Speciation Tools
12 A wide variety of analytical and chemical techniques have been used to characterize a
13 metal's speciation (as defined above) in various media (Hunt et al., 1992; Manceau et al., 1996,
14 2000a; Welter et al., 1999; Szulczewski et al., 1997; Isaure et. al., 2002; Lumsdon and Evans,
15 1995; Gupta and Chen, 1975; Ma and Uren, 1995; Charlatchka et al., 1997). Perhaps the most
16 important factor that one must keep in mind in selecting a technique is that, when dealing with
17 metal-contaminated media, one is most often looking for the proverbial "needle in a haystack."
18 Therefore, the speciation technique must not only provide the information outlined above, but it
19 must also determine that information from a medium that contains very little of the metal.
20 As illustrated in Figure AX8-1.1.5, for a Pb-contaminated soil, less than 1% (modally) of a
21 single species can be responsible for a bulk metal's concentration above an action level. This
22 factor is even more significant for other metals (i.e., As, Cd, or Hg) were action levels are often
23 below 100 mg/kg.
24 Of the techniques tested (physicochemical, extractive, and theoretical), the tools that have
25 been used most often to evaluate speciation for particle-bound metal include X-ray absorption
26 spectroscopy (XAS), X-ray diffraction (XRD), particle induced X-ray emission (PIXE and
27 jiPIXE), electron probe microanalysis (EPMA), secondary ion mass spectrometry (SIMS),
28 X-ray photoelectron spectroscopy (XPS), sequential extractions, and single chemical extractions.
29 The tools that have been used most often to evaluate speciation for metal particles in solution
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vs
SLAG
-------
1 only a few currently provide useful information on metal bioavailability at a "site" level.
2 However, one may still find other techniques essential to a detailed characterization of a selected
3 material to describe the chemical or kinetic factors controlling a metal's release, transport,
4 and/or exposure.
5 X-Ray absorption Spectroscopy (XAS). X-ray absorption spectroscopy (XAS) is a
6 powerful technique using the tunable, monochromatic (white light) X-rays produced by a
7 synchrotron (2-4 GeV) to record oscillations in atomic absorption within a few 100 eV of an
8 element's absorption edge. Spectra provide both information on chemical state and atomic
9 structure. Measurements are theoretically available for all elements and are not surface-sensitive
10 nor sample-sensitive (i.e., gases, liquids, solids, and amorphous materials are testable).
11 High-energy spectra within 30 eV of the edge, termed XANES (X-ray absorption near
12 edge structure spectroscopy (Fendorf et al., 1994; Maginn, 1998), are particularly suited for
13 determination and quantification (10 to 100 ppm) of metal in a particular oxidation state
14 (Szulczewski et al., 1997; Shaffer et al., 2001; Dupont et al., 2002). The lower-energy spectra
15 persist some 100 eV above the edge. These oscillations are termed EXAFS (extended X-ray
16 absorption fine structure) and are more commonly used for speciation analyses (Welter et al.,
17 1999; Manceau et al., 1996, 2000a; Isaure et al., 2002).
18 The main limitations to XAS techniques are (1) the lack of spatial resolution; (2) XAS
19 techniques provide only a weighted average signal of structural configurations, providing
20 information on the predominant form of the metal, while minor species, which may be more
21 bioavailable, can be overlooked; (3) access to synchrotrons is limited and the beam time required
22 to conduct a site investigation would be prohibitive; (4) a large spectral library must be
23 developed; (5) generally, poor fits to solution models are achieved when the compound list is
24 large; and (6) high atomic number elements have masking problems based on compound density.
25 X-Ray Diffraction (XRD). In X-ray diffraction, a monochromatic Fe, Mo, Cr, Co, W, or
26 Cu X-ray beam rotates about a finely powdered sample and is reflected off the interplanar
27 spacings of all crystalline compounds in the sample, fulfilling Bragg's law (nX = 2dsin0). The
28 identification of a species from this pattern is based upon the position of the lines (in terms of
29 9 or 29) and their intensities as recorded by an X-ray detector. The diffraction angle (29) is
30 determined by the spacing between a particular set of atomic planes. Identification of the species
31 is empirical, and current available databases contain more than 53,000 compounds.
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1 If a sample contains multiple compounds, interpretation becomes more difficult and
2 computer-matching programs are essential. In some instances, by measuring the intensity of the
3 diffraction lines and comparing them to standards, it is possible to quantitatively analyze
4 crystalline mixtures; however, if the species is a hydrated form or has a preferred orientation, this
5 method is only semi quantitative at best. Since this technique represents a bulk analysis, no
6 particle size or lability information can be extracted from the patterns.
7 Particle InducedX-Ray Emission (PIXE and juPIXE). Particle induced X-ray emission
8 (PIXE) uses a beam, ~4 jim in diameter, of heavy charged particles (generally He) to irradiate
9 the sample. The resulting characteristic X-rays are emitted and detected in a similar manner as
10 XRF, using Si-Li detectors. Particles generated from a small accelerator or cyclotron, with a
11 potential of 2 to 4 MeV, are commonly used. Detection limits on the order of 1 mg/kg are
12 achieved on thin-film samples. Disadvantages to its use for speciation include (1) only a small
13 volume of material can be analyzed (1 to 2 mg/cm2); (2) no particle size information is provided;
14 (3) peak overlaps associated with Si-Li detectors limit identification of species; (4) limited
15 availability; and (5) high cost. For a further review of PIXE analysis and applications, see
16 Maenhaut(1987).
17 Electron Probe Microanalysis (EPMA). Electron probe microanalysis uses a finely
18 focused (1 |im) electron beam (generated by an electron gun operating at a 2 to 30 kV
19 accelerating voltage and pico/nanoamp currents) to produce a combination of characteristic
20 X-rays for elemental quantification along with secondary electrons and/or backscatter electrons
21 for visual inspection of a sample. Elements from beryllium to uranium can be nondestructively
22 analyzed at the 50-ppm level with limited sample preparation. X-ray spectra can be rapidly
23 acquired using either wavelength dispersive spectrometers (WDS) or energy dispersive
24 spectrometers (EDS).
25 With WDS, a set of diffracting crystals, of known d-spacing, revolve in tandem with a
26 gas-filled proportional counter inside the spectrometer housing so that Bragg's law is satisfied
27 and a particular wavelength can be focused. Photon energy pulses reflecting off the crystal are
28 collected for an individual elemental line by the counter as a first approximation to
29 concentration. For quantitative analysis, these intensities are compared to those of known
30 standards and must be corrected for background, dead time, and elemental interactions (ZAF)
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1 (Goldstein et al., 1992). ZAP correction is in reference to the three components of matrix
2 effects: atomic number (Z), absorption (A), and fluorescence (F).
3 With EDS, a single Si-Li crystal detector is used in conjunction with a multichannel
4 analog-digital converter (ADC) to sort electrical pulses (with heights approximately proportional
5 to the quantum energy of the photon that generated them), producing a spectrum of energy
6 (wavelength) versus counts. The net area under a particular peak (elemental line) is proportional
7 to its concentration in the sample. For quantitative analyses, corrections similar to WDS analysis
8 must be performed. Although EDS detectors are more efficient than WDS, detection limits are
9 significantly greater (-1000 ppm), because of elevated backgrounds and peak overlaps.
10 For speciation analysis, the EDS system must NEVER be used as the primary detector, as
11 numerous errors in species identification are often made. These are generally the result of
12 higher-order X-ray line overlaps.
13 This technique has been routinely used for site characterizations (Linton et al., 1980;
14 Hunt et al., 1992; Camp, Dresser, and McKee (COM), 1994; U.S. Environmental Protection
15 Agency, 2002). Currently this technique offers the most complete data package on metal
16 speciation than any of the other tools. The method is relatively fast and inexpensive, available,
17 and provides all of the required information for bioavailability assessments (i.e., particle size,
18 species, lability, and sourcing). A number of limitations still need to be addressed including:
19 (1) its inability to quickly isolate a statistically significant population of particles in soils with
20 low bulk metal concentrations (<50 mg/kg), meaning that for some metals with low
21 concentrations of concern (i.e., Cd, Mo, Sb, Se), this method may be less useful; (2) the more
22 volatile metals (i.e., Hg, Tl) are often volatilized under the electron beam or lost during sample
23 preparation.
24 Secondary Ion Mass Spectrometry (SIMS). Secondary ion mass spectrometry (also known
25 as ion microprobes or ion probes) is a well-known technique, primarily surface focused, that uses
26 a 0.5 to 20 kV O, Ar, Ga, In, or Cs ion beam in bombarding (sputtering) the surface of a sample
27 while emitting secondary ions that are detected by either quadrapole, time-of-flight (TOF), or
28 magnetic sector mass spectrometers. Sensitivity is very high, in the ppb range for elements
29 hydrogen to uranium, providing quantitative results on elemental or isotopic metals and organic
30 compounds. With the advent of liquid metal (In and Ga) ion beams, beam sizes of less than 1
31 |im are possible, although 20 jim is more commonly used.
May 2006 AX8-15 DRAFT-DO NOT QUOTE OR CITE
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1 The major disadvantage of SIMS to species identification is that each element or isotope
2 must be tuned and analyzed sequentially. This makes the identification of a metal form highly
3 time-consuming and, thus, the characterization of a multiphase medium impractical.
4 X-Ray Photoelectron Spectroscopy (XPS). X-ray photoelectron spectroscopy or ESC A
5 (electron spectroscopy for chemical analysis, as it was previously known) is a classical surface,
6 10 to 50 A in depth, analytical technique for determinating qualitative elemental concentrations
7 of elements greater than He in atomic number and provides limited structural and oxidation state
8 information. In XPS, the high-energy (15 kV) electrons are typically produced from a dual-
9 anode (Al-Mg) X-ray tube. The excitation or photoionization of atoms within the near surface of
10 the specimen emit a spectrum of photoelectrons. The measured binding energy is characteristic
11 of the individual atom to which it was bound. Monochromatic sources are often employed to
12 improve energy resolution, allowing one to infer oxidation states of elements or structure of
13 compounds (organic and inorganic) by means of small chemical shifts in binding energies
14 (Hercules, 1970). The major disadvantages of XPS for environmental speciation studies is its
15 poor sensitivity, especially in complex matrices and its large, 100-200 jim, spatial resolution.
16 The direct speciation techniques discussed above are summarized in Table AX8-1.1.3.
17
18 Indirect Approaches
19 A more indirect approach to speciation than the methods previously described include the
20 functional or operational extraction techniques that have been used extensively over the years
21 (Tessier et al, 1979; Tessier and Campbell, 1988; Gupta and Chen, 1975). These methods use
22 either a single or sequential extraction procedure to release species associated with a particular
23 metal within the media. Single chemical extractions are generally used to determine the
24 bioavailable amount of metal in a functional class: water-soluble, exchangeable, organically
25 bound, Fe-Mn bound, or insoluble.
26 In a similar approach, sequential extractions treat a sample with a succession of reagents
27 intended to specifically dissolve different, less available phases. Many of these techniques have
28 been proposed, most of which are a variation on the classical method of Tessier et al. (1979),
29 in which metal associated with exchangeable, carbonate-bound, Fe-Mn bound, organically
30 bound, and residual species can be determined. Beckett (1989), Kheboian and Bauer (1987),
31 and Foerstner (1987) provide excellent reviews on the use and abuse of extractions. These
May 2006 AX8-16 DRAFT-DO NOT QUOTE OR CITE
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Table AX8-1.1.3. Characteristics for Direct Speciation Techniques
to
o
o
ON
>
X
oo
1
o
£>
•3
.a
s5
J
5
'S
a.
Tools ^
XRD No
EMPA Yes
SIMS No
XPS No
XAS No
PIXIE No
U O feft
•S O fi
•sS'-S ou^s-^a^
*" ~" s ^ '-5 ^ a a •"•" .aS a •'• -2 -a
* .§ >| « |'| '1| si fl si S •* 1
S^ sS(/5 ^ aa °-s — & ocs ^a * '"3 u
'3 '3 '3 ^S ^ * wa. *-S fl W« v >
«Ss ri-^^^^"-^
a. a, S. ^
&c &c &c
No No No No# No No No 3-4vol% Bulk 1 $
Yes Yes+ No Yes Yes? B-U No*** 50 ppm 0.5-1 um 2 $$
Yes No No Yes* Yes** Li-U Yes 1 ppb 10 um 4 $$$
No Yes Yes Yes* Yes** H-U No wt.% 100 um 2 $$
No Yes Yes Yes* Yes** He-U No ppb 2 um 5 $$$$
No No No Yes Yes** B-U No 10 ppm 4 um 4 $$$$
* Technique requires each element be tuned and standardized, requiring unreasonable time limits.
W ** Techniques designed and tested only on simple systems. Multiple species require lengthy analytical times and data reduction.
ITJ * * * Limited when combined with ICP/MS/L A.
H
6
0
H
O
O
H
W
O
O
HH
H
W
# Identifies crystalline
compounds and stoichiometric compositions only.
? Technique has limitations based on particle counting statistics.
+ Valance determined by charge balance of complete analyses.
-------
1 techniques can be useful in a study of metal uptake in plants, where transfer takes place
2 predominately via a solution phase. However, one must keep in mind that they are not
3 "selective" in metal species, give no particle size information and, above all, these teachable
4 fractions have never been correlated to bioavailability.
5 Solution Speciation Using Computer-Based Models. Computer-based models are either
6 based upon equilibrium constants or upon Gibb's free energy values in determining metal
7 speciation from solution chemistry conditions (concentration, pH, Eh, organic complexes,
8 adsorption/desorption sites, and temperature). Both approaches are subject to mass balance and
9 equilibrium conditions. These models have undergone a great deal of development in recent
10 years, as reliable thermodynamic data has become available and can provide some predictive
11 estimates of metal behavior. A good review of these models and their applications is provided
12 by Lumsdon and Evans (1995).
13 Speciation can be controlled by simple reactions; however, in many cases, particularly in
14 contaminated media, their state of equilibrium and reversibility are unknown. In addition, these
15 models suffer from other limitations such as a lack of reliable thermodynamic data on relevant
16 species, inadequacies in models to correct for high ionic strength, reaction kinetics are poorly
17 known, and complex reactions with co-precipitation/adsorption are not modeled.
18 The first limitation is perhaps the most significant for contaminated media. For example,
19 none of the models would predict the common, anthropogenic, Pb phases, i.e., paint, solder,
20 and slag.
21
22 AX8.1.1.4 Biotic Ligand Model
23 The biotic ligand model (BLM) is an equilibrium-based model that has been incorporated
24 into regulatory agencies guidelines (including the EPA) to predict effects of metals primarily on
25 aquatic biota and to aid in the understanding of their interactions with biological surfaces.
26 As initially presented by Paquin et al. (1999), the BLM evolved from both the gill surface
27 interaction model (GSIM) of Pagenkopf (1983) and the free ion activity model (FLAM) of Morel
28 (1983). The model can be used to define site-specific ambient water quality criteria (AWQC) by
29 providing the rational as to how metal toxicity to an aquatic organism is controlled by variations
30 in water chemistry.
May 2006 AX8-18 DRAFT-DO NOT QUOTE OR CITE
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1 By integrating the interaction of a metal in solution with its predicted speciation and
2 subsequent interaction with either a receptor site (e.g., root, gill, whole body) of an organism
3 (biotic ligand) a lethal concentration (LC50) estimate is made, replacing expensive, time
4 consuming bioassay testing. The biotic ligand is assumed to be independent and homogeneously
5 distributed and is essentially described using an affinity constant (Ks [M-l]) that have been
6 generated from laboratory studies. A current version (v 2.12) of the BLM can be downloaded
7 from: http://www.hydroqual.com/blm.
8 Currently, a limited metal/organism set ([Cu, Ag, Cd, and Zn] and [flathead minnow,
9 rainbow trout, Daphnia magna, Daphniapulex, and Ceidaphia dubia], respectively) are
10 provided. However, users are able to input site-specific metal/organism datasets if available.
11 The literature contains numerous studies on additional metals (i.e., Co, Ni, Pb, U, Sr, and Ba)
12 and aquatic organisms, references to which can be found in Slaveykova and Wilkinson (2005)
13 and Niyogi and Wood (2004). Site-specific water chemistry is entered as temperature, pH, metal
14 (Cu, Ag, Cd, and Zn), dissolved organic carbon (DOM), humic acid (HA), cations (Ca, Mg, Na,
15 and K), anions (Cl and 804), and alkalinity for speciation calculations.
16 Currently, there is no acute BLM for Pb; however the work of MacDonald et al. (2002) on
17 gill-Pb in rainbow trout and that of Slaveykova and Wilkinson (2002) on algae suggest that Ca2+,
18 DOM, and perhaps Na+ competitively inhibit Pb2+ uptake and thus exhibit a much lower (<100x)
19 affinity for the biotic ligand. Further toxicity testing must be conducted before an acute BLM for
20 Pb is established. Presently, affinity constants for Pb are limited to a few organisms
21 (TableAX8-1.1.4).
Table AX8-1.1.4. Affinity Constants for Lead
Organism
Phytoplankton
Bacteria
Fish
Cladoceran
Species
Chlorella kesslerii
Bacillus subtilis
Bacillus lichiformis
Rainbow trout
Hyalella azteca
log Ks [M-l]
5.5
3.4,5.1
4.4, 5.7
6.0
5.8,6.9
Reference
Slaveykova and Wilkinson (2002)
Daughney and Fein (1998)
Daughney and Fein (1998)
MacDonald et al. (2002)
Borgmann et al. (1993, 2004)
MacLean et al. (1996)
May 2006
AX8-19
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1 Because of assumed similarities in mechanisms of toxicity between aquatic and terrestrial
2 organisms, it is likely that the BLM approach as developed for the aquatic compartment may also
3 be applicable to the terrestrial environment. Recent research has been directed toward extending
4 the BLM to predict metal toxicity in soils (Steenbergen et al., 2005). Steenbergen et al. (2005)
5 pointed out that, until recently, the BLM concept has not been applied to predict toxicity to soil
6 organisms. The authors believe there may be two reasons for this. First, metal uptake routes in
7 soils are generally more complex than those in water, because exposure via pore water and
8 exposure via ingestion of soil particles may, in principle, both be important. Second, it remains
9 very difficult to univariately control the composition of the soil pore water and the metal
10 concentrations in the pore water, due to reequilibration of the system following modification of
11 any of the soil properties (including addition of metal salts).
12 Steenbergen et al. (2005) assessed acute copper toxicity to the earthworm Aporrectodea
13 caliginosa using the BLM. To overcome the aforementioned problems inherent in soil toxicity
14 tests they developed an artificial flow-through exposure system consisting of an inert quartz sand
15 matrix and a nutrient solution, of which the composition was univariately modified. Thus, the
16 obstacles in employing the BLM to terrestrial ecosystems seem to be surmountable, and future
17 research may provide useful information on Pb bioavailability and toxicity to terrestrial
18 organisms.
19
20 AX8.1.1.5 Soil Amendments
21 The removal of contaminated soil to mitigate exposure of terrestrial ecosystem
22 components to Pb can often present both economic and logistic problems. Because of this,
23 recent studies have focused on in situ methodologies to lower soil-Pb RBA (Brown et al.,
24 2003a,b). To date, the most common methods studied include the addition of soil amendments
25 in an effort either to lower the solubility of the Pb form or to provide sorbtion sites for fixation of
26 pore-water Pb. These amendments typically fall within the categories of phosphate, biosolid,
27 and Al/Fe/Mn-oxide amendments.
28
29 Phosphate Amendments
30 Phosphate amendments have been studied extensively and, in some cases, offer the most
31 promising results (Brown et al., 1999; Ryan et al., 2001; Cotter-Howells and Caporn, 1996;
May 2006 AX8-20 DRAFT-DO NOT QUOTE OR CITE
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1 Hettiarachchi et al., 2001, 2003; Yang et al., 2001; Ma et al., 1995). Research in this area stems
2 from early work by Nriagu (1973) and Cotter-Howells and Caporn (1996), who pointed out the
3 very low solubilities for many Pb-phosphates (Ksp -27 to -66), particularly chloropyromorphite
4 [Pbs(PO4)3Cl]. The quest to transform soluble Pb mineralogical forms into chloropyromorthite
5 continues to be the primary focus of most studies. Sources of phosphorous have included
6 phosphoric acid (HsPO/t), triple-super phosphate (TSP), phosphate rock, and/or hydroxyapatite
7 (HA). Various studies have combined one or more of these phosphorous sources with or without
8 lime, iron, and/or manganese in an attempt to enhance amendment qualities. Most amendments
9 are formulated to contain between 0.5 and 1.0% phosphorous by weight. They are then either
10 applied wet or dry and then mixed or left unmixed with the contaminated soil. Success of
11 phosphate amendments has been variable, and the degree of success appears to depend on
12 available phosphorous and the dissolution rate of the original Pb species.
13 A number of potentially significant problems associated with phosphate amendments have
14 been recognized, including both phyto- and earthworm toxicity (Ownby et al., 2005; Cao et al.,
15 2002; and Rusek and Marshall, 2000). Both of these toxicities are primarily associated with very
16 high applications of phosphorous and/or decreased soil pH. Indications of phytotoxicity are
17 often balanced by studies such as Zhu et al. (2004) that illustrate a 50 to 70% reduction in shoot-
18 root uptake of Pb in phosphate-amended soils. Additionally, the added phosphate poses the
19 potential risk of eutrophication of nearby waterways from soil runoff. Finally, Pb-contaminated
20 soils from the extractive metals industry or agricultural sites often have elevated concentrations
21 of arsenic. It has been shown (Impellitteri, 2005; Smith et al., 2002; Chaney and Ryan, 1994;
22 and Ruby et al., 1994) that the addition of phosphate to such soils would enhance arsenic
23 mobility (potentially moving arsenic down into the groundwater) through competitive anion
24 exchange. Some data (Lenoble et al., 2005) indicate that if one could amend arsenic and Pb
25 contaminated soils with iron(III) phosphate this problem can be mitigated, however the increased
26 concentrations of both phosphate and iron still exclude the application when drinking water is an
27 issue.
28
29 Biosolid Amendments
30 Historically, biosolids have been used in the restoration of coal mines (Haering et al.,
31 2000; Sopper, 1993). More recently, workers have demonstrated the feasibility of their use in
May 2006 AX8-21 DRAFT-DO NOT QUOTE OR CITE
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1 the restoration of mine tailings (Brown et al., 2003a), and urban soils (Brown et al., 2003b;
2 Farfel et al., 2005). Mine tailings are inherently difficult to remediate in that they pose numerous
3 obstacles to plant growth. They are most often (1) acidic; (2) high in metal content, thus prone to
4 phytotoxicity; (3) very low in organic content; and (4) deficient in macro- and micronutrients.
5 Stabilization (i.e., the establishment of a vegetative cover) of these environments is essential to
6 the control of metal exposure or migration from soil/dust and groundwater pathways.
7 At Bunker Hill, ID, Brown et al. (2003b) demonstrated that a mixture of high nitrogen
8 biosolids and wood pulp or ash, when surface applied at a rate of approximately 50 and
9 220 tons/ha, respectively, increased soil pH from 6.8 to approximately 8.0, increased plant
10 biomass from 0.01 mg/ha to more than 3.4 tons/ha, and resulted in a healthy plant cover within 2
11 years. Metal mobility was more difficult to evaluate. Plant concentrations of Zn and Cd were
12 generally normal for the first 2 years of the study; however, Pb concentrations in vegetation
13 dramatically increased two to three times in the first year. Additionally, macronutrients (Ca, K,
14 and Mg) decreased in plant tissue.
15 Urban soils, whether contaminated from smelting, paint, auto emissions, or industrial
16 activity, are often contaminated with Pb (Agency for Toxic Substances and Disease Registry
17 [ATSDR], 1988) and can be a significant pathway to elevated child blood Pb levels (Angle et al.,
18 1974). Typically, contaminated residential soils are replaced under Superfund rules. However,
19 urban soils are less likely to be remediated unless a particular facility is identified as the
20 contaminate source. Application of biosolids to such soils may be a cost-effective means for
21 individuals or communities to lower Pb RBAs.
22 A field study by Farfel et al. (2005) using the commercial biosolid ORGO found that,
23 over a 1-year period, Pb in the dripline soils of one residence had reduced RBAs by
24 approximately 60%. However, soils throughout the remainder of the yard showed either no
25 reduction in RBA or a slight increase. A more complex study was conducted by Brown et al.
26 (2003a) on an urban dripline soil in the lab. The study used an assortment of locally derived
27 biosolids (raw, ashed, high-Fe compost, and compost) with and without lime. All amendments
28 were incubated with approximately 10% biosolids for a little more than 30 days. In vitro and in
29 vivo data both indicated a 3 to 54% reduction in Pb RBA, with the high-Fe compost providing
30 the greatest reduction.
May 2006 AX8-22 DRAFT-DO NOT QUOTE OR CITE
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1 As with phosphate amendments, problems with biosolid application have also been
2 documented. Studies have shown that metal transport is significantly accelerated in soils
3 amended with biosolids (Al-Wabel et al., 2002; McBride et al., 1997, 1999; Lamy et al., 1993;
4 Richards et al., 1998, 2000). Some of these studies indicate that metal concentrations in soil
5 solutions up to 80 cm below the amended surface increased by 3- to 20-fold in concentration up
6 to 15 years after biosolid application. The increase in metal transport is likely the result of
7 elevated dissolved organic carbon (DOC) in the amended soil. Anodic stripping voltammetry
8 has indicated that very low percentages (2 to 18%) of the soluble metals are present as ionic or
9 inorganic complexes (McBride, 1999; Al-Wabel et al., 2002).
10
11 AX8.1.1.6 Future Needs
12 Since the 1986 Pb AQCD (U.S. Environmental Protection Agency, 1986a), considerable
13 data has been generated on the bioavailability process. The understanding of bioavailability is
14 central to improving risk assessments and designing efficient, cost-effective remediations. Four
15 key areas for future research can be identified.
16 • A set of bioavailability and speciation standards should be developed for traceability
17 and quality assurance to aid researchers in developing new or refining existing tools.
18 • An effort should be made to develop in vitro bioassays for nonhuman biota in order to
19 provide site-specific, rapid, cost-effective estimates of bioavailability/toxicity for all
20 levels of the ecosystem evaluated in a risk assessment.
21 • Research should continue on the development of in situ amendments to lower Pb
22 bioavailability, with a strong emphasis on long-term field validation studies.
23 • Finally, toxicity testing for expanding organism/metal affinity constants for both the
24 aquatic and terrestrial BLM should be continued, particularly for Pb.
25
26 AX8.1.2 Distribution of Atmospherically Delivered Lead in
27 Terrestrial Ecosystems
28 The 1986 Pb AQCD (U.S. Environmental Protection Agency, 1986a) contains only a few
29 minor sections that detail the speciation, distribution, and behavior of atmospherically delivered
30 Pb in terrestrial ecosystems. The document concluded that the majority of Pb in the atmosphere
31 at that time was from gasoline consumption: of the 34,881 tons of Pb emitted to the atmosphere
32 in 1984, 89% was from gasoline use and minor amounts were from waste oil combustion, iron
33 and steel production, and smelting. Lead in the atmosphere today, however, is not primarily
May 2006 AX8-23 DRAFT-DO NOT QUOTE OR CITE
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1 from gasoline consumption, but results largely from waste incineration, metal smelting, metal
2 production, and coal-fired power plants (Polissar et al., 2001; Newhook et al., 2003). The
3 emission source can determine the species of Pb that are delivered to terrestrial ecosystems. For
4 example, Pb species emitted from automobile exhaust is dominated by particulate Pb halides and
5 double salts with ammonium halides (e.g., PbBrCl, PbBrCbNH/tCl), while Pb emitted from
6 smelters is dominated by Pb-sulfur species (Habibi, 1973). The halides from automobile exhaust
7 break down rapidly in the atmosphere, possibly via reactions with atmospheric acids (Biggins
8 and Harrison, 1979). Lead phases in the atmosphere, and presumably the compounds delivered
9 to the surface of the earth (i.e., to vegetation and soils), are suspected to be in the form of PbSC>4,
10 PbS, and PbO (Olson and Skogerboe, 1975; Clevenger et al., 1991; Utsunomiya et al., 2004).
11 There are conflicting reports of how atmospherically derived Pb specifically behaves in
12 surface soils. This disagreement may represent the natural variability of the biogeochemical
13 behavior of Pb in different terrestrial systems, the different Pb sources, or it may be a function of
14 the different analytical methods employed. The importance of humic and fulvic acids (Zimdahl
15 and Skogerboe, 1977; Gamble et al., 1983) and hydrous Mn- and Fe-oxides (Miller and McFee,
16 1983) for scavenging Pb in soils are discussed in some detail in the 1986 Pb AQCD. Nriagu
17 (1974) used thermodynamics to argue that Pb-orthophosphates (e.g., pyromorphite) represented
18 the most stable Pb phase in many soils and sediments. He further suggested that, because of the
19 extremely low solubility of Pb-phosphate minerals, Pb deposition could potentially reduce
20 phosphorous availability. Olson and Skogerboe (1975) reported that solid-phase PbSO4
21 dominated gasoline-derived Pb speciationin surface soils from Colorado, Missouri, and Chicago,
22 while Santillan-Medrano and Jurinak (1975) suggested that Pb(OH)2, Pb(PO4)2, and PbCO3
23 could regulate Pb speciation in soils. However, insoluble organic material can bind strongly to
24 Pb and prevent many inorganic phases from ever forming in soils (Zimdahl and Skogerboe,
25 1977).
26 The vertical distribution and mobility of atmospheric Pb in soils was poorly documented
27 prior to 1986. Chapter 6 of the 1986 AQCD cited a few references suggesting that atmospheric
28 Pb is retained in the upper 5 cm of soil (Reaves and Berrow, 1984). Techniques using radiogenic
29 Pb isotopes had been developed to discern between gasoline-derived Pb and natural, geogenic
30 (native) Pb, but these techniques were mostly applied to sediments (Shirahata et al., 1980) prior
31 to the 1986 Pb AQCD. Without using these techniques, accurate determinations of the depth-
May 2006 AX8-24 DRAFT-DO NOT QUOTE OR CITE
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1 distribution and potential migration velocities for atmospherically delivered Pb in soils were
2 largely unavailable.
3 Several technological advances, combined with the expansion of existing technologies
4 after 1986 resulted in the publication of a large body of literature detailing the speciation,
5 distribution, and geochemical behavior of gasoline-derived Pb in the terrestrial environment.
6 Most notably, the development of selective chemical extraction (SCE) procedures as a rapid and
7 inexpensive means for partitioning Pb into different soil and sediment phases (e.g., Pb-oxides,
8 Pb-humate, etc.) has been exploited by a number of researchers (Tessier et al., 1979; Johnson
9 and Petras, 1998; Ho and Evans, 2000; Scheckel et al., 2003). Also, since 1986, several workers
10 have exploited synchrotron-based XAS in order to probe the electron coordination environment
11 of Pb in soils, organic matter, organisms, and sediments (Manceau et al., 1996; Xia et al., 1997;
12 Trivedi et al., 2003). X-ray absorption studies can be used for the in-situ determination of the
13 valence state of Pb and can be used to quantify Pb speciation in a variety of untreated samples.
14 Biosensors, which are a relatively new technology coupling biological material, such as an
15 enzyme, with a transducer, offer a new, simple, and inexpensive means for quantifying available
16 Pb in ecosystems (Verma and Singh, 2005). Advances in voltammetric, diffusive gradients in
17 thin films (DOT), and TCP techniques have also increased the abilities of researchers to quantify
18 Pb phases in solutions (Berbel et al., 2001; Scally et al., 2003). In addition to the development of
19 techniques for describing and quantifying Pb species in the soils and solutions, researchers have
20 used radiogenic Pb isotopes (206Pb, 207Pb, 208Pb) to quantify the distribution, speciation, and
21 transport of anthropogenic Pb in soil profiles and in vegetation (Bindler et al., 1999; Erel et al.,
22 2001; Kaste et al., 2003; Klaminder et al., 2005).
23 Over the past several decades, workers have also developed time-series data for Pb in
24 precipitation, vegetation, organic horizons, mineral soils, and surface waters. Since
25 atmospherically delivered Pb often comprises a significant fraction of the "labile" Pb (i.e., Pb not
26 associated with primary minerals), these data have been useful for developing transport and
27 residence time models of Pb in different terrestrial reservoirs (Friedland et al., 1992; Miller and
28 Friedland, 1994; Johnson et al., 1995a; Wang and Benoit, 1997). Overall, a significant amount
29 of research has been published on the distribution, speciation, and behavior of anthropogenic Pb
30 in the terrestrial environment since 1986. However, certain specific details on the behavior of Pb
31 in the terrestrial environment and its potential effects on soil microorganisms remain elusive.
May 2006 AX8-25 DRAFT-DO NOT QUOTE OR CITE
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1 AX8.1.2.1 Speciation of Atmospherically-Delivered Lead in Terrestrial Ecosystems
2 Lead in the Solid Phases
3 Lead can enter terrestrial ecosystems through natural rock weathering and by a variety of
4 anthropogenic pathways. These different source terms control the species of Pb that is
5 introduced into the terrestrial environment. While Pb is highly concentrated (percent level) in
6 certain hydrothermal sulfide deposits (e.g., PbS) that are disseminated throughout parts of the
7 upper crust, these occurrences are relatively rare. Therefore, the occurrence of Pb as a minor
8 constituent of rocks (ppm level), particularly granites, rhyelites, and argillaceous sedimentary
9 rocks is the more pertinent source term for the vast majority of terrestrial ecosystems. During
10 the hydrolysis and oxidation of Pb-containing minerals, divalent Pb is released to the soil
11 solution where it is rapidly fixed by organic matter and secondary mineral phases (Kabata-
12 Pendias and Pendias, 1992). The geochemical form of natural Pb in terrestrial ecosystems will
13 be strongly controlled by soil type (Emmanuel and Erel, 2002). In contrast, anthropogenically
14 introduced Pb has a variety of different geochemical forms, depending on the specific source.
15 While Pb in soils from battery reclamation areas can be in the form of PbSC>4 or PbSiOs, Pb in
16 soils from shooting ranges and paint spills is commonly found as PbO and a variety of Pb
17 carbonates (Vantelon et al., 2005; Laperche et al., 1996; Manceau et al., 1996). Atmospherically
18 delivered Pb resulting from fossil fuel combustion is typically introduced into terrestrial
19 ecosystems as Pb-sulfur compounds and Pb oxides (Olson and Skogerboe, 1975; Clevenger et
20 al., 1991; Utsunomiya et al., 2004). After deposition, Pb species are likely transformed.
21 Although the specific factors that control the speciation of anthropogenic Pb speciation in soils
22 are not well understood, there are many studies that have partitioned Pb into its different
23 geochemical phases. A thorough understanding of Pb speciation is critical in order to predict
24 potential mobility and bioavailability (see Section AX8.1.1).
25 Selective chemical extractions have been employed extensively over the past 20 years for
26 quantifying amounts of a particular metal phase (e.g., PbS, Pb-humate, Pb-Fe/Mn-oxide) present
27 in soil or sediment rather than total metal concentration. Sometimes selective chemical
28 extractions are applied sequentially to a particular sample. For example, the exchangeable metal
29 fraction is removed from the soil using a weak acid or salt solution (e.g., BaCb), followed
30 immediately by an extraction targeting organic matter (e.g., H2O2 or NaOCl), further followed by
31 an extraction targeting secondary iron oxides (e.g., NH2OH-HC1), and finally, a strong reagent
May 2006 AX8-26 DRAFT-DO NOT QUOTE OR CITE
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1 cocktail (e.g., HMVHCl-HF) targets primary minerals. Tessier et al. (1979) developed this
2 technique. More recently, this technique has been modified and developed specifically for
3 different metals and different types of materials (Keon et al., 2001). Alternatively, batch-style
4 selective chemical extractions have been used on soils and sediments to avoid the problems
5 associated with nonselective reagents (Johnson and Petras, 1998). Selective extractions can be a
6 relatively rapid, simple, and inexpensive means for determining metal phases in soils and
7 sediments, and the generated data can be linked to potential mobility and bioavailability of the
8 metal (Tessier and Campbell, 1987). However, some problems persist with the selective
9 extraction technique. First, extractions are rarely specific to a single phase. For example, while
10 H2O2 is often used to remove metals bound to organic matter in soils, others have demonstrated
11 that this reagent destroys clay minerals and sulfides (Ryan et al., 2002). Peroxide solutions may
12 also be inefficient in removing metals bound to humic acids, and in fact could potentially result
13 in the precipitation of metal-humate substances. In addition to the nonselectivity of reagents,
14 significant metal redistribution has been documented to occur during sequential chemical
15 extractions (Ho and Evans, 2000; Sulkowski and Hirner, 2006), and many reagents may not
16 completely extract targeted phases. While chemical extractions provide some useful information
17 on metal phases in soil or sediment, the results should be treated as "operationally defined," e.g.,
18 "H2O2-liberated Pb" rather than "organic Pb."
19 Lead forms strong coordination complexes with oxygen on mineral surfaces and organic
20 matter functional groups (Abd-Elfattah and Wada, 1981), because of its high electronegativity
21 and hydrolysis constant. Therefore, Pb is generally not readily exchangeable, i.e., the amount of
22 Pb removed from soils by dilute acid or salts is usually less that 10% (Karamanos et al., 1976;
23 Sposito et al., 1982; Miller and McFee, 1983; Johnson and Petras, 1998; Bacon and Hewitt,
24 2005). Lead is typically adsorbed to organic and inorganic soil particles strongly via inner-
25 sphere adsorption (Xia et al., 1997; Bargar et al., 1997a,b, 1998). Kaste et al. (2005) found that a
26 single extract of 0.02 M HC1 removed 15% or less Pb in organic horizons from a montane forest
27 in New Hampshire. The fact that relatively concentrated acids, reducing agents, oxidizing
28 agents, or chelating agents are required to liberate the majority of Pb from soils is used as one
29 line of evidence that Pb migration and uptake by plants in soils is expected to be low.
30 Lead that is "organically bound" in soils is typically quantified by extractions that
31 dissolve/disperse or destroy organic matter. The former approach often employs an alkaline
May 2006 AX8-27 DRAFT-DO NOT QUOTE OR CITE
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1 solution (NaOH), which deprotonates organic matter functional groups, or a phosphate solution,
2 which chelates structural cations. Extractions used to destroy organic matter often rely on H2O2
3 or NaOCl. Both organic and mineral horizons typically have significant Pb in this soil phase.
4 Miller and McFee (1983) used Na4P2O? to extract organically bound Pb from the upper 2.5 cm of
5 soils sampled from northwestern Indiana. They found that organically bound Pb accounted for
6 between 25 and 50% of the total Pb present in the sampled topsoils. Jersak et al. (1997), Johnson
7 and Petras (1998), and Kaste et al. (2005) selectively extracted Pb from spodosols from the
8 northeastern United States. Using acidified H2O2, Jersak et al. (1997) found that very little
9 (<10 %) of the Pb in mineral soils (E, B, C) sampled from New York and Vermont was organic.
10 Johnson and Petras (1998) used K4P2Oy to quantify organically bound Pb in the Oa horizon and
11 in mineral soils from the Hubbard Brook Experimental Forest in New Hampshire. They reported
12 that 60% of the total Pb in the Oa horizon was organic and that between 8 and 17% of the total
13 Pb in the mineral soil was organic. However, in the E, Bh, and Bsl horizons, organically bound
14 Pb dominated the total "labile" (non-mineral lattice) Pb. Kaste et al. (2005) used selective
15 chemical extractions on organic horizons from montane forests in Vermont and New Hampshire.
16 They found that repeated extractions with Na4P2O? removed between 60 and 100% of the Pb
17 from their samples. Caution should be used when interpreting the results of pyrophosphate
18 extractions. Although they are often used to quantify organically bound metals, this reagent can
19 both disperse and dissolve Fe phases (Jeanroy and Guillet, 1981; Shuman, 1982). Acidified
20 H2O2 has also been reported to destroy and release elements associated with secondary soil
21 minerals (Papp et al., 1991; Ryan et al., 2002).
22 Aside from organic forms, Pb is often found to be associated with secondary oxide
23 minerals in soils. Pb can be partitioned with secondary oxides by a variety of mechanisms,
24 including (1) simple ion exchange, (2) inner-sphere or outer-sphere adsorption, and (3) co-
25 precipitation and/or occlusion (Bargar et al., 1997a,b, 1998, 1999). As discussed above, very
26 little Pb is removed from soil via dilute acid or salt solutions, so adsorption and co-precipitation
27 are likely the dominant Pb interactions with secondary mineral phases. Reagents used to
28 quantify this phase are often solutions of EDTA, oxalate, or hydroxylamine hydrochloride (HH).
29 Miller and McFee (1983) used an EDTA solution followed by an HH solution to quantify Pb
30 occluded by Fe and Mn minerals, respectively, in their surface-soil samples from Indiana. They
31 reported that approximately 30% of the total soil Pb was occluded in Fe minerals, and 5 to 15%
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1 was occluded in Mn phases. In soils from the northeastern United States, Jersak et al. (1997)
2 used various strengths of HH solutions and concluded that negligible Pb was associated with
3 Mn-oxides and that 1 to 30% of the Pb was associated with Fe phases in the mineral soils in their
4 study. Johnson and Petras (1998) reported that no Pb was removed from the Oa horizon at the
5 Hubbard Brook Experimental Forest (FffiEF) by oxalate, but that 5 to 15% of the total Pb in
6 mineral soils was removed by this extraction, presumably because it was bound to amorphous
7 oxide minerals. Kaste et al. (2005), however, reported that HH removed 30 to 40% of the Pb
8 from organic horizons in their study. They concluded that Fe phases were important in
9 scavenging Pb, even in soil horizons dominated by organic matter.
10 Synchrotron radiation (X-rays) allows researchers to probe the electron configuration of
11 metals in untreated soil and sediment samples. This type of analysis has been extremely valuable
12 for directly determining the coordination environment of Pb in a variety of soils and sediments.
13 Since different elements have different electron binding energies (Eb), X-rays can be focused in
14 an energy window specific to a metal of interest. In experiments involving XAS, X-ray energy is
15 increased until a rapid increase in the amount of absorption occurs; this absorption edge
16 represents Eb. The precise energy required to dislodge a core electron from a metal (i.e., Eb) will
17 be a function of the oxidation state and covalency of the metal. X-ray absorption studies that
18 focus on the location of the absorption edge are referred to as XANES (X-ray absorption near
19 edge structure). In the energy region immediately after the absorption edge, X-ray absorption
20 increases and decreases with a periodicity that represents the wave functions of the ejected
21 electrons and the constructive and destructive interference with the wave functions of the nearby
22 atoms. X-ray absorption studies used to investigate the periodicity of the absorption after Eb are
23 referred to as EXAFS (extended X-ray absorption fine structure). Since the electron
24 configuration of a Pb atom will be directly governed by its speciation (e.g., Pb bound to organics,
25 Pb adsorbed to oxide surfaces, PbS, etc.) X-ray absorption studies provide a powerful in-situ
26 technique for determining speciation without some of the problems associated with chemical
27 extractions (Bargar et al., 1997a,b, 1998).
28 Manceau et al. (1996) used EXAFS to study soil contaminated by gasoline-derived Pb in
29 France and found that the Pb was divalent and complexed to salicylate and catechol-type
30 functional groups of humic substances. He concluded that the alkyl-tetravalent Pb compounds
31 that were added to gasoline were relatively unstable and will not dominate the speciation of Pb
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1 fallout from the combustion of leaded gasoline. The binding mechanism of Pb to organics is
2 primarily inner-sphere adsorption (Xia et al., 1997). DeVolder et al. (2003) used EXAFS to
3 demonstrate that Pb phases were shifting to the relatively insoluble PbS when contaminated
4 wetland soils were treated with sulfate. More recent XAS studies have demonstrated the
5 importance of biomineralization of Pb in soils by bacteria and nematodes (Xia et al., 1997;
6 Templeton et al., 2003a,b; Jackson et al., 2005). Templeton et al. (2003a,b) demonstrated that
7 biogenic precipitation of pyromorphite was the dominant source of Pb uptake by Burkholderia
8 cepacia biofilms below pH 4.5. Above pH 4.5, adsorption complexes began to form in addition
9 to Pb mineral precipitation.
10 In addition to XAS studies of Pb in environmental samples, numerous experimental-based
11 XAS studies have documented in detail the coordination environment of Pb adsorbed to Fe-
12 oxides, Mn-oxides, Al-oxides, and clay minerals (Manceau et al., 1996, 2000a,b, 2002; Bargar
13 et al., 1997a,b, 1998, 1999; Strawn and Sparks, 1999; Trivedi et al., 2003). Bargar et al. (1997a)
14 showed that Pb can adsorb to FeOe octahedra on three different types of sites: on corners, edges,
15 or faces. Ostergren et al. (2000a,b) showed that the presence of dissolved carbonate and sulfate
16 increased Pb adsorbtion on goethite. The relative fraction of corner-sharing complexes can be
17 greatly increased by the presence of these ligands, as bridging complexes between the metal and
18 the corners are formed (Ostergren et al., 2000a,b).
19 Recently, Jackson et al. (2005) used microfocused synchrotron-based X-ray fluorescence
20 (//SXRF) to detail the distribution of Pb and Cu in the nematode Caenorhabditis elegans. They
21 found that, while Cu was evenly distributed throughout the bodies of exposed C. elegans, Pb was
22 concentrated in the anterior pharynx region. Microfocused X-ray diffraction indicated that the
23 highly concentrated Pb region in the pharynx was actually comprised of the crystalline Pb
24 mineral, pyromorphite. The authors concluded that C. elegans precipitated pyromorphite in the
25 pharynx as a defense mechanism to prevent spreading the toxic metal to the rest of the
26 organism's body. They further suggested that, because of the high turnover rate of nematodes,
27 biomineralization could play an important role in the speciation of Pb in certain soils.
28
29 Lead Solid-Solution Partitioning
30 The concentration of Pb species dissolved in soil solution is probably controlled by some
31 combination of (a) Pb-mineral solubility equilibria, (b) adsorption reactions of dissolved Pb
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1 phases on inorganic surfaces (e.g., crystalline or amorphous oxides of Al, Fe, Si, Mn, etc.; clay
2 minerals), and (c) adsorption reactions of dissolved Pb phases on soil organic matter. Dissolved
3 Pb phases in soil solution can be some combination of Pb2+ and its hydrolysis species, Pb bound
4 to dissolved organic matter, and Pb complexes with inorganic ligands such as Cl and SO42 .
5 Alkaline soils typically have solutions supersaturated with respect to PbCOs, Pb3(CO3)2(OH)2,
6 Pb(OH)2, Pb3(PO4)2, Pb5(PO4)3(OH), and Pb4O(PO4)2 (Badawy et al., 2002). Pb-phosphate
7 minerals in particular are very insoluble, and calculations based on thermodynamic data predict
8 that these phases will control dissolved Pb in soil solution under a variety of conditions (Nriagu,
9 1974; Ruby et al., 1994). However, certain chelating agents, such as dissolved organic matter,
10 can prevent the precipitation of Pb minerals (Lang and Kaupenjohann, 2003).
11 Using a combination of desorption experiments and XAS, EXAFS, and XANES, Rouff et
12 al. (2006) found that aging of Pb-calcite suspensions resulted in changes in the solid-phase
13 distribution of Pb. Increased sorption time can reduce trace metal desorption by enhancing the
14 stability of surface complexes (Rouff et al., 2005) or by mechanisms involving microporous
15 diffusion (Backes et al., 1995), recrystallization-induced incorporation into the solid phase
16 (Ainsworth et al., 1994), and formation and stabilization of surface precipitates (Ford et al.,
17 1999). For adsorption of Pb to hydrous Fe oxides, goethite, and Pb-contaminated soils, aged
18 samples show Pb to be reversibly bound, suggesting Pb adsorption primarily to the substrates'
19 surfaces (Rouff et al., 2006). However, for Pb adsorption to calcite aging played a significant
20 role due to detection of multiple sorption mechanisms, even for short sorption times (Rouff et al.,
21 2006). pH also played a role in Pb sorption. Over long sorption periods (60 to 270 days), slow
22 continuous uptake of Pb occurred at pH 7.3 and 8.2. At pH 9.4, no further uptake occurred with
23 aging and very little desorption occurred. These results show the importance of contact time and
24 pH on Pb solid-phase partitioning, particularly in geochemical systems in which calcite may be
25 the predominant mineralogical constituent.
26 Soil solution dissolved organic matter content and pH typically have very strong positive
27 and negative correlations, respectively, with the concentration of dissolved Pb species (Sauve
28 et al., 1998, 2000a, 2003; Weng et al., 2002; Badawy et al., 2002; Tipping et al., 2003). In the
29 case of adsorption phenomena, the partitioning of Pb2+ to the solid phase is also controlled by
30 total metal loading, i.e., high Pb loadings will result in a lower fraction being partitioned to the
31 solid phase. Sauve et al. (1997, 1998) demonstrated that only a fraction of the total Pb in
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1 solution was actually Pb2+ in soils treated with leaf compost. The fraction of Pb2+ to total
2 dissolved Pb ranged from <1 to 60%, depending on pH and the availability of Pb-binding
3 ligands. Nolan et al. (2003) used Donnan dialysis to show that 2.9 to 48.8% of the dissolved Pb
4 was Pb2+ in pore waters of agricultural and contaminated soils from Australia and the United
5 States. In acidic soils, Al species can compete for sites on natural organic matter and inhibit Pb
6 binding to surfaces (Gustafsson et al., 2003).
7 Differential pulse anodic stripping voltammetry (DPASV) is a technique that is useful for
8 identifying relatively low concentrations of Pb2+ and has found many applications in adsorption
9 and partitioning experiments. This technique has been particularly useful for quantifying the Kd,
10 or partitioning ratio of Pb in the solid-to-liquid phase (Kd = [total solid-phase metal in mg kg l] I
11 [dissolved metal in mg I/1]). While the exact Kd value is a function of pH, organic matter
12 content, substrate type, total metal burden, and concentrations of competing ligands, such studies
13 typically show that Pb has very strong solid-phase partitioning. Partitioning ratios determined by
14 DPASV generally range from 103 to 106 in soils in the typical pH range (Sauve et al., 2000b).
15 Aualiitia and Pickering (1987) used thin film ASV to compare the relative affinity of Pb for
16 different inorganic particulates. They reported that Mn(IV) oxides completely adsorbed the Pb,
17 regardless of pH in the range of 3 to 9, and had the highest affinity for Pb in their study. The
18 adsorption of Pb to pedogenic Fe-oxides, Al-hydroxides, clay minerals, and Fe ores was reported
19 to be pH-dependent. Sauve et al. (1998) used DPASV to study the effects of organic matter and
20 pH on Pb adsorption in an orchard soil. They demonstrated that Pb complexation to dissolved
21 organic matter (DOM) increased Pb solubility, and that 30 to 50% of the dissolved Pb was bound
22 to DOM at pH 3 to 4, while >80% of the dissolved Pb was bound to DOM at neutral pH. They
23 concluded that in most soils, Pb in solution would not be found as Pb2+ but as bound to DOM.
24 Sauve et al. (2000a) compared the relative affinity of Pb2+ for synthetic ferrihydrite, leaf
25 compost, and secondary oxide minerals collected from soils. They reported that the inorganic
26 mineral phases were more efficient at lowering the amount of Pb2+ that was available in solution
27 than the leaf compost. Glover et al. (2002) used DPASV in studying the effects of time and
28 organic acids on Pb adsorption to goethite. They found that Pb adsorption to geothite was very
29 rapid, and remained unchanged after a period of about 4 h. Lead desorption was found to be
30 much slower. The presence of salicylate appeared to increase the amount of Pb that desorbed
31 from goethite more so than oxalate.
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1 AX8.1.2.2 Tracing the Fate of Atmospherically Delivered Lead in Terrestrial
2 Ecosystems
3 Radiogenic Pb isotopes offer a powerful tool for separating anthropogenic Pb from natural
4 Pb derived from mineral weathering (Erel and Patterson, 1994; Erel et al., 1997). This is
5 particularly useful for studying Pb in mineral soil, where geogenic Pb often dominates. The
6 three radiogenic stable Pb isotopes (206Pb, 207Pb, and 208Pb) have a heterogeneous distribution in
7 the earth's crust primarily because of the differences in the half-lives of their respective parents
8 (238U, Ti/2 = 4.7 x 109 year; 235U, Ti/2 = 0.7 x 109 year; 232Th, Ti/2 = 14 x 109 year). The result is
9 that the ore bodies from which anthropogenic Pb are typically derived are usually enriched in
10 207Pb relative to 206Pb and 208Pb when compared with Pb found in granitic rocks. Graney et al.
11 (1995) analyzed a dated core from Lake Erie, and found that the 206Pb/207Pb value in sediment
12 deposited in the late 1700s was 1.224, but in 20th-century sediment, the ratio ranged from 1.223
13 to 1.197. This shift in the Pb isotopic composition represents the introduction of a significant
14 amount of anthropogenic Pb into the environment. Bindler et al. (1999) and Emmanuel and Erel
15 (2002) analyzed the isotopic composition of Pb in soil profiles in Sweden and the Czech
16 Republic, respectively, and determined that mineral soils immediately below the organic horizon
17 had a mixture of both anthropogenic and geogenic Pb.
18 Erel and Patterson (1994) used radiogenic Pb isotopes to trace the movement of industrial
19 Pb from topsoils to groundwaters to streams in a remote mountainous region of Yosemite
20 National Park in California. They calculated that total 20th-century industrial Pb input to their
21 study site was approximately 0.4 g Pb m 2. Lead concentrations in organic material were highest
22 in the upper soil horizons, and decreased with depth. During snowmelt, Pb in the snowpack was
23 mixed with the anthropogenic and geogenic Pb already in the topsoil, and spring melts contained
24 a mixture of anthropogenic and geogenic particulate Pb. During base flows, however, 80% of
25 the Pb export from groundwater and streams was from natural granite weathering (Erel and
26 Patterson, 1994).
27 Uranium-238 series 210Pb also provides a tool for tracing atmospherically delivered Pb in
28 soils. After 222Rn (Ti/2 = 3.8 days) is produced from the decay of 226Ra (Ti/2 = 1600 years), some
29 fraction of the 222Rn escapes from rocks and soils to the atmosphere. It then decays relatively
30 rapidly to 210Pb (Ti/2 = 22.3 years), which has a tropospheric residence time of a few weeks
31 (Koch et al., 1996). Fallout 210Pb is deposited onto forests via wet and dry deposition, similar to
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1 anthropogenic Pb deposition in forests, and is thus useful as a tracer for non-native Pb in soils.
2 Lead-210 is convenient to use for calculating the residence time of Pb in soil layers, because its
3 atmospheric and soil fluxes can be assumed to be in steady state at undisturbed sites (Dorr and
4 Munnich, 1989; Dorr, 1995; Kaste et al., 2003). Atmospheric 210Pb (210Pbex hereafter,210Pb in
5 "excess" of that supported by 222Rn in the soil) must be calculated by subtracting the amount of
6 210Pb formed in soils by the in-situ decay of 222Rn from the total 210Pb (Moore and Poet, 1976;
7 Nozakietal., 1978).
8 Benninger et al. (1975) measured fallout 210Pb in soils and streamwater at Hubbard Brook
9 and at an undisturbed forest in Pennsylvania. They estimated atmospheric 210Pb export in
10 streamwaters to be <0.02% of the standing 210Pb crop in the organic horizons. They used a
11 simple steady-state model to calculate the residence time of Pb in the organic horizons to be
12 5,000 years. This overestimate of the Pb residence time in the organic horizons was likely a
13 result of the low resolution of their sampling. Since they only sampled the upper 6 cm of soil
14 and the drainage waters, they did not accurately evaluate the distribution of 210Pb in the soil
15 column in between. Dorr and Munnich (1989, 1991) used 210Pb profiles in soils of southern
16 Germany to evaluate the behavior of atmospherically delivered Pb. They calculated the vertical
17 velocity of Pb by dividing the relaxation depth (i.e., the depth at which 210Pb activity decreases to
18 1/e, or approximately 37% of its surface value) by the 210Pb mean life of 32 years. They reported
19 downward transit velocities of atmospherically deposited Pb at 0.89 ± 0.33 mm year l. The
20 downward transport of atmospheric Pb was not affected by pH or soil type. However, since Pb
21 velocities in the soil profile where identical to carbon velocities calculated with 14C, they
22 concluded that Pb movement in forest soils is probably controlled by carbon transport. Kaste
23 et al. (2003) used 210Pb to model the response time of atmospherically delivered Pb in the O
24 horizon at Camel's Hump Mountain in Vermont. They concluded that the forest floor response
25 time was between 60 and 150 years, depending on vegetation zone and elevation. Using
26 206Pb:207Pb, they also demonstrated that some gasoline-derived Pb migrated out of the O horizon
27 and into the mineral soil in the deciduous vegetation zone on the mountain, while all of the
28 atmospheric Pb was retained in the upper 20 cm of the soil profile.
29 Researchers assessing the fate of atmospheric Pb in soils have also relied on repeated
30 sampling of soils and vegetation for total Pb. This technique works best when anthropogenic Pb
31 accounts for the vast majority of total Pb in a particular reservoir. Johnson et al. (1995a), Yanai
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1 et al. (2004), and Friedland et al. (1992) used O horizon (forest floor) time-series data to evaluate
2 the movement of gasoline-derived Pb in the soil profile. These studies have concluded that the
3 distribution of Pb in the upper soil horizons has changed over the past few decades. Yanai et al.
4 (2004) documented a decline in Pb from the Oie horizon between the late 1970s to the early
5 1990s in remote forest soils in New Hampshire. Johnson et al. (1995a) and Friedland et al.
6 (1992) demonstrated that some fraction of Pb had moved from the O horizon to the mineral soil
7 during the 1980s at Hubbard Brook and at selected remote sites in the northeastern United States,
8 respectively. Evans et al. (2005) demonstrated that Pb concentrations in the litter layer (fresh
9 litter + Oi horizon) sampled in a transect from Vermont to Quebec decreased significantly
10 between 1979 and 1996, reflecting a decrease in Pb deposition to forests and upper soil horizons
11 during that time period. Miller et al. (1993) and Wang and Benoit (1997) used forest floor time-
12 series data to model the response time (e folding time, the time it takes a reservoir to decrease to
13 the l/e, (ca. 37%) of its original amount) of Pb in the forest floor. Miller et al. (1993) calculated
14 O horizon response times of 17 years for the northern hardwood forest and 77 years in the
15 spruce-fir zone on Camel's Hump Mountain in Vermont. Wang and Benoit (1997) determined
16 that the O horizon would reach steady state with respect to Pb (1.3 jig g l Pb) by 2100. Both
17 suggested that the movement of organic particulates dominated Pb transport in the soil profile.
18
19 AX8.1.2.3 Inputs/Outputs of Atmospherically Delivered Lead in Terrestrial Ecosystems
20 The concentration of Pb in contemporary rainfall in the mid-Atlantic and northeastern
21 United States is on the order of 500 to 1000 pg g"1 (Wang et al., 1995; Kim et al., 2000; Scudlark
22 et al., 2005). For comparison, rainfall measured in Los Angeles (CA) during 2003-2004
23 averaged 150 pg g l, but showed nearly an order-of-magnitude variation, presumably because of
24 the arid environmental (Sabin et al., 2005). The role of dry deposition in the total deposition of
25 Pb to terrestrial ecosystems is not constrained well. Researchers have estimated that dry
26 deposition accounts for anywhere between 10 to >90% of total Pb deposition (Galloway et al.,
27 1982; Wu et al., 1994; Sabin et al., 2005). Arid environments appear to have a much higher
28 fraction of dry deposition:total deposition (Sabin et al., 2005). Furthermore, it is possible that
29 Clean Air Act Legislation enacted in the late 1970s preferentially reduced Pb associated with
30 fine particles, so the relative contributions of dry deposition may have changed in the last few
31 decades. If the major source of Pb to a terrestrial ecosystem is resuspended parti culates from
May 2006 AX8-3 5 DRAFT-DO NOT QUOTE OR CITE
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1 transportation corridors, then the particle size fraction that dominates deposition may be
2 relatively coarse (>50 jim) relative to other atmospheric sources (Pirrone et al., 1995; Sansalone
3 et al., 2003).
4 Total contemporary loadings to terrestrial ecosystems are approximately 1 to 2 mg m 2
5 yeafl (Wu et al., 1994; Wang et al., 1995; Sabin et al., 2005). This is a relatively small annual
6 flux of Pb if compared to the reservoir of approximately 0.5 to 4 g m 2 of gasoline-derived Pb
7 that is already in surface soils over much of the United States (Friedland et al., 1992; Miller and
8 Friedland, 1994; Erel and Patterson, 1994; Marsh and Siccama, 1997; Yanai et al., 2004;
9 Johnson et al., 2004; Evans et al., 2005). While vegetation can play an important role in
10 sequestering Pb from rain and dry deposition (Russell et al., 1981), direct uptake of Pb from soils
11 by plants appears to be low (Klaminder et al., 2005). High elevation areas, particularly those
12 near the base level of clouds often have higher burdens of atmospheric contaminants (Siccama,
13 1974). A Pb deposition model by Miller and Friedland (1994) predicted 2.2 and 3.5 g Pb m"2
14 deposition for the 20th century in the deciduous zone (600 m) and the coniferous zone (1000 m),
15 respectively. More recently, Kaste et al. (2003) used radiogenic isotope measurements on the
16 same mountain to confirm higher loadings at higher elevation. They measured 1.3 and 3.4 g
17 gasoline-derived Pb m 2 in the deciduous zone and coniferous zones, respectively. Higher
18 atmospheric Pb loadings to higher elevations are attributed to (1) the higher leaf area of
19 coniferous species, which are generally more prevalent at high elevation; (2) higher rainfall at
20 higher elevation; and (3) increased cloudwater impaction at high elevation (Miller et al., 1993).
21 Although inputs of Pb to ecosystems are currently low, Pb export from watersheds via
22 groundwater and streams is substantially lower. Therefore, even at current input levels,
23 watersheds are accumulating industrial Pb. Seeps and streams at the HBEF have Pb
24 concentrations on the order of 10 to 30 pg Pb g l (Wang et al., 1995). At a remote valley in the
25 Sierra Nevada, Pb concentrations in streamwaters were on the order of 15 pg Pb g l (Erel and
26 Patterson, 1994). Losses of Pb from soil horizons are assumed to be via particulates (Dorr and
27 Munnich, 1989; Wang and Benoit, 1996, 1997). Tyler (1981) noted that Pb losses from a
28 horizon in Sweden were influenced by season; with highest Pb fluxes being observed during
29 warm, wet months. He suggested that DOC production and Pb movement were tightly linked.
30 Surface soils across the United States are enriched in Pb relative to levels expected from
31 solely natural geogenic inputs (Friedland et al., 1984; Francek, 1992; Erel and Patterson, 1994;
May 2006 AX8-36 DRAFT-DO NOT QUOTE OR CITE
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1 Marsh and Siccama, 1997; Yanai et al., 2004; Murray et al., 2004). While some of this
2 contaminant Pb is attributed to paint, salvage yards, shooting ranges, and the use of Pb arsenate
3 as a pesticide in localized areas (Francek, 1997), Pb contamination of surface soils is essentially
4 ubiquitous because of atmospheric pollution associated with the combustion of fossil fuels, waste
5 incineration, and metal smelting and production (Newhook et al., 2003; Polissar et al., 2001).
6 Surface soils in Michigan, for example, typically range from 8 to several hundred ppm Pb
7 (Francek, 1992; Murray et al., 2004). Soils collected and analyzed beneath 50 cm in Michigan,
8 however, range only from 4 to 60 ppm Pb (Murray et al., 2004). In remote surface soils from the
9 Sierra Nevada Mountains, litter and upper soil horizons are 20 to 40 ppm Pb, and approximately
10 75% of this Pb has been attributed to atmospheric deposition during the 20th century (Erel and
11 Patterson, 1994). Repeated sampling of the forest floor (O horizon) in the northeastern United
12 States demonstrates that the organic layer has retained much of the Pb load deposited on
13 ecosytems during the 20th century. Total Pb deposition during the 20th century has been
14 estimated at 1 to 3 g Pb m 2, depending on elevation and proximity to urban areas (Miller and
15 Friedland, 1994; Johnson et al., 1995a). Forest floors sampled during the 1980s and 1990s, and
16 in early 2000 had between 0.7 and 2 g Pb m"2 (Friedland et al., 1992; Miller and Friedland, 1994;
17 Johnson et al., 1995a; Kaste et al., 2003; Yanai et al., 2004; Evans et al., 2005). The pool of Pb
18 in above- and below-ground biomass at the HBEF is on the order of 0.13 g Pb m 2 (Johnson
19 etal., 1995a).
20 The amount of Pb that has leached into mineral soil appears to be on the order of 20 to
21 50% of the total anthropogenic Pb deposition. Kaste et al. (2003) and Miller and Friedland
22 (1994) demonstrated that Pb loss from the forest floor at Camel's Hump Mountain in Vermont
23 depended on elevation. While the mineral soil in the deciduous forest had between 0.4 and 0.5 g
24 Pb m 2 (out of 1 to 2 g Pb m 2 in the total soil profile), at higher elevations the thicker coniferous
25 forest floor retained more than 90% of the total Pb deposition (Kaste et al., 2003). Johnson et al.
26 (1995a) determined that the forest floor at FfflEF in the mid-1980s had about 0.75 g Pb m"2.
27 Compared to their estimated 20th-century atmospheric Pb deposition of 0.9 g Pb m 2, the forest
28 floor has retained 83% of the atmospheric Pb loadings (Johnson et al., 1995a). Johnson et al.
29 (2004) noted that gasoline-derived Pb was a significant component of the labile Pb at the HBEF.
30 They calculated that Pb fluxes to the HBEF by atmospheric pollution were essentially equivalent
31 to the Pb released by mineral weathering over the past 12,000 years. Marsh and Siccama (1997)
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1 used the relatively homogenous mineral soils underneath formerly plowed land in New
2 Hampshire, Connecticut, and Rhode Island to assess the depth-distribution of atmospheric Pb.
3 They reported that 65% of the atmospheric Pb deposited during the 20th century is in the mineral
4 soil and 35% is in the forest floor. At their remote study site in the Sierra Nevada Mountains,
5 Erel and Patterson (1994) reported that most of the anthropogenic Pb was associated with the
6 humus fraction of the litter layer and soils sampled in the upper few cm.
7 Atmospherically delivered Pb is probably present in ecosystems in a variety of different
8 biogeochemical phases. A combination of Pb adsorbtion processes and the precipitation of Pb
9 minerals will typically keep dissolved Pb species low in soil solution, surface waters, and
10 streams (Sauve et al., 2000a; Jackson et al., 2005). While experimental and theoretical evidence
11 suggest that the precipitation of inorganic Pb phases and the adsorption of Pb on inorganic
12 phases can control the biogeochemistry of contaminant Pb (Nriagu, 1974; Ruby et al., 1994;
13 Jackson et al., 2005), the influence of organic matter on the biogeochemistry of Pb in terrestrial
14 ecosystems cannot be ignored in many systems. Organic matter can bind to Pb, preventing Pb
15 migration and the precipitation of inorganic phases (Manceau et al., 1996; Xia et al., 1997; Lang
16 and Kaupenjohann, 2003). As the abundance of organic matter declines in soil, Pb adsorption to
17 inorganic soil minerals and the direct precipitation of Pb phases may dominate the
18 biogeochemistry of Pb in terrestrial ecosystems (Ostergren et al., 2000a,b; Sauve et al., 2000a).
19
20 Conclusions
21 Advances in technology since the 1986 Pb AQCD have allowed for a quantitative
22 determination of the mobility, distribution, uptake, and fluxes of atmospherically delivered Pb in
23 ecosystems. Among other things, these studies have shown that industrial Pb represents a
24 significant fraction of total labile Pb in watersheds. Selective chemical extractions and
25 synchrotron-based X-ray studies have shown that industrial Pb can be strongly sequestered by
26 organic matter and by secondary minerals such as clays and oxides of Al, Fe, and Mn. Some of
27 these studies have provided compelling evidence that the biomineralization of Pb phosphates by
28 soil organisms can play an important role in the biogeochemistry of Pb. Surface soils sampled
29 relatively recently demonstrate that the upper soil horizons (O + A horizons) are retaining most
30 of the industrial Pb burden introduced to the systems during the 20th century. The migration and
31 biological uptake of Pb in ecosystems is relatively low. The different biogeochemical behaviors
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1 of Pb reported by various studies may be a result of the many different analytical techniques
2 employed, or they may be a result of natural variability in the behavior of Pb in different
3 systems.
4
5 AX8.1.3 Terrestrial Species Response/Mode of Action
6 The 1986 Pb AQCD (U.S. Environmental Protection Agency, 1986a) reviewed the
7 literature on the uptake of Pb into plants, soil organisms, birds, and mammals. This chapter
8 expands upon the major conclusions from the EPA (U.S. Environmental Protection Agency,
9 1986a) related to those organisms. It summarizes the recent (since 1986) critical research
10 conducted on Pb uptake into terrestrial organisms (Section AX8.1.3.1), mechanisms of resistance
11 to Pb toxicity (Section AX8.1.3.2), the physiological effects of Pb (Section AX8.1.3.3), and, the
12 factors that modify organism response to Pb (Section AX8.1.3.4). A summary is presented in
13 Section AX8.1.3.5. All concentrations are expressed as mg Pb/kg dw (dry weight) unless
14 otherwise indicated.
15 Areas of research that are not addressed include those that used irrelevant exposure
16 conditions relative to airborne emissions of Pb (e.g., Pb shot, Pb paint, injection studies, studies
17 conducted on mine tailings or using hyperaccumulator plants for phytoremediation, and studies
18 conducted with hydroponic solutions) except when these studies provided critical information for
19 understanding physiologic effects.
20
21 AX8.1.3.1 Lead Uptake
22 Since the 1986 Pb AQCD (U.S. Environmental Protection Agency, 1986a), there have
23 been several studies that evaluated the uptake of Pb into plants and invertebrates. The
24 mechanisms associated with Pb uptake and translocation are described in this section. The
25 methods used by the EPA (U.S. Environmental Protection Agency, 2005a) to estimate Pb uptake
26 into plants, earthworms, and small mammals as part of Ecological Soil Screening Level (Eco-
27 SSL) development are also presented.
28 The accumulation of Pb into the various tissues of consumers (birds and mammals) is
29 discussed only when it was described relative to either environmental concentrations or
30 organismal effects. Numerous other monitoring studies measuring only the Pb concentrations in
31 various tissues of birds and mammals were not included in this chapter; their data cannot be used
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1 to develop an air quality standard without accompanying information on environmental
2 concentrations or organismal effects.
3
4 Lead Uptake into Plants
5 Plants take up Pb via their foliage and through their root systems (U.S. Environmental
6 Protection Agency, 1986a; Pahlsson, 1989). Surface deposition of Pb onto plants may represent
7 a significant contribution to the total Pb in and on the plant, as has been observed for plants near
8 smelters and along roadsides (U.S. Environmental Protection Agency, 1986a). The importance
9 of atmospheric deposition on above-ground plant Pb uptake is well-documented (Dalenberg and
10 Van Driel, 1990; Jones and Johnston, 1991; Angel ova et al., 2004). Data examined from
11 experimental grassland plots in southeast England demonstrated that atmospheric Pb is a greater
12 contributor than soil-derived Pb in crop plants and grasses (Jones and Johnston, 1991). A study
13 by Dalenberg and Van Driel (1990) showed that 75 to 95% of the Pb found in field-grown test
14 plants (i.e., the leafy material of grass, spinach, and carrot; wheat grain; and straw) was from
15 atmospheric deposition. Angelova et al. (2004) found that tobacco grown in an industrial area
16 accumulated significant amounts of Pb from the atmosphere, although uptake from soil was also
17 observed. The concentration of Pb in tobacco seeds was linearly related to the concentration of
18 Pb in the exchangeable and carbonate-bound fractions of soil, as measured using sequential
19 extraction (Angelova et al., 2004). Lead in soil is more significant when considering uptake into
20 root vegetables (e.g., carrot, potato), since, as was noted in the 1986 Pb AQCD (U.S.
21 Environmental Protection Agency, 1986a), most Pb remains in the roots of plants.
22 There are two possible mechanisms (symplastic or apoplastic) by which Pb may enter the
23 root of a plant. The symplastic route is through the cell membranes of root hairs; this is the
24 mechanism of uptake for water and nutrients. The apoplastic route is an extracellular route
25 between epidermal cells into the intercellular spaces of the root cortex. Previously, Pb was
26 thought to enter the plant via the symplastic route, probably by transport mechanisms similar to
27 those involved in the uptake of calcium or other divalent cations (i.e., transpirational mass flow,
28 diffusion, or active transport). However, it also had been speculated that Pb may enter the plant
29 via the apoplastic route (U.S. Environmental Protection Agency, 1986a). Sieghardt (1990)
30 determined that the mechanism of Pb uptake was via the symplastic route only and that the
31 apoplastic pathway of transport was stopped in the primary roots by the endodermis. He studied
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1 the uptake of Pb into two plants, Minuartia verna (moss sandwort) and Silene vulgaris (bladder
2 campion) that colonize metal-contaminated sites. In the roots of both plants, Pb was found
3 mainly in the root cortex. Active ion uptake was required to transport the Pb into the stele and
4 then into the shoots of the plant (Sieghardt, 1990).
5 Although some plants translocate more Pb to the shoots than others, most Pb remains in
6 the roots of plants. Two mechanisms have been proposed to account for this relative lack of
7 translocation to the shoots: (1) Pb may be deposited within root cell wall material, or (2) Pb may
8 be sequestered within root cell organelles (U.S. Environmental Protection Agency, 1986a).
9 Pahlsson (1989) noted that plants can accumulate large quantities of Pb from the soil but that
10 translocation to shoots and leaves is limited by the binding of Pb ions at root surfaces and cell
11 walls. In a study by Wierzbicka (1999), 21 different plant species were exposed to Pb2+ in the
12 form of Pb-chloride. The plant species included cucumber (Cucumis sativus), soy bean (Soja
13 hispida), bean (Phaseolus vulgaris), rapeseed (Brassica napus), rye (Secale cereale), barley
14 (Hordeum vulgar e), wheat (Triticum vulgar e), radish (Raphanus sativus), pea (Pisum sativum),
15 maize (Zea mays), onion (Allium cepa), lupine (Lupinus luteus), bladder campion (Silene
16 vulgaris)., Buckler mustard (Biscutella laevigata), and rough hawkbit (Leontodon hispidus).
17 Although, the amount of Pb taken up by the plant varied with species, over 90% of absorbed Pb
18 was retained in the roots. Only a small amount of Pb was translocated (~2 to 4%) to the shoots
19 of the plants. Lead in roots was present in the deeper layers of root tissues (in particular, the root
20 cortex) and not only on the root surface. There was no correlation between Pb tolerance
21 (measured as root mass increase expressed as a percentage of controls) and either root or shoot
22 tissue concentrations (Wierzbicka, 1999). The study by Wierzbicka (1999) was the first to report
23 that plants developing from bulbs, in this case the onion, were more tolerant to Pb than plants
24 developing from seeds. This tolerance was assumed to be related to the large amounts of Pb that
25 were transported from the roots and stored in the bulb of the plant (Wierzbicka, 1999).
26 Uptake of Pb from soil into plants was modeled as part of Eco-SSL development (U.S.
27 Environmental Protection Agency, 2005a). The relationship derived between Pb in the soil and
28 Pb in a plant was taken from Bechtel Jacobs Company (BJC) (1998) and is as follows:
29
31 Ln(Cp) = 0.561 * Ln(Csoil) - 1.328 (8-1)
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1 where Cp is the concentration of Pb in the plant (dry weight) and Csoil is the concentration of Pb
2 in the soil. This equation recognizes that the ratio of Pb concentration in plant to Pb
3 concentration in soil is not constant.
4
5 Invertebrates
6 There was no clear evidence suggesting a differential uptake of Pb into different species
7 of earthworm (Lumbricus terrestris, Aporrectodea rosea, and A. caliginosd) collected around a
8 smelter site near Avonmouth, England (Spurgeon and Hopkin, 1996a). This is in contrast to Pizl
9 and Josens (1995) and Terhivuo et al. (1994) who found Aporrectodea spp. accumulated more
10 Pb than Lumbricus. The authors suggested that these differences could be due to different
11 feeding behaviors, as Lumbricus feeds on organic material and Apporectodea species are
12 geophagus, ingesting large amounts of soil during feeding. The differences between species also
13 may be related to differing efficiencies in excretory mechanisms (Pizl and Josens, 1995).
14 However, the interpretation of species difference is complicated by a number of potentially
15 confounding variables, such as soil characteristics (e.g., calcium or other nutrient levels)
16 (Pizl and Josens, 1995).
17 The bioaccumulation of Pb from contaminated soil was tested using the earthworm
18 Eiseniafetida, and the amount of Pb accumulated did not change significantly until the
19 concentration within soil reached 5000 mg/kg (Davies et al., 2003). This coincided with the
20 lowest soil concentrations at which earthworm mortality was observed. The ratio of the
21 concentration of Pb in worms to the concentration in soil decreased from 0.03 at 100 mg/kg to
22 0.001 at 3000 mg/kg, but then increased quickly to 0.02 at 5000 mg/kg. The authors concluded
23 that earthworms exhibit regulated uptake of Pb at levels of low contamination (<3000 mg/kg)
24 until a critical concentration is reached, at which point this mechanism breaks down, resulting in
25 unregulated accumulation and mortality. This study was conducted using test methods where
26 soil was not allowed to equilibrate following the addition of Pb and prior to the addition of the
27 test organisms. This may have resulted in an increased bioavailability and overestimated Pb
28 toxicity relative to actual environmental conditions (Davies et al., 2003). See the discussion in
29 Section AX8.1.2 on the effects of aging on Pb sorption processes.
30 Lock and Janssen (2002) and Bongers et al. (2004) found that Pb-nitrate was more toxic
31 than Pb-chloride to survival and reproduction of the springtail Folsomia Candida. However,
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1 percolation (removal of the chloride or nitrate counted on) caused a significant decrease in Pb-
2 nitrate toxicity such that there was no difference in toxicity once the counterion was removed
3 (Bongers et al., 2004). No change in toxicity was observed for Pb-chloride once the chloride
4 was removed from the soil. Bongers et al. (2004) suggested that the nitrate ion was more toxic
5 than the chloride ion to springtails.
6 Uptake of Pb from soil into earthworms was also modeled as part of Eco-SSL
7 development (U.S. Environmental Protection Agency, 2005a). The relationship derived between
8 Pb in the soil and Pb in an earthworm was taken from Sample et al. (1999) and is as follows:
9
11 Ln(Cworm) = 0.807 * Ln(Csoil) - 0.218 (8-2)
13
14 where Cworm is the concentration of Pb in the earthworm (dry weight) and Csoil is the
15 concentration of Pb in the soil. This equation recognizes that the ratio of Pb concentration in
16 worm to Pb concentration in soil is not constant.
17
18 Wildlife
19 Research has been conducted to determine what Pb concentrations in various organs
20 would be indicative of various levels of effects. For example, Franson (1996) compiled data to
21 determine what residue levels were consistent with three levels of effects in Falconiformes (e.g.,
22 falcons, hawks, eagles, kestrels, ospreys), Columbiformes (e.g., doves, pigeons), and Galliformes
23 (e.g., turkey, pheasant, partridge, quail, chickens). The three levels of effect were (1) subclinical,
24 which are physiological effects only, such as the inhibition of 5-aminolevulinic acid dehydratase
25 (ALAD; see Section AX8.1.3.3); (2) toxic, a threshold level marking the initiation of clinical
26 signs, such as anemia, lesions in tissues, weight loss, muscular incoordination, green diarrhea,
27 and anorexia; and (3) compatible with death, an approximate threshold value associated with
28 death in field, captive, and/or experimental cases of Pb poisoning. The tissue Pb levels
29 associated with these levels of effects are presented in Table AX8-1.3.1.
30 Tissue residue levels below the subclinical levels in Table AX8-1.3.1 should be
31 considered "background" (Franson, 1996). Levels in the subclinical range are indicative of
32 potential injury from which the bird would probably recover if Pb exposure was terminated.
33 Toxic residues could lead to death. Residues above the compatible-with-death threshold are
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Table AX8-1.3.1. Tissue Lead Levels in Birds Causing Effects
(Taken from Franson, 1996)
Blood
Order (^g/dL)
Falconiformes
Subclinical 0.2-1.5
Toxic >1
Compatible with death >5
Columbiformes
Subclinical 0.2 - 2.5
Toxic >2
Compatible with death >10
Galliformes
Subclinical 0.2 - 3
Toxic >5
Compatible with death >10
Liver
(ppm wet wt.)
2-4
>3
>5
2-6
>6
>20
2-6
>6
>15
Kidney
(ppm wet wt.)
2-5
>3
>5
2-20
>15
>40
2-20
>15
>50
1 consistent with Pb-poisoning mortality (Franson, 1996). Additional information on residue
2 levels for Passeriformes (e.g., sparrows, starlings, robins, cowbirds), Charadriiformes (e.g., gulls,
3 terns), Gruiformes (e.g., cranes), Ciconiformes (e.g., egrets), Gaviformes (e.g., loons), and
4 Strigiformes (e.g., owls) is available (Franson, 1996). Scheuhammer (1989) found blood Pb
5 concentrations of between 0.18 and 0.65 |ig/mL in mallards corresponded to conditions
6 associated with greater than normal exposure to Pb but that should not be considered Pb
7 poisoning.
8 Lead concentrations in various tissues of mammals also have been correlated with toxicity
9 (Ma, 1996). The tissues commonly analysed for Pb are blood, liver, and kidney. Typical
10 baseline levels of blood Pb are approximately 4 to 8 |ig/dL for small mammals, and 2 to 6 |ig/dL
11 for mature cattle. Typical baseline levels of Pb in liver are 1 to 2 mg/kg dw for small mammals.
12 Typical baseline levels of Pb in kidney are 0.2 to 1.5 mg/kg dw for mice and voles, but shrews
13 typically have higher baseline levels of 3 to 19 mg/kg dw. Ma (1996) concluded that Pb levels
14 less than 5 mg/kg dw in liver and 10 mg/kg dw in kidney were not associated with toxicity, but
15 that levels greater than 5 mg/kg dw in liver and greater than 15 mg/kg dw in kidney could be
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1 taken as a chemical biomarker of toxic exposure to Pb in mammals. Humphreys (1991) noted
2 that the concentrations of Pb in liver and kidney can be elevated in animals with normal blood Pb
3 concentrations (and without exhibiting clinical signs of Pb toxicity), because Pb persists in these
4 organs longer than in blood.
5 Uptake of Pb from soil into small mammals was also modeled as part of Eco-SSL
6 development (U.S. Environmental Protection Agency, 2005a). The relationship derived between
7 Pb in the soil and Pb in the whole-body of a small mammal was taken from Sample et al. (1998)
8 and is as follows:
10
12 Ln(Cmammal) = 0.4422 * Ln(Csoil) + 0.0761 (8'3)
14
15 Where Cmammal is the concentration of Pb in small mammals (dry weight) and Csoil is the
16 concentration of Pb in the soil. Similar to the uptake equations for plants (Eq. 8-1) and
17 earthworms (Eq 8-2), the equation for mammalian uptake recognizes that the ratio of Pb
18 concentration in small mammals to Pb concentration in soil is not constant.
19
20 AX8.1.3.2 Resistance Mechanisms
21 Many mechanisms related to heavy metal tolerance in plants and invertebrates have been
22 described, including avoidance (i.e., root redistribution, food rejection), exclusion (i.e., selective
23 uptake and translocation), immobilization at the plant cell wall, and excretion (i.e., foliar
24 leakage, moulting) (Tyler et al., 1989; Patra et al., 2004). The following section reviews the
25 recent literature on the resistance mechanisms of plants and invertebrates through mitigation of
26 Pb (1) toxicity or (2) exposure.
27
28 Detoxification Mechanisms
29 Lead sequestration in cell walls may be the most important detoxification mechanism in
30 plants. Calcium may play a role in this detoxification by regulating internal Pb concentrations
31 through the formation of Pb-containing precipitates in the cell wall (Antosiewicz, 2005). Yang
32 et al. (2000) screened 229 varieties of rice (Oryza saliva) for tolerance or sensitivity to Pb and
33 found that the oxalate content in the root and root exudates was increased in Pb-tolerant varieties.
34 The authors suggested that the oxalate reduced Pb bioavailability, and that this was an important
35 tolerance mechanism (Yang et al., 2000). Sharma et al. (2004) found Pb-sulfur and Pb-sulfate in
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1 the leaves, and Pb-sulfur in the roots of Sesbania drummondii (Rattlebox Drummond), a Pb
2 hyperaccumulator plant grown in Pb-nitrate solution. They hypothesized that these sulfur
3 ligands were indicative of glutathione and phytochelatins, which play a role in heavy metal
4 homeostasis and detoxification (Sharma et al., 2004).
5 Sea pinks (Armeria maritimd) grown on a metal-contaminated site (calamine spoils more
6 than 100 years old) accumulated 6x the concentrations of Pb in brown (dead and withering)
7 leaves than green leaves (Szarek-Lukaszewska et al., 2004). The concentration of Pb in brown
8 leaves was similar to that in roots. This greater accumulation of Pb into older leaves was not
9 observed in plants grown hydroponically in the laboratory. The authors hypothesized that this
10 sequestering of Pb into the oldest leaves was a detoxification mechanism (Szarek-Lukaszewska
11 etal.,2004).
12 Terrestrial invertebrates also mitigate Pb toxicity. Wilczek et al. (2004) studied two
13 species of spider, the web-building Agelena labyrinthica and the active hunter wolf spider
14 Pardosa lugubris. The activity of metal detoxifying enzymes (via the glutathione metabolism
15 pathways) was greater in A. labyrinthica and in females of both species (Wilczek et al., 2004).
16 Marinussen et al. (1997) found that earthworms can excrete 60% of accumulated Pb very
17 quickly once exposure to Pb-contaminated soils has ended. However, the remainder of the body
18 burden is not excreted, possibly due to the storage of Pb in waste nodules that are too large to be
19 excreted (Hopkin, 1989). Gintenreiter et al. (1993) found that Lepidoptera larvae (in this case,
20 the gypsy moth Lymantria dispar) eliminated Pb, to some extent, in the meconium (the fluid
21 excreted shortly after emergence from the chrysalis).
22 Lead, in the form of pyromorphite (Pbs^O^Cl), was localized in the anterior pharynx
23 region of the nematode Ceanorhabditis elegans (Jackson et al., 2005). The authors hypothesized
24 that the nematode may detoxify Pb via its precipitation into pyromorphite, which is relatively
25 insoluble (Jackson et al., 2005).
26
27 Avoidance Response
28 Studies with soil invertebrates hypothesize that these organisms may avoid soil with high
29 Pb concentrations. For example, Bengtsson et al. (1986) suggested that the lower Pb
30 concentrations in earthworm tissues may be a result of lowered feeding activity of worms at
31 higher Pb concentrations in soil.
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1 AX8.1.3.3 Physiological Effects of Lead
2 Several studies have measured decreased blood ALAD activity in birds and mammals
3 exposed to Pb (U.S. Environmental Protection Agency, 1986a). Recent studies on the
4 physiological effects of Pb to consumers have focused on heme synthesis (as measured by
5 ALAD activity and protoporphyrin concentration), lipid peroxidation, and production of fatty
6 acids. Effects on growth are covered in Section AX8.1.4.
7 Biochemically, Pb adversely affects hemoglobin synthesis in birds and mammals. Early
8 indicators of Pb exposure in birds and mammals include decreased blood ALAD concentrations
9 and increased protoporphyrin IX activity. The effects of Pb on blood parameters and the use of
10 these parameters as sensitive biomarkers of exposure has been well documented (Eisler, 1988;
11 U.S. Environmental Protection Agency, 2005b). However, the linkage between these
12 biochemical indicators and ecologically relevant effects is less well understood. Low-level
13 inhibition of ALAD is not generally considered a toxic response, because this enzyme is thought
14 to be present in excess concentrations; rather, it may simply indicate that the organism has
15 recently been exposed to Pb (Henny et al., 1991).
16 Schlick et al. (1983) studied ALAD inhibition in mouse bone marrow and erythrocytes.
17 They estimated that an absorbed dose of between 50 and 100 jig Pb-acetate/kg body weight per
18 day would result in long-term inhibition of ALAD.
19 Beyer et al. (2000) related blood Pb to sublethal effects in waterfowl along the Coeur
20 d'Alene River near a mining site in Idaho. The sublethal effects measured included, among
21 others, red blood cell ALAD activity and protoporphyrin levels in the blood. As found in other
22 studies, ALAD activity was the most sensitive indicator of Pb exposure, decreasing to 3% of the
23 reference value at a blood Pb concentration of 0.68 mg/kg ww (wet weight). Protoporphyrin
24 concentrations showed a 4.2-fold increase at this same concentration.
25 Henny et al. (1991) studied osprey along the Coeur d'Alene River. There were no
26 observations of death, behavioral abnormalities, or reduced productivity related to Pb exposure,
27 although inhibition of blood ALAD and increased protoporphyrin concentrations were measured
28 in ospreys. Henny et al. (1991) hypothesized that no impacts to osprey were observed, even
29 though swan mortality was documented in the area because swans feed at a lower trophic level
30 (i.e., Pb does not biomagnify, and thus is found at higher concentrations in lower trophic level
31 organisms).
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1 Hoffman et al. (2000a) also studied the effects of Coeur d'Alene River sediment on
2 waterfowl, focusing on mallard ducklings for 6 weeks after hatching. The study revealed that a
3 90% reduction in ALAD activity and a greater than 3-fold increase in protoporphyrin
4 concentration occurred when blood Pb reached a concentration of 1.41 mg/kg ww as a result of
5 the ducklings being fed a diet composed of 12% sediment (3449 mg/kg Pb). Those ducklings
6 fed a diet composed of 24% sediment were found to have a mean blood Pb concentration of 2.56
7 mg/kg ww and a greater than 6-fold increase in protoporphyrin concentration. Hoffman et al.
8 (2000b) also studied Canada Geese (Branta canadensis) goslings in a similar fashion. The
9 results revealed that, while blood Pb concentrations in goslings were approximately half (0.68
10 mg/kg ww) of those found in ducklings under the same conditions (12% diet of 3449 mg/kg
11 sediment Pb), goslings showed an increased sensitivity to Pb exposure. Goslings experienced a
12 90% reduction in ALAD activity and a 4-fold increase in protoporphyrin concentration, similar
13 to conditions found in the ducklings, although blood Pb concentrations were half those found in
14 the ducklings. More serious effects were seen in the goslings when blood Pb reached 2.52
15 mg/kg, including decreased growth and mortality.
16 Redig et al. (1991) reported a hawk LOAEL (lowest observed adverse effect level) of 0.82
17 mg/kg-day for effects on heme biosynthetic pathways. Lead dosages as high as 1.64 to 6.55
18 mg/kg-day caused neither mortality nor clinical signs of toxicity. A dose of 6.55 mg/kg-day
19 resulted in blood Pb levels of 1.58 |ig/mL. There were minimal changes in immune function
20 (Redig etal., 1991).
21 Repeated oral administration of Pb resulted in biochemical alterations in broiler chickens
22 (Brar et al., 1997a,b). At a dose of 200 mg/kg-day Pb-acetate, there were significant increases in
23 plasma levels of uric acid and creatinine and significant declines in the levels of total proteins,
24 albumin, glucose, and cholesterol. Brar et al. (1997a) suggested that increased uric acid and
25 creatinine levels could be due to an accelerated rate of protein catabolism and/or kidney damage.
26 They also suggested that the decline in plasma proteins and albumin levels may be caused by
27 diarrhea and liver dysfunction due to the Pb exposure. Brar et al. (1997b) also found that
28 significant changes in plasma enzymes may be causing damage to other organs.
29 Lead can cause an increase in tissue lipid peroxides and changes in glutathione
30 concentrations, which may be related to peroxidative damage of cell membranes (Mateo and
31 Hoffman, 2001). There are species-specific differences in resistance to oxidative stress (lipid
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1 peroxidation), which may explain why Canada geese are more sensitive to Pb poisoning than
2 mallards (Mateo and Hoffman, 2001). Lead also caused an increase in the production of the fatty
3 acid arachidonic acid, which has been associated with changes in bone formation and immune
4 response (Mateo et al., 2003a). The effects observed by Mateo et al. (2003a,b) were associated
5 with very high concentrations of Pb in the diet (1840 mg Pb/kg diet), much higher than would be
6 found generally in the environment, and high enough that birds decreased their food intake.
7 Lead also induces lipid peroxidation in plants. Rice plants exposed to a highly toxic level
8 of Pb (1000 jiM in nutrient solution) showed elevated levels of lipid peroxides, increased activity
9 of superoxide dismutase, guaiacol peroxidase, ascorbate peroxidase, and glutatione reductase
10 (Verma and Dubey, 2003). The elevated levels of these enzymes suggest the plants may have an
11 antioxidative defense mechanism against oxidative injury caused by Pb (Verma and Dubey,
12 2003).
13
14 AX8.1.3.4 Factors that Modify Organism Response
15 Research has demonstrated that Pb may affect survival, reproduction, growth,
16 metabolism, and development in a wide range of species. These effects may be modified by
17 chemical, biological, and physical factors. The factors that modify responses of organisms to Pb
18 are described in the following sections.
19
20 Genetics
21 Uptake and toxicity of Pb to plants are influenced strongly by the type of plant. Liu et al.
22 (2003) found that Pb uptake and translocation by rice plants differed by cultivar (a cultivated
23 variety of plant produced by selective breeding) but was not related to genotype. Twenty
24 cultivars were tested from three genotypes. The differences in Pb concentrations among
25 cultivars were smallest when comparing concentrations in the grains at the ripening stage. This
26 study also found that toxicity varied by cultivar; at 800 mg Pb/kg soil, some cultivars were
27 greatly inhibited, some were significantly improved, and others showed no change.
28 Dearth et al. (2004) compared the response of Fisher 344 (F344) rats and Sprague-Dawley
29 (SD) rats to exposure via gavage to 12 mg Pb/mL as Pb-acetate. Blood Pb levels in the F344
30 dams were higher than those of the SD dams. Lead delayed the timing of puberty and
31 suppressed hormone levels in F344 offspring. These effects were not observed in the offspring
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1 of SD rats, even when the dose was doubled. The authors conclude that F344 rats are more
2 sensitive to Pb (Dearth et al., 2004).
3
4 Biological Factors
5 Several biological factors may influence Pb uptake and organism response, including
6 organism age, sex, species, feeding guild, and, for plants, the presence of mycorrhizal fungi.
7 Monogastric animals are more sensitive to Pb than ruminants (Humphreys, 1991).
8 Younger organisms may be more susceptible to Pb toxicity (Eisler, 1988; Humphreys,
9 1991). Nestlings are more sensitive to the effects of Pb than older birds, and young altricial birds
10 (species unable to self-regulate body heat at birth, such as songbirds), are considered more
11 sensitive than precocial birds (species that have a high degree of independence at birth, such as
12 quail, ducks, and poultry) (Scheuhammer, 1991).
13 Sex can also have an effect on the accumulation of Pb by wildlife (Eisler, 1988). Female
14 birds accumulate more Pb than males (Scheuhammer, 1987; Tejedor and Gonzalez, 1992).
15 These and other authors have related this to the increased requirement for calcium in laying
16 females.
17 Different types of invertebrates accumulate different amounts of Pb from the environment
18 (U.S. Environmental Protection Agency, 1986a). There may be species-and sex-specific
19 differences in accumulation of Pb into invertebrates, specifically arthropods. This has been
20 shown by Wilczek et al. (2004) who studied two species of spider, the web-building
21 A. labyrinthica and the active hunter wolf spider P. lugubris. The body burdens of Pb in the
22 wolf spider were higher than in the web-building spider, and this may be due to the more
23 effective use of glutathione metabolism pathways in A. labyrinthica. Body burdens of females
24 were lower than those of males in both species. This was also observed in spiders by Rabitsch
25 (1995a). Females are thought to be able to detoxify and excrete excess metals more effectively
26 than males (Wilczek et al., 2004). Lead accumulation has been measured in numerous species of
27 arthropods with different feeding strategies. Differences were observed between species
28 (Janssen and Hogervorst, 1993; Rabitsch, 1995a) and depending upon sex (Rabitsch, 1995a),
29 developmental stage (Gintenreiter et al., 1993; Rabitsch, 1995a), and season (Rabitsch, 1995a).
30 Uptake of Pb may be enhanced by symbiotic associations between plant roots and
31 mycorrhizal fungi. Similar to the mechanism associated with increased uptake of nutrients,
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1 mycorrhizal fungi also may cause an increase in the uptake of Pb by increasing the surface area
2 of the roots, the ability of the root to absorb particular ions, and the transfer of ions through the
3 soil (U.S. Environmental Protection Agency, 1986a). There have been contradictory results
4 published in the literature regarding the influence of mycorrhizal organisms on the uptake and
5 toxicity of Pb to plants (see review in Pahlsson, 1989). Lin et al. (2004) found that the
6 bioavailability of Pb increased in the rhizosphere of rice plants, although the availability varied
7 with Pb concentration in soil. Bioavailability was measured as the soluble plus exchangeable Pb
8 fraction from sequential extraction analysis. The authors hypothesized that the enhanced
9 solubility of Pb may be due to a reduced pH in the rhizosphere or, more likely, the greater
10 availability of organic ligands, which further stimulates microbial growth (Lin et al., 2004).
11 Increased bioavailability of Pb in soil may increase the uptake of Pb into plants, although
12 this was not assessed by Lin et al. (2004). However, Dixon (1988) found that red oak
13 (Quercus rubrd) seedlings with abundant ectomycorrhizae had lower Pb concentrations in their
14 roots than those seedlings without this fungus, although only at the 100 mg Pb/kg sandy loam
15 soil concentration (no differences were found at lower Pb concentrations). Lead in soil also was
16 found to be toxic to the ectomycorrhizal fungi after 16 weeks of exposure to 50 mg Pb/kg or
17 more (Dixon, 1988). Malcova and Gryndler (2003) showed that maize root exudates from
18 mycorrhizal fungi can ameliorate heavy metal toxicity until a threshold metal concentration was
19 surpassed. This may explain the conflicting results in the past regarding the uptake and toxicity
20 of Pb to plants with mycorrhizal fungi.
21 The type of food eaten is a major determinant of Pb body burdens in small mammals, with
22 insectivorous animals accumulating more Pb than herbivores or granivores (U.S. Environmental
23 Protection Agency, 1986a). In fact, the main issue identified by the EPA (U.S. Environmental
24 Protection Agency, 1986a) related to invertebrate uptake of Pb was not toxicity to the
25 invertebrates, but accumulation of Pb to levels that may be toxic to their consumers. Several
26 authors suggest that shrews are a good indicator of metal contamination, because they tend to
27 accumulate higher levels of metals than herbivorous small mammals (see data summary in
28 Sample et al. (1998)). Shrews accumulate higher levels of metals in contaminated habitats,
29 because their diet mainly consists of detritivores (i.e., earthworms) and other soil invertebrates in
30 direct contact with the soil (Beyer et al., 1985).
31
May 2006 AX8-51 DRAFT-DO NOT QUOTE OR CITE
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1 Physical/Environmental Factors
2 Plants
3 The uptake and distribution of Pb into higher plants from the soil is affected by various
4 chemical and physical factors including the chemical form of Pb, the presence of other metal
5 ions, soil type, soil pH, cation exchange capacity (CEC), the amount of Fe/Mn-oxide films
6 present, organic matter content, temperature, light, and nutrient availability. A small fraction of
7 Pb in soil may be released into the soil water, which is then available to be taken up by plants
8 (U.S. Environmental Protection Agency, 1986a).
9 The form of Pb has an influence on its toxicity to plants. For example, Pb-oxide is less
10 toxic than more bioavailable forms such as Pb-chloride or Pb-acetate. In a study by Khan and
11 Frankland (1983), radish plants were exposed to Pb-oxide and Pb-chloride in a loamy sand at pH
12 5.4, in a 42-day study. In a tested concentration range of 0 to 5000 mg/kg, root growth was
13 inhibited by 24% at 500 mg/kg for Pb-chloride and an ECso of 2400 mg/kg was calculated from a
14 dose-response curve. Plant growth ceased at 5000 mg/kg and shoots exhibited an ECso of
15 2800 mg/kg. For Pb-oxide exposure (concentration range of 0 to 10,000 mg/kg), reported results
16 indicate an ECso of 12,000 mg/kg for shoot growth and an ECso of 10,000 mg/kg for root growth.
17 There was no effect on root growth at 500 mg/kg and a 26% reduction at 1000 mg/kg Pb oxide.
18 Soil pH is the most influential soil property with respect to uptake and accumulation of Pb
19 into plant species. This is most likely due to increased bioavailability of Pb created by low soil
20 pH. At low soil pH conditions, markedly elevated Pb toxicity was reported for red spruce
21 (P. rubens) (Seller and Paganelli, 1987). At a soil pH of 4.5, ryegrass (Lolium hybridum)
22 and oats (Avena sativd) had significantly higher Pb concentrations after 3 months of growth
23 compared to plants grown at pH 6.4 (Allinson and Dzialo, 1981).
24
25 Invertebrates
26 The uptake of Pb into invertebrates depends on the physical environment and parameters
27 such as pH, calcium concentration, organic matter content, and CEC. Greater accumulation is
28 found generally when the soil pH or organic content is lower (U.S. Environmental Protection
29 Agency, 1986a).
30 Soil pH has a significant influence on uptake of Pb into invertebrates. Peramaki et al.
31 (1992) studied the influence of soil pH on uptake into the earthworm Aporrectodea caliginosa.
May 2006 AX8-52 DRAFT-DO NOT QUOTE OR CITE
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1 Lead accumulation was lowest at the highest pH values, but there was no statistical difference
2 due to variability in the data. Variability in the response also was found by Bengtsson et al.
3 (1986), who reared earthworms (Dendrobaena rubida) in acidified soils at pH 4.5, 5.5, or 6.5.
4 Lead uptake into worms was pH-dependent, although the highest concentrations were not always
5 found at the lowest pH. There was no clear relationship between Pb concentration in cocoons
6 and soil pH, and Pb concentrations were higher in the hatchlings than in the cocoons. As has
7 been reported in many other studies (Neuhauser et al., 1995), concentration factors (ratio of Pb in
8 worm to Pb in soil) were lower at higher Pb concentrations in soil. The authors attribute some of
9 this to a lowered feeding activity in worms at higher Pb concentrations (Bengtsson et al., 1986).
10 Beyer et al. (1987) and Morgan and Morgan (1988) recognized that other factors beyond
11 soil pH could influence the uptake of Pb into earthworms, which may be the cause of the
12 inconsistencies reported by several authors. Both studies evaluated worm uptake of Pb relative
13 to pH, soil calcium concentration, and organic matter content. Morgan and Morgan (1988) also
14 considered CEC, and Beyer et al. (1987) considered concentrations of phosphorus, potassium, or
15 magnesium in soil. Both studies found that calcium concentrations in soil were correlated with
16 soil pH. Morgan and Morgan (1988) also found that CEC was correlated with percentage
17 organic matter. Soil pH (coupled with CEC) and soil calcium were found to play significant
18 roles in the uptake of Pb into worms (Beyer et al., 1987; Morgan and Morgan, 1988). Beyer
19 et al. (1987) noted that concentrations of phosphorus in soil had no effect.
20
21 Nutritional Factors
22 Diet is a significant modifier of Pb absorption and of toxic effects in many species of
23 birds and mammals (Eisler, 1988). Dietary deficiencies in calcium, zinc, iron, vitamin E, copper,
24 thiamin, phosphorus, magnesium, fat, protein, minerals, and ascorbic acid increased Pb
25 absorption and its toxic effects (Eisler, 1988).
26 Mateo et al. (2003b) studied intraspecies sensitivity to Pb-induced oxidative stress, by
27 varying the vitamin E content of mallard diets. Vitamin E can protect against peroxidative
28 damage and was found to decrease the lipid peroxidation in nerves of birds; however, it did not
29 alleviate any sign of the Pb poisoning. The authors hypothesize that inhibition of antioxidant
30 enzymes and interaction with sulfhydryl groups of proteins may have a greater influence on Pb
31 toxicity than lipid peroxidation (Mateo et al., 2003b). The effects observed by Mateo et al.
May 2006 AX8-53 DRAFT-DO NOT QUOTE OR CITE
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1 (2003b) were associated with very high concentrations of Pb in diet (1840 mg Pb/kg diet), much
2 higher than would be found generally in the environment, and high enough that the birds
3 decreased their food intake.
4 Mallard ducklings were exposed to Pb-contaminated sediment and either a low nutrition
5 or optimal nutrition diet (Douglas-Stroebel et al., 2005). Lead exposure combined with a
6 nutritionally inferior diet caused more changes in behavior (as measured by time bathing, resting,
7 and feeding) than Pb exposure or low-nutrition diet alone. These effects may be due to the low-
8 nutrition diet being deficient in levels of protein, amino acids, calcium, zinc, and other nutrients.
9 Zebra finches (Taeniopygia guttatd) were exposed to Pb-acetate via drinking water at
10 20 mg/L for 38 days, along with either a low- or high-calcium diet (Snoeijs et al., 2005). Lead
11 uptake into tissues was enhanced by a low-calcium diet. Lead did not affect body weight,
12 hematocrit, or adrenal stress response. Lead suppressed the humoral immune response only in
13 females on a low-calcium diet, suggesting increased susceptibility of females to Pb (Snoeijs
14 et al., 2005).
15
16 Interactions with Other Pollutants
17 Lead can interact with other pollutants to exert toxicity in an antagonistic (less than
18 additive), independent, additive, or synergistic (more than additive) manner. Concurrent
19 exposure to Pb and additional pollutant(s) can affect the ability of plants to uptake Pb or the
20 other pollutant. However, the uptake and toxic response of plants exposed to Pb combined with
21 other metals is inconsistent (Pahlsson, 1989). Therefore, no generalizations can be made about
22 the relative toxicity of metal mixtures. For example, An et al. (2004) conducted acute, 5-day
23 bioassays on cucumber exposed to Pb, Pb + copper, Pb + cadmium, or Pb + copper + cadmium
24 in a sandy loam soil of pH 4.3. Shoot and root growth were measured. Depending on the tissue
25 and metal combination, additivity, synergism, or antagonism was observed in the responses to
26 these metals. In fact, the response in roots was not consistent with the response in shoots for the
27 binary mixtures. However, the combined effects were greater in the roots than the shoots, which
28 may be explained by the tendency for Pb and other heavy metals to be retained in the roots of
29 plants. In addition, the pattern of metal bioaccumulation into plant tissue did not always
30 correlate with the toxic response. However, antagonism was observed in the response of roots
31 and shoots exposed to all three metals, and this was reflected in the decreased accumulation of
May 2006 AX8-54 DRAFT-DO NOT QUOTE OR CITE
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1 metals into plant tissues. The authors hypothesized that this may be due to the formation of less
2 bioavailable metal complexes (An et al., 2004).
3 He et al. (2004) found that selenium and zinc both inhibited the uptake of Pb into Chinese
4 cabbage (Brassica rapct) and lettuce (Lactuca saliva). Zinc applied at 100 mg/kg or selenium
5 applied at 1 mg/kg decreased the uptake of Pb (present in soil at 10 mg/kg as Pb-nitrate) into
6 lettuce by 15% and 20%, respectively, and into Chinese cabbage by 23 and 20%, respectively.
7 Selenium compounds were evaluated to determine whether they could change the
8 inhibition of ALAD in liver, kidney, or brain of mice exposed to Pb-acetate (Perottoni et al.,
9 2005). Selenium did not affect the inhibition of ALAD in the kidney or liver, but it did reverse
10 the ALAD inhibition in mouse brain.
11 Co-occurrence of cadmium with Pb resulted in reduced blood Pb concentrations in rats
12 (Garcia and Corredor, 2004). The authors hypothesized that cadmium may block or antagonize
13 the intestinal absorption of Pb, or the metallothionein induced by cadmium may sequester Pb.
14 However, this was not observed in pigs, where blood Pb concentrations were greater when
15 cadmium was also administered (Phillips et al., 2003). The effect on growth rate also was
16 additive when both metals were given to young pigs (Phillips et al., 2003).
17
18 AX8.1.3.5 Summary
19 The current document expands upon and updates knowledge related to the uptake,
20 detoxification, physiological effects, and modifying factors of Pb toxicity to terrestrial
21 organisms.
22
23 Surface Deposition onto Plants
24 Recent work (Dalenberg and Van Driel, 1990; Jones and Johnston, 1991; Angelova et al.,
25 2004) has supported previous results and conclusions that surface deposition of Pb onto above-
26 ground vegetation from airborne sources may be significant (U.S. Environmental Protection
27 Agency, 1986a). Similarly, it has been well documented previously that Pb in soil also is taken
28 up by plants, although most remains in the roots, there is little translocation to shoots, leaves, or
29 other plant parts (U.S. Environmental Protection Agency, 1986a). More recent work continues
30 to support this finding (Sieghardt, 1990), and one study found increased tolerance in species with
31 bulbs, possibly due to the storage of Pb in the bulb (Wierzbicka, 1999).
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1 Uptake Mechanism into Plants
2 Lead was thought previously to be taken up by plants via the symplastic route (through
3 cell membranes), although it was unknown whether some Pb also may be taken up via the
4 apoplastic route (between cells) (U.S. Environmental Protection Agency, 1986a). Recent work
5 has shown that the apoplastic route of transport is stopped in the primary roots by the endodermis
6 (Sieghardt, 1990), supporting the previous conclusion that the symplastic route is the most
7 significant route of tranport into plant cells.
8
9 Species Differences in Uptake into Earthworms
10 Different species of earthworm accumulated different amounts of Pb, and this was not
11 related to feeding strategy (U.S. Environmental Protection Agency, 1986a). This is supported by
12 recent work, which has shown Aporrectodea accumulated more than Lumbricus (Terhivuo et al.,
13 1994; Pizl and Josens, 1995), although this is not consistently observed (Spurgeon and Hopkin,
14 1996a).
15
16 Speciation and Form of Lead
17 Recent work supports previous conclusions that the form of metal tested, and its
18 speciation in soil, influence uptake and toxicity to plants and invertebrates (U.S. Environmental
19 Protection Agency, 1986a). The oxide form is less toxic that the chloride or acetate forms,
20 which are less toxic than the nitrate form of Pb (Khan and Frankland, 1983; Lock and Janssen,
21 2002; Bongers et al., 2004). However, these results must be interpreted with caution, as the
22 counterion (e.g., the nitrate ion) may be contributing to the observed toxicity (Bongers et al.,
23 2004).
24
25 Detoxification in Plants
26 Lead may be deposited in root cell walls as a detoxification mechanism (U.S.
27 Environmental Protection Agency, 1986a), and this may be influenced by calcium concentrations
28 (Antosiewicz, 2005). Yang et al. (2000) suggested that the oxalate content in root and root
29 exudates reduced the bioavailability of Pb in soil, and that this was an important tolerance
30 mechanism. Other hypotheses put forward recently include the presence of sulfur ligands
May 2006 AX8-56 DRAFT-DO NOT QUOTE OR CITE
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1 (Sharma et al., 2004) and the sequestration of Pb in old leaves (Szarek-Lukaszewska et al., 2004)
2 as detoxification mechanisms.
3
4 Detoxification in Invertebrates
5 Lead detoxification has not been studied extensively in invertebrates. Glutathione
6 detoxification enzymes were measured in two species of spider (Wilczek et al., 2004). Lead may
7 be stored in waste nodules in earthworms (Hopkin, 1989) or as pyromorphite in the nematode
8 (Jackson et al., 2005).
9
10 Physiological Effects
11 The effects on heme synthesis (as measured by ALAD activity and protoporphyrin
12 concentration, primarily) have been well-documented (U.S. Environmental Protection Agency,
13 1986a) and continue to be studied (Schlick et al., 1983; Scheuhammer, 1989; Henny et al., 1991;
14 Redig et al., 1991; Beyer et al., 2000; Hoffman et al., 2000a,b). However, Henny et al. (1991)
15 caution that changes in ALAD and other enzyme parameters are not always related to adverse
16 effects, but simply indicate exposure. Other effects on plasma enzymes, which may damage
17 other organs, have been reported (Brar et al., 1997a,b). Lead also may cause lipid peroxidation
18 (Mateo and Hoffman, 2001), which may be alleviated by vitamin E, although Pb poisoning may
19 still result (Mateo et al., 2003b). Changes in fatty acid production have been reported, which
20 may influence immune response and bone formation (Mateo et al., 2003a).
21
22 Response Modification
23 Genetics, biological factors, physical/environmental factors, nutritional factors, and other
24 pollutants can modify terrestrial organism response to Pb. Fisher 344 rats were found to be more
25 sensitive to Pb than Sprague-Dawley rats (Dearth et al., 2004). Younger animals are more
26 sensitive than older animals (Eisler, 1988; Scheuhammer, 1991), and females generally are more
27 sensitive than males (Scheuhammer, 1987; Tejedor and Gonzalez, 1992; Snoeijs et al., 2005).
28 Monogastric animals are more sensitive than ruminants (Humphreys, 1991). Insectivorous
29 mammals may be more exposed to Pb than herbivores (Beyer et al., 1985; Sample et al., 1998),
30 and higher tropic-level consumers may be less exposed than lower trophic-level organisms
May 2006 AX8-57 DRAFT-DO NOT QUOTE OR CITE
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1 (Hennyetal., 1991). Nutritionally-deficient diets (including low calcium) cause increased
2 uptake of Pb (Snoeijs et al., 2005) and greater toxicity (Douglas-Stroebel et al., 2005) in birds.
3 Mycorrhizal fungi may ameliorate Pb toxicity until a threshold is surpassed (Malcova and
4 Gryndler, 2003), which may explain why some studies show increased uptake into plants (Lin
5 et al., 2004) while others show no difference or less uptake (Dixon, 1988). Lower soil pH
6 generally increases uptake of Pb into plants and soil invertebrates. However, calcium content,
7 organic matter content, and cation exchange capacity of soils also have had a significant
8 influence on uptake of Pb into plants and invertebrates (Beyer et al., 1987; Morgan and Morgan,
9 1988).
10 Interactions of Pb with other metals are inconsistent, depending on the endpoint
11 measured, the tissue analyzed, the animal species, and the metal combination (Phillips et al.,
12 2003; An et al., 2004; He et al., 2004; Garcia and Corredor, 2004; Perottoni et al., 2005).
13
14 AX8.1.4 Exposure-Response of Terrestrial Species
15 Section AX8.1.3 summarized the most important factors related to uptake of Pb by
16 terrestrial organisms, the physiological effects of Pb, and the factors that modify terrestrial
17 organism responses to Pb. Section AX8.1.4 outlines and highlights the critical recent
18 advancements in the understanding of the toxicity of Pb to terrestrial organisms. This section
19 begins with a summary of the conclusions from the 1986 Pb AQCD (U.S. Environmental
20 Protection Agency, 1986a) and then summarizes the more recent critical research conducted on
21 effects of Pb on primary producers, consumers, and decomposers. All concentrations are
22 expressed as mg Pb/kg soil dw, unless otherwise indicated.
23 The summary of recent critical advancements in understanding toxicity relies heavily on
24 the work completed by a multi-stakeholder group, consisting of federal, state, consulting,
25 industry, and academic participants, led by the EPA to develop Ecological Soil Screening Levels
26 (Eco-SSLs). Eco-SSLs describe the concentrations of contaminants in soils that are protective of
27 ecological receptors (U.S. Environmental Protection Agency, 2005a). They were developed to
28 identify contaminants requiring further evaluation in an ecological risk assessment and were not
29 designed to be used as cleanup target levels. Eco-SSLs were derived for terrestrial plants, soil
30 invertebrates, birds, and mammals. Detailed procedures using an extensive list of acceptability
31 and exclusion criteria (U.S. Environmental Protection Agency, 2005a) were used in screening
May 2006 AX8-58 DRAFT-DO NOT QUOTE OR CITE
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1 the toxicity studies to ensure that only those that met minimum quality standards were used to
2 develop the Eco-SSLs. In addition, two peer reviews were completed during the Eco-SSL
3 development process. The first was a consultation with the EPA Science Advisory Board (SAB)
4 in April 1999, and the second was a peer review workshop in July 2000, which was open to
5 the public.
6 Areas of research that were not addressed are effects from irrelevant exposure conditions
7 relative to airborne emissions of Pb (e.g., Pb shot, Pb paint, injection studies, studies conducted
8 on mine tailings, and studies conducted with hydroponic solutions); mixture toxicity (addressed
9 in Section AX8.1.3); issues related to indirect effects (e.g., effects on predator/prey interactions,
10 habitat alteration, etc.); and human health-related research (e.g., hypertension), which is
11 addressed in other sections of this document.
12 The toxicity data presented herein should be reviewed with a note of caution regarding
13 their relevance to field conditions. Laboratory studies, particularly those using Pb-spiked soil,
14 generally do not allow the soil to equilibrate following the addition of Pb and prior to the
15 addition of test organisms. This may result in increased bioavailability and overestimation of Pb
16 toxicity relative to actual environmental conditions (Davies et al., 2003).
17
18 AX8.1.4.1 Summary of Conclusions from the 1986 Lead Criteria Document
19 The previous Pb AQCD, Volume II (U.S. Environmental Protection Agency, 1986a)
20 reviewed the literature on the toxicity of Pb to plants, soil organisms, birds, and mammals. The
21 main conclusions from this document are provided below.
22
23 Primary Producers
24 Commonly reported effects of Pb on vascular plants include the inhibition of
25 photosynthesis, respiration, and/or cell elongation, all of which reduce plant growth. However, it
26 was noted that studies of other effects on plant processes such as maintenance, flowering, and
27 hormone development had not been conducted; therefore, no conclusion could be reached
28 concerning effects of Pb on these processes.
29 The EPA (U.S. Environmental Protection Agency, 1986a) concluded that most plants
30 experience reduced growth when Pb concentrations in soil moisture (the film of moisture
31 surrounding soil particles in the root zone of soil) exceed 2 to 10 mg/kg. It also was concluded
May 2006 AX8-59 DRAFT-DO NOT QUOTE OR CITE
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1 that most plants would experience reduced growth (inhibition of photosynthesis, respiration, or
2 cell elongation) in soils of > 10,000 mg/kg when soil composition and pH are such that
3 bioavailability of Pb in the soil is low (see Section AX8.1.3 for details on factors affecting
4 bioavailability of Pb in soil). Acid soils or soils with low organic matter tend to increase Pb
5 bioavailability and would inhibit plants at much lower Pb concentrations (e.g., as low as
6 <100 mg/kg).
7 Many effect levels have been reported at Pb concentrations much lower than
8 10,000 mg/kg soil. For example, effects on rye grass (Lolium rigidum) exposed to Pb in soil
9 included inhibition of germinating root elongation (at <2.5 mg/kg), absence of root growth
10 (at 5 mg/kg), or 55% inhibition of seed germination (at 20 to 40 mg/kg). Stunted growth in
11 radish (Raphanus sativus) was observed at 1000 mg/kg soil, with complete growth inhibition at
12 5000 mg/kg, when Pb was added as Pb-chloride; effects were less severe when the Pb was added
13 as Pb-oxide.
14
15 Consumers
16 The EPA (U.S. Environmental Protection Agency, 1986a) concluded that food is the
17 largest contributor of Pb to animals, with inhalation rarely accounting for more than 10 to 15%
18 of daily intake of Pb and drinking water exposures being quite low. It also was concluded that a
19 regular dose of 2 to 8 mg/kg-day causes death in most animals. Grazing animals may consume
20 more than 1 mg/kg-day in habitats near smelters and roadsides, but no toxic effects were
21 documented in these animals.
22
23 Decomposers
24 Lack of decomposition has been observed as a particular problem around smelter sites.
25 Lead concentrations between 10,000 and 40,000 mg/kg soil can eliminate populations of
26 decomposer bacteria and fungi (U.S. Environmental Protection Agency, 1986a). Lead may
27 affect decomposition processes by direct toxicity to specific groups of decomposers, by
28 deactivating enzymes excreted by decomposers to break down organic matter, or by binding with
29 the organic matter and rendering it resistant to the action of decomposers.
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1 Microorganisms are more sensitive than plants to Pb in soil. Delayed decomposition may
2 occur at between 750 and 7500 mg/kg soil (depending on soil type and other conditions).
3 Nitrification is inhibited by 14% at 1000 mg/kg soil.
4
5 U.S. Environmental Protection Agency Staff Review of 1986 Criteria Document
6 The EPA reviewed the 1986 Pb AQCD and presented an overall summary of conclusions
7 and recommendations (U.S. Environmental Protection Agency, 1990). The major conclusion
8 was that available laboratory and field data indicated that high concentrations of Pb can affect
9 certain plants and alter the composition of soil microbial communities. It was noted that few
10 field studies were available in which Pb exposures and associated effects in wildlife were
11 reported.
12
13 AX8.1.4.2 Recent Studies on the Effects of Lead on Primary Producers
14 Several studies published since 1986 have reported terrestrial plant exposure to Pb in soil,
15 many of which were reviewed during the development of the Eco-SSLs (U.S. Environmental
16 Protection Agency, 2005b). The relevant information from the Eco-SSL document (U.S.
17 Environmental Protection Agency, 2005b) is summarized below. A literature search and review
18 also was conducted to identify critical papers published since 2002, which is when the literature
19 search was completed for Eco-SSL development, and no new papers were identified as critical to
20 the understanding of Pb toxicity to terrestrial primary producers.
21 Effects observed in studies conducted since the 1986 Pb AQCD are similar to those
22 reported previously and include decreased photosynthetic and transpiration rates and decreased
23 growth and yield (U.S. Environmental Protection Agency, 2005b). The phytotoxicity of Pb is
24 considered relatively low, due to the limited availability and uptake of Pb from soil and soil
25 solution and minimal translocation of Pb from roots to shoots (Pahlsson, 1989). Although many
26 laboratory toxicity studies have reported effects on plants, there are few reports of phytotoxicity
27 from Pb exposure under field conditions. For example, Leita et al. (1989) and Sieghardt (1990)
28 reported high concentrations of Pb and other metals in soil and vegetation collected around
29 mining areas in Europe, with no toxicity symptoms observed in plants or fruit.
30 The literature search completed for the terrestrial plant Eco-SSL development identified
31 439 papers for detailed review, of which 28 met the minimum criteria (U.S. Environmental
May 2006 AX8-61 DRAFT-DO NOT QUOTE OR CITE
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1
2
3
4
5
6
7
Protection Agency, 2005a). Thirty ecotoxicological endpoints were gleaned from these 28
papers and were further evaluated; most of those evaluated growth (biomass), which was
considered the most sensitive and ecologically relevant endpoint (U.S. Environmental Protection
Agency, 2005b). Five of the endpoints, representing four species tested under three different
combinations of pH and organic matter content, were used to develop the Eco-SSL of 120 mg/kg
(115 mg/kg rounded to two significant digits) (Table AX8-1.4.1).
Table AX8-1.4.1. Plant Toxicity Data Used to Develop the Eco-SSL
Plant Species
Loblolly pine (Pinus taeda)
Red maple (Acer rubrum)
Berseem clover
(Trifolium alexandrium)
Berseem clover
Rye grass (Lolium rigidum)
Soil pH
4
4
6.3
6.7
5.6
% Organic
Matter
2.5
2.5
0.94
3.11
0.1
Toxicity
Parameter
MATC* (growth)
MATC (growth)
MATC (growth)
MATC (growth)
MATC (growth)
Geometric Mean
Pb in Soil
(mg/kg dw)
144
144
316
141
22
115
*MATC = Maximum Acceptable Threshold Concentration, or the geometric mean of the NOAEC
(no-observed-adverse-effect concentration) and LOAEC (lowest-observed-adverse-effect concentration).
Source: U.S. Environmental Protection Agency (2005b).
9 The 25 ecotoxicological endpoints that were not used to develop the Eco-SSL for plants
10 are presented in Table AX8-1.4.2. The first six endpoints were considered eligible for Eco-SSL
11 derivation but were not used; the remainder did not meet all of the requirements to be considered
12 for inclusion in the Eco-SSL derivation process.
13
14
15
AX8.1.4.3 Recent Studies on the Effects of Lead on Consumers
Since the 1986 Pb AQCD (U.S. Environmental Protection Agency, 1986a), there have
16 been several studies in which birds and mammals were exposed to Pb via ingestion (primarily
17 through dietary Pb). Many of these were reviewed during development of the Eco-SSLs (U.S.
May 2006
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Table AX8-1.4.2. Plant Toxicity Data Not Used to Develop the Eco-SSL
Plant Species Soil pH
Studies eligible for Eco-SSL derivation,
Berseem clover
(Trifolium alexandrium)
Tomato
(Lycopericon esculentum)
Tomato
Fenugreek
(Trigonella foenum-graecum)
Spinach (Spinacea oleracea)
Com (Zea mays)
Sow thistle (Sonchus oleraceus)
but not used
6.7
7.73
8.20
8.3
6.7
6.5
7.23
% Organic
Matter
3.11
1.70
0.86
0.5
3.0
2.1
1.6
Toxicity Parameter
MATC
MATC
MATC
MATC
MATC
MATC
MATC
Pb in Soil
(mg/kg dw)
141
71
71
283
424
158
2,263
Studies not eligible for Eco-SSL derivation
Loblolly pine (Finns taeda)
Red oak (Quercus rubra)
Spinach
Alfalfa (Medicago sativa)
Alfalfa
Alfalfa
Radish (Raphanus sativus)
Radish
Radish
Onion (Allium cepa)
Radish
Carrot (Daucus carota)
Peas (Pisum sativum)
Barley (Hordeum vulgare)
Alfalfa
Tomato
Spinach
Radish
Radish
5.5
6
6.7
6.4
6.9
6.9
6.9
6.9
6.9
8.3
5.1
7.0
7.0
6.0
6.9
7.45
6.7
6.2
7.1
3.4
1.5
0.0
1.0
1.7
1.7
1.0
1.0
1.0
0.5
8.0
0.6
0.6
2.5
4.8
2.06
8.0
8.0
8.0
NOAEC
LOAEC
NOAEC
NOAEC
NOAEC
NOAEC
LOAEC
LOAEC
LOAEC
LOAEC
NOAEC
NOAEC
NOAEC
NOAEC
NOAEC
MATC
NOAEC
NOAEC
NOAEC
480
100
600
250
250
250
500
100
100
50
600
85
85
1,000
250
35
600
600
600
*MATC = Maximum Acceptable Threshold Concentration, or the geometric mean of the NOAEC
(no-observed-adverse-effect concentration) and LOAEC (lowest-observed-adverse-effect concentration).
Source: U.S. Environmental Protection Agency (2005b).
May 2006
AX8-63
DRAFT-DO NOT QUOTE OR CITE
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1 Environmental Protection Agency, 2005b). The relevant information from the Eco-SSL
2 document (U.S. Environmental Protection Agency, 2005b) is described below. A literature
3 search and review was conducted to identify critical papers published since 2002. These recent
4 critical papers are described briefly below. No studies were found that used inhalation exposures
5 to evaluate endpoints such as survival, growth, and reproduction in birds or mammals. All
6 studies described below exposed organisms via ingestion (drinking water or diet) or gavage.
7 The Eco-SSLs for avian and mammalian consumers are presented as Pb concentrations in
8 soil. These concentrations were calculated by assuming exposure to Pb via incidental soil
9 ingestion and ingestion of Pb-contaminated food, and using a NOAEL as the TRY (U.S.
10 Environmental Protection Agency, 2005a). A simplified version of the equation used to
1 1 calculate the Eco-SSL is:
12
14 HQ = rrCgnjj x IRsoiT) + (C^ x IR^I / BW (8-4)
16 TRY
17
18 where:
19
20 HQ = hazard quotient (1 mg Pb/kg bw/day)
21 CSoii = concentration of Pb in soil (mg Pb/kg soil)
22 IRsoii = incidental soil ingestion rate (kg soil/day)
23 Cfood = concentration of Pb in food (mg Pb/kg food)
24 IRfood = food ingestion rate (kg food/day)
25 BW = body weight (kg)
26 TRY = toxicity reference value (mg Pb/kg bw/day)
27
28 Food ingestion was estimated by modeling the uptake of Pb from soil into each diet
29 component (e.g., vegetation, invertebrates, etc.). Bioavailability of Pb in soil and food was
30 assumed to be 100%. The Eco-SSL is equivalent to the concentration of Pb in soil that results in
31 an HQ = 1 . The two factors that may have the most significant influence on the resulting Eco-
32 SSL are the assumption of 100% bioavailability of Pb in soil and diet and the selection of the
33 TRY. The toxicity data that were reviewed to develop the TRY are presented in the following
34 subsections.
35 Representative avian and mammalian wildlife species were selected for modeling Pb
36 exposures to wildlife with different diets and calculating the Eco-SSL. The avian species
May 2006 AX8-64 DRAFT-DO NOT QUOTE OR CITE
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1 selected were dove (herbivore), woodcock (insectivore), and hawk (carnivore). The mammalian
2 species selected were vole (herbivore), shrew (insectivore), and weasel (carnivore). The lowest
3 of the three back-calculated soil concentrations, which resulted in an HQ = 1, was selected as the
4 Eco-SSL. For Pb, the lowest values were for the insectivorous species of bird and mammal.
5
6 Avian Consumers
1 Effects on birds observed in studies conducted since the 1986 Pb AQCD (U.S.
8 Environmental Protection Agency, 1986a) are similar to those reported previously: mortality,
9 changes in juvenile growth rate and weight gain, effects on various reproductive measures, and
10 changes in behavior (U.S. Environmental Protection Agency, 2005b). Reproductive effects
11 following Pb exposure included declines in clutch size, number of young hatched, and number of
12 young fledged as well as decreased fertility or eggshell thickness. Few significant reproductive
13 effects have been reported in birds at Pb concentrations below 100 mg/kg in the diet
14 (Scheuhammer, 1987).
15 The literature search completed for Eco-SSL development identified 2,429 papers for
16 detailed review for either avian or mammalian species, of which 54 met the minimum criteria for
17 further consideration for avian Eco-SSL development (U.S. Environmental Protection Agency,
18 2005b). The 106 toxicological data points for birds that were further evaluated included
19 biochemical, behavioral, physiological, pathological, reproductive, growth, and survival effects.
20 Growth and reproduction data were used to derive the Eco-SSL (Table AX8-1.4.3; Figure
21 AX8-1.4.1). The geometric mean of the NOAELs was calculated as 10.9 mg/kg-day, which was
22 higher than the lowest bounded LOAEL (the term "bounded" means that both a NOAEL and
23 LOAEL were obtained from the same study). Therefore, the highest bounded NOAEL that was
24 lower than the lowest bounded LOAEL for survival, growth, or reproduction (1.63 mg Pb/kg bw-
25 day) was used as the TRV (U.S. Environmental Protection Agency, 2005b). The TRV was used
26 to back-calculate the Eco-SSL of 11 mg/kg soil for avian species (U.S. Environmental Protection
27 Agency, 2005b). For more information on the rationale for selecting TRVs, please refer to U.S.
28 Environmental Protection Agency (2003).
29 Many of the toxicity data presented in the Eco-SSL document (U.S. Environmental
30 Protection Agency, 2005b) are lower than those discussed in the 1986 Pb AQCD. The TRV and
31 resulting Eco-SSL were derived using many conservative assumptions. For example, the EPA
May 2006 AX8-65 DRAFT-DO NOT QUOTE OR CITE
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Table AX8-1.4.3. Avian Toxicity Data Used to Develop the Eco-SSL
^H
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ON
^
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6
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n
s.**
H
O
O
H
W
O
o
HH
H
W
Avian
Species
Reproduction
Japanese
quail
Chicken
Chicken
Mallard
American
kestrel
Japanese
quail
Japanese
quail
Japanese
quail
Japanese
quail
Japanese
quail
Chicken
Ringed
turtle dove
Japanese
quail
Japanese
quail
No. of
Doses
4
3
4
2
3
5
5
3
5
4
5
2
2
2
Route of
Exposure
FD
FD
FD
FD
FD
FD
FD
FD
FD
FD
FD
DR
FD
FD
Exposure
Duration
5
4
30
76
6
5
5
32
12
12
10
11
1
27
Dura-
tion
Units Age
w 6
w NR
d 22
d NR
mo 1-6
w 6
w 1
d NR
w 0
w NR
w NR
w NR
w 14
d NR
Age
Units Lifestage Sex
w LB F
NR LB F
w LB F
NR SM F
yr AD F
d JV M
d JV M
NR AD F
d LB B
NR LB F
NR LB F
NR AD M
w JV F
NR AD F
Effect
Type
REP
REP
EGG
EGG
REP
REP
REP
REP
REP
REP
REP
REP
REP
REP
Effect
Measure
PROG
PROG
ALWT
ESTH
RSUC
TEWT
TEWT
PROG
EGPN
PROG
PROG
TEWT
TPRD
PROG
NOAEL
Response (nig/kg
Site bw/day)
WO 0.194
WO 1.63
EG 2.69
EG 5.63
WO 12.0
TE 12.6
TE 67.4
WO 125
EG
WO
WO
TE
WO
WO
LOAEL
(mg/kg
bw/day)
1.94
3.26
4.04
126
135
0.110
0.194
3.26
11.8
93.1
377
-------
Table AX8-1.4.3 (cont'd). Avian Toxicity Data Used to Develop the Eco-SSL
to
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\^
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H
6
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0
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O
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W
o
O
HH
H
W
Avian
Species
Growth
Japanese
quail
Japanese
quail
Japanese
quail
Japanese
quail
Chicken
Chicken
Japanese
quail
Japanese
quail
Japanese
quail
Japanese
quail
Chicken
Duck
American
kestrel
Chicken
Japanese
quail
No. of
Doses
3
3
2
3
4
4
5
5
5
5
2
3
4
2
5
Route of
Exposure
FD
FD
FD
FD
FD
FD
FD
FD
FD
FD
FD
GV
GV
FD
FD
Exposure
Duration
5
2
2
4
4
4
12
12
2
1
21
3
10
20
14
Dura-
tion
Units
w
w
w
w
w
w
w
w
w
w
d
mo
d
d
d
Age
i
i
i
0
4
4
0
1
6
1
1
24
1
1
1
Age
Units
d
d
d
d
w
w
d
w
d
d
d
w
d
d
d
Lifestage
JV
JV
JV
JV
JV
JV
JV
JV
JV
JV
JV
MA
JV
JV
JV
Sex
F
B
NR
F
NR
NR
F
F
NR
NR
B
F
NR
B
B
Effect
Type
GRO
GRO
GRO
GRO
GRO
GRO
GRO
GRO
GRO
GRO
GRO
GRO
GRO
GRO
GRO
Effect
Measure
BDWT
BDWT
BDWT
BDWT
BDWT
BDWT
BDWT
BDWT
BDWT
BDWT
BDWT
BDWT
BDWT
BDWT
BDWT
Response
Site
wo
wo
wo
wo
wo
wo
wo
wo
wo
wo
wo
wo
wo
wo
wo
NOAEL
(mg/kg
bw/day)
1.56
2.77
4.64
5.93
6.14
7.10
11.1
11.2
12.6
13.5
14.2
20.0
25.0
28.4
34.5
LOAEL
(mg/kg
bw/day)
15.6
59.3
61.4
71.0
111
112
126
67.4
125
-------
Table AX8-1.4.3 (cont'd). Avian Toxicity Data Used to Develop the Eco-SSL
to
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ON
X
oo
ON
oo
O
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6
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0
H
O
O
H
W
O
O
HH
H
W
Dura-
Avian
Species
American
kestrel
Chicken
Mallard
Chicken
Chicken
Japanese
quail
Chicken
Chicken
Chicken
Chicken
Chicken
Chicken
Chicken
Chicken
Chicken
AD = adult
thinning; F
or numbers
Source: U.
No. of Route of
Doses Exposure
4 FD
5 FD
4 FD
5 FD
2 FD
3 FD
2 FD
3 FD
4 FD
2 FD
2 OR
2 FD
2 FD
2 FD
2 FD
; ALWT = albumin weight; B
= female; FD = food; GRO =
; REP = reproduction; RSUC
Exposure
Duration
60
2
8
20
3
32
19
2
14
20
4
7
2
7
14
tion
Units Age
d
w
d
d
w
d
d
w
d
d
w
d
w
d
d
1-2
1
9
1
1
NR
1
1
8
1
NR
1
1
1
8
Age
Units
yr
d
d
d
d
NR
d
d
d
d
NR
d
d
d
d
Lifestage
AD
JV
JV
JV
JV
AD
JV
JV
JV
JV
JV
JV
JV
IM
JV
Sex
B
M
NR
M
M
F
M
M
M
M
B
M
M
NR
M
NOAEL LOAEL
Effect Effect Response (mg/kg (mg/kg
Type Measure Site bw/day) bw/day)
GRO BDWT
GRO BDWT
GRO BDWT
GRO BDWT
GRO BDWT
GRO BDWT
GRO BDWT
GRO BDWT
GRO BDWT
GRO BDWT
GRO BDWT
GRO BDWT
GRO BDWT
GRO BDWT
GRO BDWT
WO
WO
WO
WO
WO
WO
WO
WO
WO
WO
WO
WO
WO
wo
wo
54.3
61.3 123
66.9
38.2
53.1
64.3
76.3
124
152
163
200
262
270
273
282
= both; BDWT = body weight changes; d = days; DR = drinking water; EG = egg; EGG = effects on eggs; EGPN = egg production; ESTH = eggshell
growth; GV = gavage; JV = juvenile; LB = laying bird; MA = mature; M = male; mo = months; NR = not reported; OR = other oral; PROG = progeny counts
= reproductive success; SM = sexually mature; TE = testes; TEWT = testes weight; TPRD = total production; w = weeks; WO = whole organism; yr = years.
S. Environmental Protection Agency (2005b)
-------
IUUU
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ro 100
^
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0 1
Q
n -i
A
oooo
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AA^o o o o
A Q
oo°A o .»• Qo
• °
• •
o A •••!
• • ii
«** Legend:
A . A
A • Growth NOAEL
0 Growth LOAEL
A Reproduction NOAEL
A Reproduction LOAEL
A A - Eco-SSL (mg/kg dw)
A
Reproduction and Growth
Figure AX8-1.4.1. Avian reproduction and growth toxicity data considered in
development of the Eco-SSL.
Source: U.S. Environmental Protection Agency (2005b).
1 (U.S. Environmental Protection Agency, 2005b) recognizes that toxicity is observed over a wide
2 range of doses (<1 to >100 mg Pb/kg bw/day), even when considering only reproductive effects
3 in the same species. In addition, the TRY of 1.63 mg/kg-day is lower than most of the reported
4 doses that have been associated with measured effects. This is true for not only survival, growth,
5 and reproductive effects but also biochemical, behavioral, physiological, and pathological
6 effects, which generally are observed at lower concentrations than effects on growth or
7 reproduction. In addition, the Eco-SSL was back-calculated using conservative modeling
8 assumptions. Therefore, the Eco-SSL of 11 mg/kg may be considered a conservative value.
9 Very little research has been done to expand the knowledge of the toxicity of Pb to birds
10 since the Eco-SSL work was done. However, several studies have been conducted on waterfowl.
11 Toxicity data for waterfowl (in particular, mallards) were included in the soil Eco-SSL
12 development process (Table AX8-1.4.3), although mallards may be more exposed to
13 contaminants in sediment than soil. Effects on waterfowl may vary depending on the form of Pb,
May 2006
AX8-69
DRAFT-DO NOT QUOTE OR CITE
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1 characteristics of the sediment, the foraging strategy of the species (which may vary during
2 reproduction), and the nutritional status of the animal. Sediment is recognized as an important
3 route of exposure for waterfowl, particularly those species that dabble (i.e., forage on
4 invertebrates in the sediment) (Beyer et al., 2000; Douglas-Stroebel et al., 2005). Douglas-
5 Stroebel et al. (2005) found that mallard ducklings exposed to Pb-contaminated sediment and a
6 low nutrition diet exhibited more changes in behavior (as measured by time bathing, resting, and
7 feeding) than Pb exposure or low nutrition exposure alone. These effects may be due to the low
8 nutrition diet being deficient in levels of protein, amino acids, calcium, zinc, and other nutrients.
9 Beyer et al. (2000) related blood Pb to sublethal effects in waterfowl along the Coeur
10 d'Alene River near a mining site in Idaho. The authors suggested that 0.20 mg/kg ww blood Pb
11 represents the no-effect level. This no-effect blood concentration corresponds to a sediment Pb
12 concentration of 24 mg/kg. A sediment concentration of 530 mg/kg, associated with a blood Pb
13 concentration of 0.68 mg/kg ww, is suggested to be the lowest-effect concentration. These
14 results are consistent with those of Scheuhammer (1989) who found blood Pb concentrations of
15 0.18 |ig/mL to 0.65 |ig/mL in mallards corresponded to conditions associated with greater than
16 normal exposure to Pb, but that that should not be considered Pb poisoning. The study by Beyer
17 et al. (2000) related blood Pb to waterfowl mortality and concluded that some swan mortality
18 may occur at blood Pb levels of 1.9 mg/kg ww, corresponding to a sediment Pb concentration of
19 1800 mg/kg. Using the mean blood level of 3.6 mg/kg ww from all moribund swans in the
20 study, it was predicted that half of the swans consuming sediment at the 90th percentile rate
21 would die with chronic exposure to sediment concentrations of 3600 mg/kg.
22
23 Mammalian Consumers
24 Effects on mammals observed in studies conducted since the 1986 AQCD (U.S.
25 Environmental Protection Agency, 1986a) are similar to those reported previously: mortality,
26 effects on reproduction, developmental effects, and changes in growth (U.S. Environmental
27 Protection Agency, 2005b). Very little research has been done to expand the knowledge of the
28 toxicity of Pb to mammalian wildlife, since the Eco-SSL work was done. Most studies
29 conducted on mammals use laboratory animals to study potential adverse effects of concern for
30 humans, and such studies are summarized in other sections of this document.
May 2006 AX8-70 DRAFT-DO NOT QUOTE OR CITE
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1 Of the 2,429 papers identified in the literature search for Eco-SSL development, 219 met
2 the minimum criteria for further consideration for mammalian Eco-SSL development (U.S.
3 Environmental Protection Agency, 2005b). The 343 ecotoxicological endpoints for mammals
4 that were further evaluated included biochemical, behavioral, physiological, pathological,
5 reproductive, growth, and survival effects. Growth and reproduction data were used to derive
6 the Eco-SSL (Table AX8-1.4.4, Figure AX8-1.4.2). The geometric mean of the NOAELs was
7 calculated as 40.7 mg/kg-day, which was higher than the lowest bounded LOAEL for survival,
8 growth, or reproduction. Therefore, the highest bounded NOAEL that was lower than the lowest
9 bounded LOAEL for survival, growth, or reproduction (4.7 mg Pb/kg bw-day) was used as the
10 TRY (U.S. Environmental Protection Agency, 2005b). The TRY was used to back-calculate the
11 Eco-SSL of 56 mg/kg soil (U.S. Environmental Protection Agency, 2005b). For more
12 information on the rationale for selecting TRVs, please refer to U.S. Environmental Protection
13 Agency (2003).
14 A review of the data presented in the Eco-SSL document (U.S. Environmental Protection
15 Agency, 2005b) reveals that effects on survival generally are observed at Pb doses much greater
16 than those reported in the 1986 Pb AQCD, where it was concluded that most animals would die
17 when consuming a regular dose of 2 to 8 mg Pb/kg bw-day (U.S. Environmental Protection
18 Agency, 1986a). However, the data presented in the Eco-SSL document (U.S. Environmental
19 Protection Agency, 2005b) generally do not support this. While five studies reported decreased
20 survival at these levels, 34 other studies reported no mortality or a LOAEL for mortality at
21 significantly higher doses (U.S. Environmental Protection Agency, 2005b). The five studies that
22 supported this low toxic level were conducted on three species (mouse, rat, and cow) and used
23 either gavage or drinking water as the exposure method. The 34 other studies included data on
24 these three species as well as five other species (rabbit, dog, pig, hamster, and shrew) and
25 included gavage and drinking water as well as food ingestion exposure methods. The NOAELs
26 for survival ranged from 3.5 to 3200 mg/kg-day (U.S. Environmental Protection Agency,
27 2005b). Therefore, the review of data in the Eco-SSL document suggests effects on survival of
28 wildlife generally would occur at doses greater than the 2 to 8 mg/kg-day reported to be toxic to
29 most animals in the 1986 Pb AQCD (U.S. Environmental Protection Agency, 1986a).
30
May 2006 AX8-71 DRAFT-DO NOT QUOTE OR CITE
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Table AX8-1.4.4. Mammalian Toxicity Data Used to Develop the Eco-SSL
to
o
o^
>
X
-------
Table AX8-1.4.4 (cont'd). Mammalian Toxicity Data Used to Develop the Eco-SSL
to
o
Oi
X
oo
1
OJ
o
f?
H
6
o
0
H
O
o
H
W
O
O
H
W
Mammalian
Species
Hamster
Hamster
Rat
Rat
Rat
Rat
Rat
Mouse
Mouse
Mouse
Rat
Rat
Rat
Rat
Rat
Rat
Mouse
Rat
Rat
Mouse
Mouse
Rat
No.
of
Doses
2
2
4
5
3
4
2
3
7
7
4
5
2
2
4
4
4
3
2
2
4
2
Route of
Exposure
DR
DR
DR
GV
DR
DR
DR
FD
FD
FD
DR
DR
GV
DR
FD
DR
GV
FD
DR
DR
GV
DR
Exposure
Duration
51
14
37
12
68
77
21
8
30
30
21
10
102
9
4
13
60
339
9
6
52
120
Duration
Units
d
d
d
d
d
d
d
w
d
d
d
w
d
mo
d
w
d
d
mo
mo
d
d
Age
15
11
NR
NR
25
25
NR
2
NR
NR
NR
NR
30
NR
NR
NR
NR
26-27
21
21
2
1
Age
Units
w
w
NR
NR
d
d
NR
mo
NR
NR
NR
NR
d
NR
NR
NR
NR
d
d
d
mo
d
Lifestage
GE
GE
GE
GE
GE
GE
LC
GE
LC
LC
GE
AD
GE
SM
LC
JV
AD
JV
JV
JV
GE
GE
Sex
F
F
F
F
F
F
F
M
F
F
F
M
F
M
F
M
F
B
F
F
F
M
Effect
Type
REP
REP
REP
REP
REP
REP
REP
REP
REP
REP
REP
REP
REP
REP
REP
REP
REP
REP
REP
REP
REP
REP
Effect
Measure
PROG
PROG
PRWT
RSEM
PRWT
PRWT
PRWT
SPCV
PRWT
PRWT
DEYO
TEWT
PRWT
RHIS
PRWT
PERT
RPRD
PRWT
DEYO
DEYO
PROG
SPCL
Response
Site
wo
wo
wo
EM
WO
wo
wo
TE
WO
WO
WO
MT
WO
TE
WO
WO
0V
WO
WO
WO
EM
TE
NOAEL
(mg/kg
bw/day)
64.8
64.9
90.1
100
115
116
120
144
202
202
276
294
441
600
601
639
—
—
LOAEL
(mg/kg
bw/day)
—
—
270
150
—
—
—
1,440
506
506
552
587
—
—
1,500
2.00
2.49
2.94
3.62
5.50
6.76
-------
Table AX8-1.4.4 (cont'd). Mammalian Toxicity Data Used to Develop the Eco-SSL
to
o
Oi
X
oo
1
o
f?
H
6
o
0
H
O
o
H
W
O
O
H
W
Mammalian
Species
Mouse
Mouse
Rat
Rat
Rat
Rat
Mouse
Mouse
Mouse
Mouse
Mouse
Rat
Rat
Rat
Rat
Rat
Rat
Rat
Rat
Rat
Rat
Rat
No.
of
Doses
2
2
2
2
2
3
2
2
2
2
4
2
5
2
3
2
2
4
2
4
3
2
Route of
Exposure
DR
GV
FD
GV
DR
DR
DR
DR
DR
DR
FD
DR
DR
DR
DR
DR
DR
FD
FD
FD
FD
DR
Exposure
Duration
5
2
102
3
18
90
23
62
18
12
18
4
6
22
30
13
21
3
1
3
25
21
Duration
Units
w
w
d
mo
d
d
d
d
w
w
d
w
w
d
d
w
d
w
w
w
d
d
Age
NR
NR
NR
8
NR
NR
NR
NR
6-8
9
NR
99
4
NR
52
NR
80
NR
19
NR
NR
NR
Age
Units
NR
NR
NR
w
NR
NR
NR
NR
w
w
NR
d
mo
NR
d
NR
d
NR
w
NR
NR
NR
Lifestage
AD
JV
GE
SM
GE
AD
GE
GE
LC
SM
GE
JV
GE
GE
JV
GE
JV
LC
LC
LC
LC
LC
Sex
M
M
F
M
F
M
F
F
F
M
F
M
F
F
M
F
F
F
F
F
F
F
Effect
Type
REP
REP
REP
REP
REP
REP
REP
REP
REP
REP
REP
REP
REP
REP
REP
REP
REP
REP
REP
REP
REP
REP
Effect
Measure
TEDG
SPCL
PROG
TEDG
PRWT
SPCL
PRWT
PRWT
PRWT
PRFM
PRWT
SPCL
RHIS
PRWT
GREP
PRWT
PRWT
PRWT
PRWT
PRWT
PRWT
PRWT
Response
Site
TE
SM
WO
TE
WO
SM
WO
wo
wo
wo
wo
SM
WO
wo
PG
WO
WO
wo
wo
wo
wo
wo
NOAEL LOAEL
(mg/kg (mg/kg
bw/day) bw/day)
— 16.6
— 46.4
— 49.6
— 50.0
— 55.5
— 61.2
— 78.6
— 99.8
— 137
— 139
— 154
— 171
— 175
— 178
— 198
— 200
— 218
— 221
— 222
— 230
— 258
— 330
-------
Table AX8-1.4.4 (cont'd). Mammalian Toxicity Data Used to Develop the Eco-SSL
to
o
Oi
X
oo
1
o
f?
H
6
o
0
H
O
o
H
W
O
O
H
W
Mammalian
Species
Rat
Rat
Rat
Rat
Rat
Mouse
Mouse
Rat
Mouse
Rat
Mouse
Rat
Rat
Rat
Mouse
Rat
Rat
Mouse
Rat
Rat
Rat
Mouse
No.
of
Doses
2
2
2
2
2
2
2
2
2
2
2
2
2
2
2
2
2
2
2
2
2
4
Route of
Exposure
DR
DR
DR
DR
DR
DR
DR
DR
DR
DR
DR
DR
DR
DR
FD
DR
DR
DR
FD
FD
FD
DR
Exposure
Duration
30
17
24
12
30
44
14
50
45
22
48
9
9
3
7
126
20
4
2
7
21
18
Duration
Units
d
d
d
d
d
d
d
d
d
d
d
mo
mo
w
d
d
w
d
w
d
d
w
Age
52
NR
NR
NR
27
NR
NR
24
50-100
NR
NR
3
NR
14
NR
1
10
NR
NR
NR
NR
11
Age
Units
d
NR
NR
NR
d
NR
NR
d
d
NR
NR
mo
NR
w
NR
d
w
NR
NR
NR
NR
w
Lifestage
JV
GE
LC
GE
JV
GE
LC
JV
GE
GE
GE
SM
SM
LC
GE
GE
GE
LC
LC
LC
LC
JV
Sex
M
F
F
F
M
F
F
F
F
F
F
M
M
F
F
F
F
F
F
F
F
F
Effect
Type
REP
REP
REP
REP
REP
REP
REP
REP
REP
REP
REP
REP
REP
REP
REP
REP
REP
REP
REP
REP
REP
REP
Effect
Measure
SPCL
PRWT
PRWT
PRWT
SPCL
PRWT
PRWT
RBEH
ODVP
PRWT
PRWT
SPCL
TEDG
PRWT
RSUC
PROG
PRWT
PRWT
PRWT
PRWT
PRWT
TEWT
Response
Site
SM
WO
wo
wo
SM
WO
wo
wo
wo
wo
wo
TE
TE
WO
EM
WO
WO
WO
wo
wo
wo
wo
NOAEL LOAEL
(mg/kg (mg/kg
bw/day) bw/day)
— 354
— 360
— 360
— 362
— 364
— 381
— 381
— 381
— 404
— 420
— 437
— 579
— 600
— 635
— 646
— 651
— 750
— 762
— 828
— 833
— 991
— 1,370
-------
Table AX8-1.4.4 (cont'd). Mammalian Toxicity Data Used to Develop the Eco-SSL
to
o
ON
X
oo
1
ON
O
f?
H
6
o
0
H
O
o
H
W
O
O
H
W
Mammalian
Species
Rat
Mouse
Rat
Rat
Rat
Rat
Mouse
Rat
Growth
Horse
Rat
Rat
Rat
Cattle
Rat
Rat
Rat
Dog
Rat
Rat
Cattle
Rat
No.
of
Doses
2
2
2
2
2
M
2
2
2
2
6
5
4
3
2
4
3
3
4
3
2
Route of
Exposure
FD
DR
FD
FD
FD
FD
DR
FD
FD
FD
DR
DR
OR
DR
DR
DR
FD
DR
DR
OR
OR
Exposure
Duration
30
14
16
7
25
27
14
17
15
21
21
7
7
14
332
7
7
30
23
84
6
Duration
Units
d
w
d
d
d
d
w
d
w
d
d
d
w
d
d
w
mo
d
d
d
w
Age
NR
NR
NR
NR
NR
NR
21
NR
20-21
0
NR
50
1
21
28
21
NR
22-24
22
NR
NR
Age
Units
NR
NR
NR
NR
NR
NR
d
NR
w
d
NR
d
w
d
d
d
NR
d
d
NR
NR
Lifestage
LC
GE
LC
LC
LC
LC
JV
LC
JV
JV
GE
AD
JV
JV
JV
GE
JV
JV
JV
JV
AD
Sex
F
B
F
F
F
C
B
F
M
F
F
F
M
F
B
F
NR
M
F
M
M
Effect
Type
REP
REP
REP
REP
REP
REP
REP
REP
GRO
GRO
GRO
GRO
GRO
GRO
GRO
GRO
GRO
GRO
GRO
GRO
GRO
Effect
Measure
PRWT
PROG
PROG
PRWT
PRWT
PROG
PROG
PRWT
BDWT
BDWT
BDWT
BDWT
BDWT
BDWT
BDWT
BDWT
BDWT
BDWT
BDWT
BDWT
BDWT
Response
Site
wo
wo
wo
wo
wo
wo
wo
wo
wo
wo
wo
wo
wo
wo
wo
wo
wo
wo
wo
wo
wo
NOAEL
(mg/kg
bw/day)
—
—
—
—
—
—
—
—
0.15
0.5
1.00
1.27
1.99
2.40
2.98
4.70
4.71
5.64
5.80
7.79
9.10
LOAEL
(mg/kg
bw/day)
1,770
1,990
2,570
2,570
2,570
2,840
3,630
6,170
—
5.00
13.0
—
8.90
28.2
29.0
—
-------
Table AX8-1.4.4 (cont'd). Mammalian Toxicity Data Used to Develop the Eco-SSL
to
o
Oi
X
oo
1
o
f?
H
6
o
0
H
O
o
H
W
O
O
H
W
Mammalian
Species
Rat
Rat
Rabbit
Rat
Rat
Rat
Rat
Rat
Mouse
Rat
Rat
Rat
Rat
Rat
Sheep
Rat
Cattle
Rat
Rat
Rat
Rat
Rat
No.
of
Doses
2
3
3
2
2
2
2
2
3
4
3
4
7
4
5
2
4
2
5
4
2
6
Route of
Exposure
GV
DR
GV
DR
DR
FD
OR
DR
DR
GV
FD
GV
DR
DR
FD
DR
FD
GV
DR
GV
FD
DR
Exposure
Duration
8
6
10
140
6
10
6
7
14
9
339
29
10
56
84
10
7
28
4
12
4
10
Duration
Units
w
mo
d
d
w
w
w
w
d
w
d
d
w
d
d
w
w
d
w
d
w
w
Age
NR
NR
1
21
NR
NR
NR
NR
0
10
26-27
NR
NR
70
NR
NR
16
2
94
2
NR
NR
Age
Units
NR
NR
d
d
NR
NR
NR
NR
d
w
d
NR
NR
d
NR
NR
w
d
d
d
NR
NR
Lifestage
JV
AD
JV
JV
JV
JV
AD
JV
JV
JV
JV
SM
JV
LC
JV
JV
JV
JV
JV
JV
JV
JV
Sex
F
M
F
M
M
M
M
M
NR
M
B
F
M
F
M
M
M
B
M
B
M
M
Effect
Type
GRO
GRO
GRO
GRO
GRO
GRO
GRO
GRO
GRO
GRO
GRO
GRO
GRO
GRO
GRO
GRO
GRO
GRO
GRO
GRO
GRO
GRO
Effect
Measure
BDWT
BDWT
BDWT
BDWT
BDWT
BDWT
BDWT
BDWT
BDWT
BDWT
BDWT
BDWT
BDWT
BDWT
BDWT
BDWT
BDWT
BDWT
BDWT
BDWT
BDWT
BDWT
Response
Site
wo
wo
wo
wo
wo
wo
wo
wo
wo
wo
wo
wo
wo
wo
wo
wo
wo
wo
wo
wo
wo
wo
NOAEL
(mg/kg
bw/day)
10.0
10.6
10.7
10.7
15.1
15.4
15.5
16.1
16.3
18.0
18.3
18.9
24.3
32.5
32.7
38.5
43.0
50.0
71.5
75.0
100
120
LOAEL
(mg/kg
bw/day)
—
532
50.4
—
—
—
—
—
163
180
—
—
—
—
—
178
225
—
383
-------
Table AX8-1.4.4 (cont'd). Mammalian Toxicity Data Used to Develop the Eco-SSL
to
o
Oi
X
OO
1
oo
o
f?
H
6
O
0
H
O
o
H
W
O
O
H
W
Mammalian
Species
Mouse
Mouse
Mouse
Rat
Rat
Rat
Mouse
Rat
Rat
Rat
Rat
Rat
Mouse
Rat
Rat
Rat
Rat
Rat
Mouse
Mouse
Rat
Rat
No.
of
Doses
3
2
2
3
2
4
3
4
2
2
4
4
5
2
4
5
2
2
4
7
2
2
Route of
Exposure
FD
DR
DR
DR
DR
GV
DR
GV
GV
DR
FD
DR
DR
DR
GV
GV
FD
GV
GV
FD
DR
DR
Exposure
Duration
4
18
12
30
4
18
6
18
91
21
1
30
10
30
14
14
14
102
12
30
126
20
Duration
Units
w
w
w
d
w
d
w
d
d
d
w
d
w
d
d
d
mo
d
d
d
d
w
Age
3
6-8
NR
52
99
3
7
2
NR
80
NR
NR
NR
52
14
20
0
30
6
NR
1
10
Age
Units
mo
w
NR
d
d
d
w
d
NR
d
NR
NR
NR
d
d
d
d
d
d
NR
d
w
Lifestage
JV
LC
GE
JV
JV
JV
SM
JV
JV
JV
LC
JV
JV
JV
JV
JV
JV
LC
JV
LC
GE
GE
Sex
B
F
M
M
B
M
M
B
M
F
F
M
M
M
NR
NR
NR
F
M
F
F
F
Effect
Type
GRO
GRO
GRO
GRO
GRO
GRO
GRO
GRO
GRO
GRO
GRO
GRO
GRO
GRO
GRO
GRO
GRO
GRO
GRO
GRO
GRO
GRO
Effect
Measure
BDWT
BDWT
BDWT
BDWT
BDWT
BDWT
BDWT
BDWT
BDWT
BDWT
BDWT
BDWT
BDWT
BDWT
BDWT
BDWT
BDWT
BDWT
BDWT
BDWT
BDWT
BDWT
Response
Site
wo
wo
wo
wo
wo
wo
wo
wo
wo
wo
wo
wo
wo
wo
wo
wo
wo
wo
wo
wo
wo
wo
NOAEL
(mg/kg
bw/day)
136
137
139
169
171
180
187
200
200
218
230
285
362
364
400
400
431
441
534
632
651
750
LOAEL
(mg/kg
bw/day)
1360
—
—
508
—
—
373
—
—
460
—
—
—
800
800
—
1264
—
-------
Table AX8-1.4.4 (cont'd). Mammalian Toxicity Data Used to Develop the Eco-SSL
to
o
Oi
X
oo
1
VO
o
f?
H
6
O
0
H
O
O
H
W
O
O
H
W
Mammalian
Species
Mouse
Rat
Rat
Cattle
Rat
Rat
Rat
Rat
Rat
Rat
Mouse
Dog
Shrew
Rat
Pig
Rat
Rat
Rat
Rat
Rat
Rat
Rat
No.
of
Doses
7
4
2
2
3
4
2
2
2
2
2
2
4
3
2
2
2
2
2
2
2
4
Route of
Exposure
FD
FD
DR
FD
DR
DR
DR
DR
DR
DR
GV
OR
FD
GV
FD
GV
FD
GV
DR
DR
GV
GV
Exposure
Duration
28
18
9
283
92
7
5
26
14
10
3
5
31
58
13
29
5
6
30
50
28
14
Duration
Units
d
d
d
d
d
d
d
d
d
d
w
w
d
d
w
d
w
d
d
d
d
d
Age
NR
NR
21
7
25
25
26
22
26
26
NR
<1
NR
2
4
2
NR
1
27
24
2
18
Age
Units Lifestage
NR LC
NR LC
d JV
mo JV
d GE
d GE
d JV
d JV
d JV
d JV
NR JV
yr JV
NR JV
d JV
w JV
d JV
NR MA
d JV
d JV
d JV
d JV
d JV
Sex
F
F
M
M
F
F
F
F
F
F
M
NR
B
B
NR
F
NR
B
M
M
M
NR
Effect
Type
GRO
GRO
GRO
GRO
MPH
GRO
GRO
GRO
MPH
GRO
GRO
GRO
GRO
GRO
GRO
GRO
GRO
GRO
GRO
GRO
GRO
GRO
Effect
Measure
BDWT
BDWT
BDWT
BDWT
GMPH
BDWT
BDWT
BDWT
Other
BDWT
BDWT
BDWT
BDWT
BDWT
BDWT
BDWT
BDWT
BDWT
BDWT
BDWT
BDWT
BDWT
Response
Site
wo
wo
wo
wo
TB
WO
WO
WO
TA
WO
WO
WO
WO
wo
wo
wo
wo
wo
wo
wo
wo
wo
NOAEL LOAEL
(mg/kg (mg/kg
bw/day) bw/day)
1260 2530
1500
— 3.30
— 15.0
— 28.7
— 29.0
— 29.0
— 29.5
— 29.9
— 30.4
— 46.4
— 50.0
— 61.5
— 100
— 173
— 200
— 272
— 328
— 354
— 371
— 400
— 400
-------
Table AX8-1.4.4 (cont'd). Mammalian Toxicity Data Used to Develop the Eco-SSL
to
o
o
Oi
X
OO
1
oo
o
DRAFT-DO I
^
0
H
O
O
H
W
O
O
HH
H
W
No.
Mammalian of
Species Doses
Mouse
Rat
Rat
Mouse
Rat
Rat
Rat
Rat
Rat
Rat
Rat
2
4
2
4
2
2
2
4
2
3
2
Route of Exposure
Exposure Duration
DR
FD
DR
DR
FD
GV
FD
GV
FD
GV
FD
45
1
6
10
21
18
2
14
2
14
14
NOAEL
Duration Age Effect Effect Response (mg/kg
Units Age Units Lifestage Sex Type Measure Site bw/day)
d 50-100
w NR
w 14
w 11
d NR
d 2
w 0
d 24
w 60-80
d 16
d 60
d
NR
w
w
NR
d
d
d
d
d
d
GE F GRO
LC F GRO
LC F GRO
JV F GRO
LC F GRO
JV B GRO
JV NR GRO
JV NR GRO
JV M GRO
JV NR GRO
JV M GRO
BDWT WO —
BDWT WO —
BDWT WO —
BDWT WO —
BDWT WO —
BDWT WO —
BDWT WO —
BDWT WO —
BDWT WO —
BDWT WO —
BDWT WO —
LOAEL
(mg/kg
bw/day)
404
442
638
748
991
1000
1430
1600
2390
2400
2650
AD = adult; B = both; BDWT = body weight changes; d = days; DEYO = death of young; DR = drinking water; F = female; FD = food; PERT = fertility; GMPH = general morphology;
GRO = growth; GV = gavage; JV = juvenile; LC = lactation; M = male; MA = mature; mo = months; MPH = morphology; NR = not reported; ODVP = offspring development; OR = other oral;
PG = prostate gland; PROG = progeny counts or numbers; PRWT = progeny weight; RBPH = reproductive behavior; REP = reproduction; RHIS = reproductive organ histology; RSEM = resorbed
embryos; RSUC = reproductive success (general); RT = reproductive tissue; SM = sexually mature; SPCL = sperm cell counts; SPCV = sperm cell viability; TA = tail; TB = tibia; TE = testes;
TEDG = testes degeneration; TEWT = testes weight; w = weeks; WO = whole organism; yr = years.
Source: U.S. Environmental Protection Agency (2005b).
-------
10000
Legend:
• Growth NOAEL
o Growth LOAEL
A Reproduction NOAEL
A Reproduction LOAEL
- Eco-SSL (mg/kg dw)
0.1
Reproduction and Growth
Figure AX8-1.4.2. Mammalian reproduction and growth toxicity data considered in
development of the Eco-SSL.
Source: U.S. Environmental Protection Agency (2005b).
1 AX8.1.4.4 Recent Studies on the Effects of Lead on Decomposers
2 Recent studies on effects of Pb to two groups of decomposers are summarized in this
3 subsection. Effects on terrestrial invertebrates, such as earthworms and springtails, are described
4 first, followed by effects on microorganisms.
5
6 Effects on Invertebrates
1 Since the 1986 Pb AQCD, there have been several studies in which terrestrial
8 invertebrates were exposed to Pb in soil. Many of these were reviewed during the development
9 of the Eco-SSLs (U.S. Environmental Protection Agency, 2005b). The relevant information
10 from the Eco-SSL document is described below.
11 A literature search and review was conducted to identify critical papers published since
12 2002. Effects on earthworms and other invertebrates observed in studies conducted since the
May 2006 AX8-81 DRAFT-DO NOT QUOTE OR CITE
-------
1 1986 Pb AQCD are similar to those reported previously: mortality and decreased growth and
2 reproduction (Lock and Janssen, 2002; Davies et al., 2002; Rao et al., 2003; Bongers et al., 2004;
3 Nursita et al., 2005; U.S. Environmental Protection Agency, 2005b).
4 The literature search completed for terrestrial invertebrate Eco-SSL development
5 identified 179 papers for detailed review, of which 13 met the minimum criteria for further
6 consideration (U.S. Environmental Protection Agency, 2005b). Most of the 18 ecotoxicological
7 endpoints that were further evaluated measured reproduction or survival as the ecologically
8 relevant endpoint. Four of these, representing one species under three different pH test
9 conditions were used to develop the Eco-SSL of 1700 mg/kg soil (Table AX8-1.4.5).
10
11
Table AX8-1.4.5. Invertebrate Toxicity Data Used to Develop the Eco-SSL
Invertebrate Species
Collembola
(Folsomia Candida)
Collembola
Collembola
Collembola
Soil pH
6.0
4.5
5.0
6.0
% Organic
Matter
10
10
10
10
Toxicity Parameter
MATC1 (reproduction)
MATC (reproduction)
MATC (reproduction)
MATC (reproduction)
Geometric Mean
Pb in Soil
(mg/kg dw)
3162
3162
894
894
1682
* MATC = Maximum Acceptable Threshold Concentration, or the geometric mean of the NOEC (no-observed-
effect concentration) and LOEC (lowest-observed-effect concentration).
Source: U.S. Environmental Protection Agency (2005b).
12 In a study designed to test the toxicity of Pb to the earthworm Eiseniafetida, Davies
13 et al. (2002) found that the 28-day LC50 (± 95% confidence intervals) for Pb in soils
14 contaminated with Pb(NC>3)2 was 4379 + 356 mg/kg. Twenty-eight day ECso values (± 95%
15 confidence intervals) for weight change and cocoon production were 1408 + 198 and 971 + 633
16 mg/kg, respectively. Significant mortalities were noted at concentrations of 2000 mg/kg. These
May 2006 AX8-82 DRAFT-DO NOT QUOTE OR CITE
-------
1 data are consistent with those reported in the Eco-SSL document (U.S. Environmental Protection
2 Agency, 2005b) for the same species of earthworm.
3 Nursita et al. (2005) found no mortality and no adverse effects on reproduction (i.e.,
4 number of juveniles) of the collembolan Proisotoma minuta exposed for 42 days to 300, 750,
5 1500, or 3000 mg Pb/kg as Pb-nitrate in an acidic (pH = 4.88) sandy loam soil. It was noted that
6 the soils were allowed to equilibrate for 4 weeks after adding the Pb-nitrate before the organisms
7 were added. The observation of no effect at 3000 mg/kg is consistent with that of Sandifer and
8 Hopkin (1996). Sandifer and Hopkin (1996) determined a NOEC (no-observed-effect
9 concentration) and LOEC (lowest-observed-effect concentration) for collembolan reproduction
10 of 2000 and 5000 mg/kg, respectively. (A MATC [maximum-acceptable-threshold
11 concentration] of 3162 mg/kg was used to develop the Eco-SSL).
12 The remaining 14 toxicity endpoints that were not used to develop the Eco-SSL for
13 invertebrates are presented in Table AX8-1.4.6. None of these endpoints was considered eligible
14 for Eco-SSL derivation.
15 Lock and Janssen (2002) exposed the potworm Enchytraeus albidus to Pb, as Pb-nitrate.
16 The 21-day LCso was 4530 mg/kg, and the 42-day ECso for juvenile reproduction was
17 320 mg/kg. The Fl generation was then grown to maturity in the same concentration soil and
18 subsequently used in a reproduction test. The ECso for the Fl generation (394 mg/kg) was
19 similar to that of the P generation. The authors concluded that the two-generation assay did not
20 increase the sensitivity of the test (Lock and Janssen, 2002). None of the 18 toxicity endpoints
21 evaluated in detail during development of the Eco-SSLs used this species. The LCso reported for
22 the potworm was higher than reported for nematodes and similar to that reported for the
23 earthworm. The ECso for reproduction was lower than reported for the earthworm or collembola.
24 Recent work by Bongers et al. (2004) cautioned against attributing all toxicity observed in
25 a spiked-soil toxicity test to Pb. They found that the counterion may also contribute to the
26 toxicity of Pb in the springtail Folsomia Candida. This may have implications on the
27 interpretation of the Eco-SSL data, because the toxicity of the counterion (nitrate) was not taken
28 into account during Eco-SSL development. Percolation (removal of the counterion) had no
29 statistically significant effect on Pb-chloride toxicity (LCso = 2900 mg/kg for both non-
30 percolated and percolated soil; ECso for reproduction = 1900 mg/kg or 2400 mg/kg for non-
31 percolated or percolated soil, respectively). However, percolation did have a significant effect
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Table AX8-1.4.6. Invertebrate Toxicity Data Not Used to Develop the Eco-SSL
Invertebrate Species
Nematode
Nematode
Nematode
Nematode
Nematode
Nematode
Earthworm
Earthworm
Earthworm
Earthworm
Nematode
Nematode
Nematode
Nematode
Soil pH
4
4
4
4
6.2
5.1
6.3
6.1
6.0
6.5
4
4
6.1
6.2
% Organic
Matter
1.14
1.14
4.2
4.2
1.7
3.0
10.0
10.0
10.0
10.0
10
10
3.4
2.2
Toxicity
Parameter
LC50
LC50
LC50
LC50
LC50
LC50
EC50
EC50
LC50
ILL
LC50
NOAEC
LC50
LC50
Pb in Soil
(mg/kg dw)
285
297
847
1341
1554
891
1940
1629
3716
1.16
1434
2235
13.9
11.6
*NOAEC (no-observed-adverse-effect concentration); LC50 (concentration lethal to 50% of test population);
EC50 (effect concentration for 50% of test population); ILL (incipient lethal level).
Source: U.S. Environmental Protection Agency (2005b).
1 on Pb-nitrate toxicity (LCso = 980 mg/kg or 2200 mg/kg for non-percolated or percolated soil,
2 respectively; ECso for reproduction = 580 mg/kg or 1700 mg/kg for non-percolated or percolated
3 soil, respectively). Lead nitrate was more toxic than Pb-chloride for survival and reproduction.
4 However, the toxicity of Pb, from chloride or nitrate, was not significantly different after the
5 counterion was percolated out of the test soil. It is noted that the soil was left for 3 weeks to
6 equilibrate before testing. Lock and Janssen (2002) also found that Pb-nitrate was more toxic
7 than Pb-chloride, and they used Pb-nitrate in their experiments because 1000 mg/kg Pb-chloride
8 did not produce any mortality in their range-finding tests. This difference in chloride and nitrate
9 toxicity has not been found for earthworms (Neuhauser et al., 1985; Bongers et al., 2004).
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1 Rao et al. (2003) exposed the earthworm Eisenia fetida to Pb-oxide in an artificial soil
2 with a pH of 6 at the LCso concentration of 11 mg/kg. Exposure for 14 days resulted in a number
3 of effects including body fragmentation, protrusions, rupture of the cuticle, etc. Many of these
4 effects may trigger defensive mechanisms. For example, fragmentation of the affected posterior
5 region was followed by regeneration and a new ectoderm layer was formed to cover affected
6 areas, both of which processes may serve to prevent soil bacteria from further affecting the
7 earthworm (Rao et al., 2003 ).
8
9 Effects on Microorganisms and Microbial Processes
10 Microorganisms and microbial processes were not included in the Eco-SSL development
11 process (see Attachment 1-2 of OSWER Directive 92857-55 dated November 2003 in U.S.
12 Environmental Protection Agency [2005a]). Many reasons were given, including that it is
13 unlikely that site conditions would only pose unacceptable risk to microbes and not be reflected
14 as unacceptable risks to higher organisms; that the significance of laboratory-derived effects data
15 to the ecosystem is uncertain; and that the spatial (across millimeter distances) and temporal
16 (within minutes to hours) variation makes understanding ecological consequences challenging.
17 Microbial endpoints often vary dramatically based on moisture, temperature, oxygen, and many
18 non-contaminant factors. Therefore, the recommendation arising from the Eco-SSL
19 development process was that risks to microbes or microbial processes not be addressed through
20 the chemical screening process but that they should be addressed within a site-specific risk
21 assessment (U.S. Environmental Protection Agency, 2005b).
22 Few studies on the effects of Pb to microbial processes have been published since 1986.
23 As the direct toxicity to fungi and bacterial populations are difficult to determine and interpret,
24 indicators for soil communities are often measured as proxies for toxicity (e.g., urease activity in
25 soil). Recent studies of this nature (Doelman and Haanstra, 1986; Wilke, 1989; Haanstra and
26 Doelman, 1991) are summarized in this subsection. The Pb concentrations in these recent
27 studies (1000 to 5000 mg/kg) are consistent with those reported in the 1986 Pb AQCD (U.S.
28 Environmental Protection Agency, 1986a) as associated with effects on microbial processes (750
29 to 7500 mg/kg).
30 The effects of Pb-chloride on the processes of nitrification and nitrogen mineralization
31 were studied in a 28-day experiment by Wilke (1989). The authors reported that nitrification
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1 was increased by 12 and 16% at levels of 1000 and 4000 mg/kg, respectively, and that nitrogen
2 mineralization was reduced by 32 and 44% at concentrations of 1000 and 4000 mg/kg,
3 respectively.
4 The effects of Pb on arylsulfatase (Haanstra and Doelman, 1991) and urease activity
5 (Doelman and Haanstra, 1986) in soil were investigated. LCsos for decreases in arylsulfatase
6 activity were reported at Pb concentrations of 3004 and 4538 mg/kg in a silty loam soil, at pH 6
7 and 8, respectively. The LCso for a decrease in urease activity was 5060 mg Pb/kg in a sandy
8 loam soil.
9 In laboratory microcosm studies Cotrufo et al. (1995) found that decomposition of oak
10 (Quercus ilex) leaf litter was reduced at elevated Pb (-20 mg Pb g"1 C) levels after 8 months
11 compared to controls (~2 mg Pb g"1 C). The researchers found soil respiration and amount of
12 soil mycelia correlated negatively with soil Pb, Zn and Cr concentration.
13
14 AX8.1.4.5 Summary
15 The current document expands upon and updates knowledge related to the effects of Pb
16 on terrestrial primary producers, consumers, and decomposers.
17
18 Primary Producers
19 The effects of Pb on terrestrial plants include decreased photosynthetic and transpiration
20 rates in addition to decreased growth and yield. The phytotoxicity of Pb is considered to be
21 relatively low, and there are few reports of phytotoxicity from Pb exposure under field
22 conditions. Recently, phytotoxicity data were reviewed for the development of the Eco-SSL
23 (U.S. Environmental Protection Agency, 2005b). Many of the toxicity data presented in the Eco-
24 SSL document (U.S. Environmental Protection Agency, 2005b) are lower than those discussed in
25 the 1986 Pb AQCD, although both documents acknowledged that toxicity is observed over a
26 wide range of concentrations of Pb in soil (tens to thousands of mg/kg soil). This may be due to
27 many factors, such as soil conditions (e.g., pH, organic matter) and differences in bioavailability
28 of the Pb in spiked soils perhaps due to lack of equilibration of the Pb solution with the soil after
29 spiking. Most phytotoxicity data continue to be developed for agricultural plant species (i.e.,
30 vegetable and grain crops). Few data are available for trees or native herbaceous plants,
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1 although two of the five toxicity endpoints used to develop the Eco-SSL were for trees and two
2 were for clover.
3
4 Consumers
5 Effects of Pb on avian and mammalian consumers include decreased survival,
6 reproduction, and growth as well as effects on development and behavior. There remain few
7 field effects data for consumers, except from sites with multiple contaminants, for which it is
8 difficult to attribute toxicity specifically to Pb. Avian and mammalian toxicity data recently
9 were reviewed for the development of Eco-SSLs (U.S. Environmental Protection Agency,
10 2005b). Many of the toxicity data presented in the Eco-SSL document (U.S. Environmental
11 Protection Agency, 2005b) are lower than those discussed in the 1986 Pb AQCD, although the
12 EPA (U.S. Environmental Protection Agency, 2005b) recognizes that toxicity is observed over a
13 wide range of doses (<1 to >1000 mg Pb/kg bw-day). Most toxicity data for birds have been
14 derived from chicken and quail studies, and most data for mammals have been derived from
15 laboratory rat and mouse studies. Data derived for other species would contribute to the
16 understanding of Pb toxicity, particularly for wildlife species with different gut physiologies. In
17 addition, data derived using environmentally realistic exposures, such as from Pb-contaminated
18 soil and food, may be recommended. Finally, data derived from inhalation exposures, which
19 evaluate endpoints such as survival, growth, and reproduction, would contribute to understanding
20 the implications of airborne releases of Pb.
21
22 Decomposers
23 Effects of Pb on soil invertebrates include decreased survival, growth, and reproduction.
24 Effects on microorganisms include changes in nitrogen mineralization and enzyme activities.
25 Recent data on Pb toxicity to soil invertebrates and microorganisms are consistent with those
26 reported in the 1986 Pb AQCD (U.S. Environmental Protection Agency, 1986a), with toxicity
27 generally observed at concentrations of hundreds to thousands of mg/kg soil. Studies on
28 microbial processes may be influenced significantly by soil parameters, and the significance of
29 the test results is not clear.
30
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1 Ecological Soil Screening Levels (Eco-SSLs)
2 Eco-SSLs are concentrations of contaminants in soils that are protective of ecological
3 receptors (U.S. Environmental Protection Agency, 2005a). They were developed following
4 rigorous scientific protocols and were subjected to two rounds of peer review. The Eco-SSLs for
5 terrestrial plants, birds, mammals, and soil invertebrates are 120, 11, 56, and 1700 mg Pb/kg soil,
6 respectively.
7
8 AX8.1.5 Effects of Lead on Natural Terrestrial Ecosystems
9 The concept that organisms are part of larger systems that include both biotic and abiotic
10 components of the environment dates back to the naturalists of the Victorian era. However, the
11 breakthrough in what we now consider the ecosystem approach to ecology occurred in the 1950s
12 and 1960s when E.P. and H.T. Odum pioneered the quantitative analysis of ecosystems
13 (Odum, 1971). This approach encouraged the calculation of energy flows into, out of, and
14 within explicitly defined ecosystems. The rapid development of computer technology aided in
15 the growth of ecosystem ecology by allowing the development and use of increasingly complex
16 models for estimating fluxes that could not be directly measured.
17 It was not long before the quantitative analysis of ecosystems was extended to examine
18 the flows of nutrients and other chemical compounds. In temperate terrestrial systems, the
19 watershed was identified as a convenient and informative experimental unit (Bormann and
20 Likens, 1967). A major conceptual breakthrough in the watershed approach was that drainage
21 water chemistry could be used as an indicator of the "health" of the ecosystem. In a system
22 limited by nitrogen, for example, elevated concentrations of NOs in drainage waters indicate
23 that the ecosystem is no longer making optimal use of available nutrients.
24 The ecosystem approach can also be used effectively in the study of trace element
25 biogeochemistry. Input-output budgets can be used to determine whether an ecosystem is a net
26 source or sink of a trace element. Changes to the input-output balance over time can be used to
27 assess the effects of natural or experimental changes in deposition, land use, climate, or other
28 factors. In addition, examination of fluxes within the ecosystem (in plant uptake, soil solutions,
29 etc.) can be used to understand the processes that are most influential in determining the fate and
30 transport of the trace element.
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1 Many published ecosystem studies include data for 1 to 3 years, the typical duration of
2 research grant funding or doctoral dissertation research. While these studies enrich our
3 understanding of terrestrial ecosystems, the most valuable studies are those that are maintained
4 over many years. Natural variations in climate, pests, animal migrations, and other factors can
5 make inferences from short-term studies misleading (Likens, 1989). To nurture long-term
6 research, the National Science Foundation supports a network of Long-Term Ecological
7 Research (LTER) sites that represent various biomes.
8 This section describes terrestrial ecosystem research on Pb, focusing on work done since
9 the 1986 Pb AQCD (U.S. Environmental Protection Agency, 1986a) and highlighting key long-
10 term studies. Unfortunately, there are few studies that feature long-term data on trace metal
11 behavior at multiple levels of organization. Therefore, this examination of the effects of Pb on
12 terrestrial ecosystems combines insights from long- and short-term investigations as well as
13 studies at scales including whole ecosystems, communities, populations, and individual species.
14
15 AX8.1.5.1 Effects of Terrestrial Ecosystem Stresses on Lead Cycling
16 Terrestrial ecosystems may respond to stressors in a variety of ways, including reductions
17 in the vigor and/or growth of vegetation, reductions in biodiversity, and effects on microbial
18 processes. Each of these effects may lead to the "leakage" of nutrients, especially nitrogen, in
19 drainage waters. The reduced vigor or growth of vegetation results in a lower uptake of nitrogen
20 and other nutrients from soils. Reduced biodiversity accompanied by lower total net primary
21 productivity for the ecosystem would also result in a lower nutrient uptake. Effects of stress in
22 microbial populations are less obvious. If the stress reduces microbial activity rates, then
23 nutrients bound in soil organic matter (e.g., organic nitrogen compounds) will likely be
24 mineralized at a lower rate and retained in the system. On the other hand, disturbances such as
25 clear-cutting, ice-storm damage, and soil freezing can result in substantial nutrient losses from
26 soils (Bormann et al., 1968; Likens et al., 1969; Mitchell et al., 1996; Groffman et al., 2001;
27 Houlton et al., 2003).
28 Since the movement and fate of Pb in terrestrial ecosystems is strongly related to the
29 organic matter cycle (Section AX8.1.2), stressors that could lead to disruption or alteration of the
30 soil organic matter pool are of particular concern in assessing effects of ecosystem stress on Pb
31 cycling. By binding soluble Pb, soil organic matter acts as a barrier to the release of Pb to
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1 drainage waters (Wang et al., 1995; Kaste et al., 2003; Watmough and Hutchinson, 2004). As a
2 result, concentrations of Pb in soil solutions and drainage waters tend to be low (Driscoll et al.,
3 1988; Wang et al., 1995; Bacon and Bain, 1995; Johnson et al., 1995a). Through decomposition
4 and leaching, soluble organic matter is released to solution, and with it, some Pb is also
5 mobilized. Wang and Benoit (1996) found that essentially all of the Pb in soil solutions in a
6 hardwood forest in New Hampshire was bound to dissolved organic matter (DOM). This release
7 of soluble Pb does not typically result in elevated surface water Pb concentrations, because
8 (1) organic matter has a relatively long residence time in most temperate soils (Gosz et al., 1976;
9 Schlesinger, 1997), so only a small fraction of the organic matter pool is dissolved at any time;
10 (2) DOM-Pb complexes solubilized in upper soil horizons may be precipitated or adsorbed lower
11 in the soil profile; (3) the DOM to which Pb is bound may be utilized by microbes, allowing the
12 associated Pb to bind anew to soil organic matter. Together, these factors tend to moderate the
13 release of Pb to surface waters in temperate terrestrial ecosystems. However, stressors or
14 disturbances that result in increased release of DOM from soils could result in the unanticipated
15 release of Pb to groundwater and/or surface waters.
16
17 Acidification
18 The effect of acidification on ecosystem cycling of Pb is difficult to predict. Like most
19 metals, the solubility of Pb is increased at lower pH (Stumm and Morgan, 1995), suggesting that
20 enhanced mobility of Pb should be found in ecosystems under acidification stress. However,
21 reductions in pH may also decrease the solubility of DOM, via protonation of carboxylic
22 functional groups (Tipping and Woof, 1990). Because of the importance of complexation with
23 organic matter to Pb mobility in soils, lower DOM concentrations resulting from acidification
24 may offset the increased solubility of the metal.
25 In a study of grassland and forest soils at the Rothamsted Experiment Station in England,
26 long-term (i.e., >100 years) soil acidification significantly increased the mobility of Pb in the soil
27 (Blake and Goulding, 2002). However, the increased mobility was only observed in very acid
28 soils, those with pH of <4.5. The fraction of exchangeable Pb (extracted with 0.1 M CaCb)
29 increased from about 3% to 15% of the total Pb in the most acidified soils. Similarly, the
30 fraction of organically bound Pb increased from about 2% of total Pb in neutral soils to 12% of
31 total Pb in the most acidified soils. Similarly, Nouri and Reddy (1995) observed higher levels of
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1 diethylenetriaminepentaacetic-acid-[DTPA]-extractable Pb in soils in a loblolly pine forest
2 treated with simulated acid rain, but only in the most acidic treatment, with simulated rain with a
3 pHof3.5.
4 Although acidification may increase the mobility of Pb in soils, it is not clear that this Pb
5 is actually moving through or out of the soil profile. In an examination of running waters in
6 Sweden, Johansson et al. (1995) found no relationship between acidification and Pb
7 concentrations and concluded that Pb concentrations were governed by the DOM concentration,
8 which masked any association with acidification. In an in situ lysimeter study, Bergkvist (1986)
9 measured lower concentrations of Pb in soil solutions draining experimentally acidified plots
10 than in unacidified plots. In a laboratory study using large soil columns, Merino and Garcia-
11 Rodeja (1997) observed no effect of experimental acidification on the release of Pb to soil
12 solution. Thus, while acidification may increase the potential mobility of Pb in soils, as
13 indicated by increases in labile soil fractions such as exchangeable and DTPA-extractable Pb, the
14 actual movement of Pb in the soil is limited by DOM solubilization and transport. It is worth
15 noting that in all of these studies, significant effects of acidification were observed for other trace
16 metals (Bergkvist, 1986; Johansson et al., 1995; Merino and Garcia-Rodeja, 1997).
17 Acidification may enhance Pb export to drainage water in very sandy soils, soils with
18 limited ability to retain organic matter. Studies in the McDonald's Branch watershed in the
19 New Jersey pine barrens, where soil texture is similar to beach sands, suggested little Pb
20 retention in the mineral soil (Swanson and Johnson, 1980; Turner et al., 1985). If acidification
21 results in the mobilization of Pb and organic matter into these mineral soils, then increased
22 streamwater Pb concentrations would likely follow.
23
24 Forest Harvesting
25 Forest harvesting represents a severe disruption of the organic matter cycle in forest
26 ecosystems. Litter inputs are severely reduced for several years after cutting (e.g., Hughes and
27 Fahey, 1994). The removal of the forest canopy results in reduced interception of precipitation,
28 and, therefore, increased water flux to the soil surface. Also, until a new canopy closes, the soil
29 surface is exposed to increased solar radiation and higher temperatures. Together, the higher
30 moisture and temperature in surface soils tend to increase the rate of organic matter
31 decomposition. Several studies have estimated decreases of up to 40% in the organic matter
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1 content of forest floor soils after clear-cutting (Covington, 1981; Federer, 1984; Johnson et al.,
2 1995b). This loss of organic matter from the forest floor could result in the mobilization of
3 organically complexed Pb. However, observations from clear-cut sites in the United States and
4 Europe indicate that forest harvesting causes little or no mobilization of Pb from forest soils.
5 At the Hubbard Brook Experimental Forest in New Hampshire, whole-tree harvesting, the
6 most intensive form of clear-cutting, resulted in very small increases in Pb concentrations in soil
7 solutions draining the Oa soil horizon despite substantial reductions in the organic matter mass of
8 that horizon (Fuller et al., 1988; Johnson et al., 1995b). These increases were associated with
9 similarly small increases in dissolved organic carbon (DOC) concentrations in the Oa horizon
10 soil water. Output of Pb from the watershed stream was unaffected by clear-cutting. Similarly,
11 Berthelsen and Steinnes (1995) observed small decreases in the Pb content of the Oa horizon
12 ("H" in the European system of soil classification) in clear-cut sites in Norway, compared to
13 uncut reference sites. This mobilization of Pb from the Oa horizon was accompanied by an
14 increase in the Pb content of the upper mineral soil horizons. The Pb decline in the Oa horizon
15 was accompanied by a decrease in the organic matter content, leading the authors to attribute the
16 Pb dynamics to leaching with DOM. In a study conducted in Wales, Durand et al. (1994)
17 observed lower Pb outputs from a stream draining a clear-cut watershed than from where the
18 stream drained the upper reaches of the watershed, which were uncut. The DOC and H+ outputs
19 were also lower in the clear-cut area. These patterns persisted in all 5 years of the study.
20 Forest harvesting is a severe form of ecosystem disturbance, and, thus, it is somewhat
21 surprising that studies of clear-cutting have shown little or no effect on Pb mobility or loss from
22 forest ecosystems. Perhaps the strong complexation behavior of Pb with natural organic matter
23 results in the retention of Pb in forest soils. Even in cases where Pb is mobilized in forest floor
24 soils (Fuller et al., 1988; Berthelsen and Steinnes, 1995), there is no evidence of loss of Pb from
25 the ecosystem, indicating that mineral soils are efficient in capturing and retaining any Pb that is
26 mobilized in the forest floor. Therefore, the principal risk associated with forest harvesting is the
27 loss of Pb in particulate form to drainage waters through erosion.
28
29 Land Use and Industry
30 Changes in land use also represent potentially significant changes in the cycling of
31 organic matter in terrestrial ecosystems. Conversion of pasture and croplands to woodlands
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1 changes the nature and quantity of organic matter inputs to the soil. In temperate climates, forest
2 ecosystems tend to accumulate organic matter in an O horizon on the forest floor, whereas
3 organic matter in grasslands and agricultural fields is concentrated in an A horizon at the soil
4 surface. Andersen et al. (2002) compared the trace metal concentrations in arable fields in
5 Denmark to nearby sites that had been converted to forest land. After 34 years of afforestation,
6 the soils showed no significant difference in Pb concentration or fractionation, despite significant
7 acidification of the soils. Afforestation had no effect on the soil carbon concentration,
8 suggesting that land use change may have little effect on Pb cycling unless soil carbon pools are
9 affected.
10 Similarly, the introduction of industrial activity may have consequences for organic
11 matter cycling, and subsequently, Pb mobilization. In a rare long-term study of polluted soils,
12 Egli et al. (1999) studied the changes in trace metal concentrations in forest soils at a site in
13 western Switzerland between 1969 and 1993. The site is 3 to 6 km downwind from an aluminum
14 industrial plant that operated between the 1950s and 1991. In the 24-year period of study, the
15 site experienced significant declines in organic carbon in surface (0 to 5 cm depth) and
16 subsurface (30 to 35 cm) soils. In the 30 to 35 cm layer, the organic carbon concentration
17 declined by more than 75%. Extractable Pb (using an ammonium acetate and EDTA mixture)
18 declined by 35% in the same layer. The authors suggested that the Pb lost from the soil had been
19 organically bound. While this study indicates that loss of soil carbon can induce the mobilization
20 and loss of Pb from terrestrial ecosystems, it is also worth noting that the decline in soil Pb was
21 considerably smaller than the decline in organic carbon. This suggests that Pb mobilized during
22 organic matter decomposition can resorb to remaining organic matter or perhaps to alternate
23 binding sites (e.g., Fe and Mn oxides).
24 The effects of industries that emit Pb to the atmosphere are discussed in Sections
25 AX8.1.5.2 and AX8.1.5.3 below.
26
27 Climate Change
28 Atmospheric Pb is not likely to contribute significantly to global climate change. Lead
29 compounds have relatively short residence times in the atmosphere, making it unlikely that they
30 will reach the stratosphere. Also, Pb compounds are not known to absorb infrared radiation and,
31 therefore, are unlikely to contribute to stratospheric ozone depletion or global warming.
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1 Climate change does, however, represent a disturbance to terrestrial ecosystems.
2 Unfortunately, the potential linkages between climate-related stress and Pb cycling are poorly
3 understood. As in the previous examples, effects related to alterations in organic matter cycling
4 may influence Pb migration. For example, an increase in temperature leading to increased rates
5 of organic matter decomposition could lead to temporary increases in DOM concentrations and
6 smaller steady-state pools of soil organic matter. Either of these factors could result in increased
7 concentrations of Pb in waters draining terrestrial ecosystems.
8 Climate change may also affect the fluctuations of temperature and/or precipitation in
9 terrestrial ecosystems. For example, there is some evidence for recent increases in the frequency
10 of soil freezing events in the northeastern United States (Mitchell et al., 1996). Soil freezing
11 occurs when soils have little or no snow cover to insulate them from cold temperatures and
12 results in an increased release of nitrate and DOC from the O horizons of forest soils (Mitchell
13 et al., 1996; Fitzhugh et al., 2001). Increased DOC losses from O horizons subjected to freezing
14 may also increase Pb mobilization.
15 Increased fluctuations in precipitation may induce more frequent flooding, with
16 potentially significant consequences for Pb contamination of floodplain ecosystems. Soils
17 collected from the floodplain of the Elbe River, in Germany, contained elevated concentrations
18 of Pb and other trace metals (Kriiger and Grongroft, 2003). Tissues of plants from floodplain
19 sites did not, however, contain higher Pb concentrations than control sites. More frequent or
20 more severe flooding would likely result in increased inputs of Pb and other metals to floodplain
21 soils.
22
23 AX8.1.5.2 Effects of Lead Exposure on Natural Ecosystem Structure and Function
24 The effects of Pb exposure on natural ecosystems are confounded by the fact that Pb
25 exposure cannot be decoupled from other factors that may also affect the ecosystem under
26 consideration. Principal among these factors are other trace metals and acidic deposition.
27 Emissions of Pb from smelting and other industrial activities are accompanied by other trace
28 metals (e.g., Zn, Cu, and Cd) and sulfur dioxide (802) that may cause toxic effects independently
29 or in concert with Pb. Reductions in the use of alkyl-Pb additives in gasoline have resulted in
30 significant decreases in Pb deposition to natural ecosystems in the northeastern United States
31 (Johnson et al., 1995a). However, the period in which Pb deposition has declined (ca. 1975 to
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1 the present) has also seen significant reductions in the acidity (i.e., increased pH) of precipitation
2 in the region (Likens et al., 1996; Driscoll et al., 1998). Therefore, changes in ecosystem Pb
3 fluxes may be the result of reduced Pb inputs and/or reduced acidity.
4 Experimental manipulation studies do not suffer from these confounding effects, because
5 Pb can be added in specific amounts, with or without other compounds. Unfortunately,
6 ecosystem-level manipulations involving Pb additions have not been undertaken. Therefore, we
7 must use observations from field studies of Pb behavior in sites exposed to various forms of Pb
8 pollution to assess the effects of Pb on terrestrial ecosystems. This section includes a discussion
9 of effects of Pb in the structure and function of terrestrial ecosystems. Effects on energy flows
10 (food chain effects) and biogeochemical cycling are discussed in Section AX8.1.5.3.
11
12 Sites Affected by Nearby Point Sources of Lead
13 Natural terrestrial ecosystems near smelters, mines, and other industrial plants have
14 exhibited a variety of effects related to ecosystem structure and function. These effects include
15 decreases in species diversity, changes in floral and faunal community composition, and
16 decreasing vigor of terrestrial vegetation.
17 All of these effects were observed in ecosystems surrounding the Anaconda smelter in
18 southwestern Montana, which operated between 1884 and 1980 (Galbraith et al., 1995). Soils in
19 affected areas around the Anaconda smelter were enriched in Pb, arsenic, copper, cadmium, and
20 zinc; had very low pH; and were determined to be phytotoxic to native vegetation (Kapustka et
21 al., 1995). The elevated soil arsenic and metal concentrations occurred despite significantly
22 lower organic matter concentrations in affected soils relative to reference sites (Galbraith et al.,
23 1995). Line-transect measurements indicated that affected sites had an average of 6.9 species per
24 10-m of transect, compared to 20.3 species per 10-m in the reference areas. More than 60% of
25 the reference sites supported coniferous (58%) or deciduous (3%) forest communities, whereas
26 less than 1% of the affected sites retained functioning forest stands. Abundant dead timber and
27 stumps confirmed that the affected sites were once as forested as the reference sites. Affected
28 grassland sites were also less diverse and had higher abundances of invasive species than
29 reference grasslands. More than 50% of the affected sites were classified as bare ground. The
30 occurrence of bare ground was significantly correlated with the phytotoxicity scores derived by
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1 Kapustka et al. (1995), indicating a link between phytotoxicity and the loss of vegetation in the
2 affected area.
3 Because of the plant community changes near the Anaconda smelter, the vertical diversity
4 of habitats in the affected ecosystems decreased, with only shrubs and soil remaining as viable
5 habitats. Galbraith et al. (1995) also used the Bureau of Land Management's habitat evaluation
6 procedure (HEP) to estimate habitat suitability indices (HSI) for two indicator species, marten
7 (Martes americand) and elk (Cervus elaphus). The HSI value ranges from 0 (poor habitat) to 1
8 (ideal habitat). In sites affected by the Anaconda smelter, HSI values for marten averaged 0.0,
9 compared to 0.5 to 0.8 for the reference sites. For elk, affected sites had an average HSI of 0.10,
10 compared to 0.31 at reference sites.
11 Similar observations were made in the area surrounding Palmerton, Pennsylvania, where
12 two zinc smelters operated between 1898 and 1980. Soils in the area were enriched in Cd, Zn,
13 Pb, and Cu, with concentrations decreasing with distance from the smelter sites (Beyer et al.,
14 1985; Storm et al., 1994). Smelting was determined to be the principal source of Pb in soils in
15 residential and undeveloped areas around Palmerton (Ketterer et al., 2001), which lies on the
16 north side of a gap in Blue Mountain, a ridge running roughly east-west in east-central
17 Pennsylvania. Much of the north-facing side of Blue Mountain within 3 km of the town is bare
18 ground or sparsely vegetated, whereas the surrounding natural landscape is predominantly oak
19 forest (Sopper, 1989; Storm et al., 1994). Biodiversity in affected areas is considerably lower
20 than at reference sites, a pattern attributed to emissions from the smelters (Beyer et al., 1985;
21 Sopper, 1989). The history is complicated, however, by the land use history of the area.
22 Logging and fire in the early 20th century may also have played a role in the changes in the
23 terrestrial ecosystems (Jordan, 1975). Extensive logging occurred after the smelters began
24 operation, suggesting that some of the logging may have been salvage logging in affected areas.
25 Regardless, the smelter emissions appear to have inhibited the regrowth of ecosystems compared
26 to those in nearby unaffected areas. As in Anaconda, MT, the changes in the structure and
27 function of the Palmerton ecosystem changed its suitability as a habitat for fauna that would
28 normally inhabit the area. Storm et al. (1994) did not find amphibians or common invertebrates
29 in two study sites nearest to the smelters. In the larger study area, they documented elevated
30 concentrations of Pb, Cd, Cu, and Zn in tissues of species ranging in size from red-backed
31 salamanders (Pletheron cenereus) to white-tailed deer (Odocoilius virginianus).
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1 Metal pollution around a Pb-Zn smelter near Bristol, England has not resulted in the loss
2 of oak woodlands within 3 km of the smelter, despite significant accumulation of Pb, Cd, Cu,
3 and Zn in soils and vegetation (Martin and Bullock, 1994). However, the high metal
4 concentrations have favored the growth of metal-tolerant species in the woodland.
5 The effects of Pb on terrestrial ecosystems near smelters and other industrial sites
6 decrease downwind from the Pb source. Several studies using the soil Pb burden as an indicator
7 have shown that much of the contamination occurs within a radius of 20 to 50 km around the
8 emission source (Miller and McFee, 1983; Martin and Bullock, 1994; Galbraith et al., 1995;
9 Spurgeon and Hopkin, 1996a; see also Section AX8.1.2.). For example, the concentration of Pb
10 in forest litter declined downwind from a Pb-Zn smelter near Bristol, UK, from 2330 to 3050
11 ppm in a stand 2.9 km from the smelter to 45 to 110 ppm in a stand 23 km from the smelter
12 (Martin and Bullock, 1994). Thus, while sites near point sources of Pb may experience profound
13 effects on ecosystem structure and function, the extent of those effects is limited spatially. Most
14 terrestrial ecosystems are far enough from point sources that long-range Pb transport is the
15 primary mechanism for Pb inputs.
16
17 Sites Affected by Long-Range Lead Transport
18 Because the effects of anthropogenic Pb emissions tend to be restricted in geographic
19 extent, most natural terrestrial ecosystems in the U.S. sites have Pb burdens derived primarily
20 from long-range atmospheric transport. Pollutant Pb represents a large fraction of the Pb in
21 many of these ecosystems. In particular, many of these sites have accumulated large amounts of
22 Pb in soils. For example, at the Hubbard Brook Experimental Forest in New Hampshire, the
23 amount of Pb in the forest floor was estimated to have increased from about 1.35 kg ha l in 1926
24 (before the introduction of alkyl-Pb additives in gasoline) to 10.5 kg ha l in 1977 (Johnson et al.,
25 1995a). They also estimated the atmospheric Pb deposition from 1926 to 1987 to be 8.7 kg ha'1,
26 an amount that could account for nearly all of the increase in Pb in the forest floor during the
27 period. The input of precipitation Pb to the Hubbard Brook ecosystem in the six decades
28 spanning 1926 to 1987 was more than half of the total Pb estimated to have been released by
29 mineral weathering in the entire 12,000- to 14,000-year post-glacial period (14.1 kg ha'1:
30 [Johnson et al., 2004]). Other studies employing Pb budgets (Miller and Friedland, 1994;
31 Watmough et al., 2004), and Pb isotopes (Bacon et al., 1995, 1996; Watmough et al., 1998;
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1 Bindler et al., 1999; Hansmann and Koppel, 2000; Kaste et al., 2003), have also shown that
2 pollutant Pb, primarily from gasoline combustion, represents a quantitatively significant fraction
3 of labile Pb in temperate soils, especially in the upper, organic-rich horizons.
4 Despite years of elevated atmospheric Pb inputs and elevated concentrations in soils, there
5 is little evidence that sites affected primarily by long-range Pb transport have experienced
6 significant effects on ecosystem structure or function. Low concentrations of Pb in soil
7 solutions, the result of strong complexation of Pb by soil organic matter, may explain why few
8 ecological effects have been observed. At Hubbard Brook, for example, the concentration of Pb
9 in soil solutions draining the Oa horizon is <0.1 |iM and is even lower in solutions draining
10 mineral-soil horizons (Driscoll et al., 1988; Wang et al., 1995). Friedland and Johnson (1985)
11 measured similar concentrations in soil solutions collected from deciduous and spruce-fir stands
12 on Camel's Hump Mountain in Vermont. In an undeveloped, forested watershed in Maryland,
13 Scudlark et al. (2005) found that atmospheric input of some elements (Al, Cd, Ni, Zn) is
14 effectively transmitted through the watershed, whereas other elements (Pb, As, Se, Fe, Cr, Cu)
15 are strongly sequestered, in the respective order noted.
16 In ecosystems where Pb concentrations in soil solutions are low, toxicity levels for
17 vegetation are not likely to be reached regardless of the soil Pb concentration. Furthermore,
18 mycorrhizal infection of tree roots appears to reduce the translocation of Pb from roots to shoots
19 (Marschner et al., 1996; Jentschke et al., 1998). In a study of mycorrhizal and non-mycorrhizal
20 Norway spruce (Picea abies (L.) Karst), mycorrhizal infection of roots was not affected by Pb
21 dose. Some, but not all, species of mycorrhizae showed reductions in the amount of
22 extrametrical mycelium with Pb exposure but only at solution concentrations of 5 jiM, a level at
23 least 50 times greater than typical concentrations in forest soils. In a related study, the growth
24 rate of mycorrhizal fungi was unaffected at solution Pb concentrations of 1 and 10 jiM, but
25 decreased at 500 |iM (Marschner et al., 1999).
26 Low soil solution Pb concentrations and the influence of mycorrhizal symbionts also
27 result in low uptake of Pb by terrestrial vegetation. The net flux of Pb into vegetation in the
28 northern hardwood forest at Hubbard Brook in the 1980s was estimated as only 1 g ha'1 year"1
29 (Johnson et al., 1995a), representing 3% of the precipitation input. Klaminder et al. (2005) also
30 measured a Pb uptake of 1 g ha'1 year"1 in a spruce-pine forest in northern Sweden. Despite
31 plant uptake fluxes being very low, they are sensitive to differences and changes in Pb
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1 deposition. Berthelsen et al. (1995) observed decreases in the Pb content of stem, twig, leaf, and
2 needle tissues of a variety of tree species in Norway between 1982 and 1992, when atmospheric
3 Pb deposition declined by approximately 70%. They also observed significantly lower Pb
4 concentrations in tree tissues collected in northern Norway versus southern Norway, where
5 atmospheric Pb deposition is greater.
6 Even at subtoxic concentrations, Pb and other metals may influence species diversity in
7 terrestrial ecosystems. However, little work has been done on the effect of low-level metal
8 concentrations on species diversity. In one study, plant species diversity was positively
9 correlated to the concentration of available Pb in natural and artificial urban meadows in Britain
10 (McCrea et al., 2004). The authors hypothesized that Pb may inhibit phosphorous uptake by
11 dominant species, allowing less abundant (but more Pb-tolerant) ones to succeed.
12
13 AX8.1.5.3 Effects of Lead on Energy Flows and Biogeochemical Cycling
14 In terrestrial ecosystems, energy flow is closely linked to the carbon cycle. The principal
15 input of energy to terrestrial ecosystems is through photosynthesis, in which CC>2 is converted to
16 biomass carbon. Because of this link between photosynthesis and energy flow, any effect that Pb
17 has on the structure and function of terrestrial ecosystems (as discussed in Section AX8.1.5.3.)
18 influences the flow of energy into the ecosystem. This section focuses on how Pb influences
19 energy transfer within terrestrial ecosystems, which begin with the decomposition of litter and
20 other detrital material by soil bacteria and fungi, and cascade through the various components of
21 the detrital food web. Because the mobility of Pb in soils is closely tied to organic matter
22 cycling, decomposition processes are central to the biogeochemical cycle of Pb. This section
23 concludes with a discussion of how biogeochemical cycling of Pb has changed in response to the
24 changing Pb inputs to terrestrial ecosystems.
25
26 Effects of Lead on Detrital Energy Flows
27 Lead can have a significant effect on energy flows in terrestrial ecosystems. At some sites
28 severely affected by metal pollution, death of vegetation can occur, dramatically reducing the
29 input of carbon to the ecosystem (Jordan, 1975; Galbraith et al., 1995). Subsequently, wind and
30 erosion may remove litter and humus, leaving bare mineral soil, a nearly sterile environment in
31 which very little energy transfer can take place (Little and Martin, 1972; Galbraith et al., 1995).
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1 At Pb-affected sites that can retain a functioning forest stand, the rate of decomposition of
2 litter may be reduced, resulting in greater accumulation of litter on the forest floor than in
3 unpolluted stands. Numerous investigators have documented significant declines in litter
4 decomposition rates (Cotrufo et al., 1995; Johnson and Hale, 2004) and/or the rate of carbon
5 respiration (Laskowski et al., 1994; Cotrufo et al., 1995; Saviozzi et al., 1997; Niklinska et al.,
6 1998; Palmborg et al., 1998; Aka and Darici, 2004) in acid- and metal-contaminated soils or soils
7 treated with Pb. The resulting accumulation of organic matter on the soil surface can be
8 dramatic. For example, an oak woodland 3 km from a smelter in Bristol, England had a litter
9 layer mass 10 times greater than the mass in a similar stand 23 km from the smelter (Martin and
10 Bullock, 1994).
11 Lower decomposition rates in polluted ecosystems are the result of the inhibition of soil
12 bacteria and fungi and its effects on microbial community structure (Baath, 1989). Kuperman
13 and Carreiro (1997) observed 60% lower substrate-induced respiration in heavily polluted
14 grassland soils near the U.S. Army's Aberdeen Proving Ground in Maryland. This decline in
15 carbon respiration was associated with 81% lower bacterial biomass and 93% lower fungal
16 biomass. Similar declines in the activities of carbon-, nitrogen-, and phosphorus-acquiring
17 enzymes were also observed. Such dramatic effects have only been observed in highly
18 contaminated ecosystems. In a less contaminated grassland site near a Pb factory in Germany,
19 Chander et al. (2001) observed a lower ratio of microbial biomass carbon to soil organic carbon
20 in polluted soils. The ratio of basal respiration to microbial biomass (the "metabolic quotient,"
21 qCC>2) declined with increasing metal concentration, though this observation depended on the
22 procedure for measuring microbial biomass (substrate-induced respiration versus fumigation-
23 extraction). The combined effect of lower microbial biomass per unit soil carbon and similar or
24 lower qCC>2 on polluted sites indicates that the ability of soil microorganisms to process carbon
25 inputs is compromised by metal pollution.
26 The type of ecosystem also plays a role in determining the effects of Pb and other metals
27 on the microbial processing of litter. Forest soils in temperate zones accumulate organic matter
28 at the soil surface to a greater degree than in grasslands. This organic-rich O horizon can support
29 a large microbial biomass; but it is also an effective trap for Pb inputs, because of the association
30 between Pb and soil organic matter. At highly contaminated forest sites, microbial biomass and
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1 enzyme activities may be depressed (Fritze et al., 1989; Baath et al., 1991), causing slower
2 decomposition of the litter.
3 In addition to effects on decomposition and carbon transformations, Pb and other trace
4 metals can also influence key nitrogen cycling processes. Studies in the 1970s demonstrated that
5 Pb and other metals inhibit the mineralization of nitrogen from soil organic matter and
6 nitrification (Liang and Tabatabai, 1977, 1978), resulting in lower nitrogen availability to plants.
7 More recent research has documented significant inhibitory effects of Pb and other metals on the
8 activities of several enzymes believed to be crucial to nitrogen mineralization in soils (Senwo
9 and Tabatabai, 1999; Acosta-Martinez and Tabatabai, 2000; Ekenler and Tabatabai, 2002). This
10 suggests that the inhibitory effect of Pb and other metals is broad-based, and not specific to any
11 particular metabolic pathway. In reducing environments, the rate of denitrification is also
12 depressed by trace metals. Fu and Tabatabai (1989) found that 2.5 |imol g'1 of Pb (ca.
13 500 mg/kg"1) was sufficient to cause 0, 27, and 52% decreases in nitrogen reductase activity in
14 three different soils.
15 Metal pollution can also affect soil invertebrate populations. Martin and Bullock (1994)
16 observed lower abundances of a variety of woodlice, millipedes, spiders, insects, and earthworms
17 in an oak woodland site 3 km from a Pb-Zn smelter in Bristol, England, compared to a reference
18 site 23 km from the smelter. The differences were most dramatic when expressed per unit mass
19 of litter. Several species that were abundant in the reference site were not found in the
20 contaminated woodland. For example, the abundance of the woodlice Trichoniscus pusillus
21 was 151 individuals per m2 in the reference woodland, but none were found in the contaminated
22 soils. This was also true of 2 of the 3 millipede species, and 4 of the 5 earthworm species
23 studied. At six sites within 1 km from the smelters, no earthworms were present at all (Spurgeon
24 and Hopkin, 1996a). Contamination at this site has apparently reduced both the population and
25 biodiversity of the soil invertebrate community.
26 The effect of metal pollution on soil invertebrates may be a threshold-type response. In a
27 study conducted in woodlands near two zinc smelters in Noyelles-Godault, in northern France,
28 soils at the most polluted site were devoid of mites and millipedes, while the remaining sites had
29 diversity measures similar to control sites (Grelle et al., 2000).
30 While Pb pollution affects the population and diversity of soil fauna, there is little
31 evidence of significant bioaccumulation ofPb in the soil food web (see also Section AX8.1.3.).
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1 In the Bristol, England study, Pb concentrations in earthworms were lower than soil Pb
2 concentrations and much lower than litter Pb concentrations (Martin and Bullock, 1994). Litter-
3 dwelling mites had Pb concentrations that were 10% of the average litter concentration. The
4 predator centipedes Lithobius forficatus and L. variegatus had mean Pb concentrations of
5 18.6 and 44.0 mg kg"1, respectively, two orders of magnitude lower than the Pb concentration of
6 litter (2193 mg kg"1) and lower than the concentrations of their known prey species. In a study
7 conducted in a Norway spruce forest affected primarily by automobile exhaust from a nearby
8 highway, earthworms had Pb concentrations similar to the soil (Roth, 1993). Almost all of the
9 litter decomposers, however, had Pb concentrations that were less than 20% of the litter. All but
10 3 of the zoophagous arthropods had Pb concentrations that were less than 40% of their prey; the
11 remaining 3 had Pb concentrations similar to their prey. Because of the absence of significant
12 bioaccumulation in the soil food web, predator species will be affected by Pb pollution primarily
13 through effects on the abundance of their prey (Spurgeon and Hopkin, 1996b).
14 Taken as a whole, ecosystem-level studies of the soil food web indicate that Pb can affect
15 energy flows in terrestrial ecosystems through two principal mechanisms. In the most severely
16 polluted sites, the death of primary producers directly decreases the flow of energy into the
17 ecosystems. More commonly, the accumulation of toxic levels of Pb or other metals in litter and
18 soil decreases the rate of litter decomposition through decreases in microbial biomass and/or
19 respiration. These reductions can subsequently affect higher trophic levels that depend on these
20 organisms. It is important to note that sites that have exhibited significant disruption to energy
21 flows and the terrestrial food web are sites that have experienced severe metal contamination and
22 adverse effects from SC>2 from smelters or other metals-related activities.
23
24 Lead Dynamics in Terrestrial Ecosystems
25 Lead inputs to terrestrial ecosystems in the United States have declined dramatically in the
26 past 30 years, primarily because of the almost complete elimination of alkyl-Pb additives in
27 gasoline in North America. Also, Pb emissions from smelters have declined as older plants have
28 been shut down or fitted with improved emissions controls. Unfortunately, there are few long-
29 term data sets of precipitation inputs to terrestrial ecosystems. At the Hubbard Brook
30 Experimental Forest, in New Hampshire, Pb input in bulk deposition declined by more than 97%
31 between 1976 and 1989 (Johnson et al., 1995a). Studies of freshwater sediments also indicate a
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1 dramatic decline in Pb inputs since the mid-1970s (Graney et al., 1995; Johnson et al., 1995a;
2 Farmer et al., 1997; Brannvall et al., 2001a,b).
3 Reported concentrations of Pb in waters draining natural terrestrial ecosystems have
4 always been low (Wang et al., 1995; Bacon and Bain, 1995; Johnson et al., 1995a; Vinogradoff
5 et al., 2005), generally less than 1 ng I/1, even at moderately polluted sites (Laskowski et al.,
6 1995). Consequently, most terrestrial ecosystems in North America and Europe remain sinks for
7 Pb despite reductions in atmospheric Pb deposition of more than 95%. At Hubbard Brook, for
8 example, the input of Pb in bulk precipitation declined from 325 g ha"1 year"1 between 1975 and
9 1977 compared to 29 g ha"1 year"1 between 1985 and 1987 (Johnson et al., 1995a). During the
10 same period, the output of Pb in stream water declined from 6 g ha'1 year"1 to 4 g ha l year"1.
11 Thus, despite the decline in Pb input, 85% of the incoming Pb was still retained in the terrestrial
12 ecosystem in the later time period. Similar observations have been made in Europe, where the
13 use of leaded gasoline has also declined in the last few decades. At the Glensaugh Research
14 Station in Scotland, the input of Pb to the forest ecosystem was estimated as 42.6 g ha"1 year"1
15 between 2001 and 2003, about six times the stream export of 7.2 g ha"1 year"1 (Vinogradoff et
16 al., 2005). Similarly, Huang and Matzner (2004) reported a throughfall flux of 16.5 g ha"1 year"1
17 at the forested Lehstenbach catchment in Bavaria, about six times the efflux in runoff of 2.82 g
18 ha"1 year"1.
19 Lead pollution has resulted in the accumulation of large Pb burdens in terrestrial
20 ecosystems (see also Section AX8.1.2). Despite reductions in emissions, this accumulation of Pb
21 continues, though at markedly lower rates. The large pool of Pb bound in soils may potentially
22 be a threat to aquatic ecosystems (see Section AX8.2), depending on its rate of release from the
23 soil. Early estimates of the residence time of Pb in the forest floor ranged from 220 to 5,000
24 years (Benninger et al., 1975; Friedland and Johnson, 1985; Turner et al., 1985). However, more
25 recent literature suggests that Pb is transported more rapidly within soil profiles than previously
26 believed. The pool of Pb in forest floor soils of the northeastern United States declined
27 significantly in the late 20th century. Friedland et al. (1992) reported a 12% decline in the
28 amount of Pb in forest floor soils at 30 sites in the region between 1980 and 1990, a much greater
29 decline than would be expected for a pool with a residence time of 220 to 5,000 years.
30 At Hubbard Brook, the pool of Pb in the forest floor declined by 29% between 1977 and 1987,
31 an even more rapid rate of loss than reported by Friedland et al. (1992). More recently, Evans
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1 et al. (2005) reported significant declines in the Pb content of forest floor soils in the
2 northeastern United States and eastern Canada between 1979 and 1996. The magnitude of the
3 decrease in Pb content was greatest at their sites in southern Vermont, and smallest at sites on the
4 Gaspe Peninsula in Quebec, reflecting the historic gradient in Pb deposition in the region. At the
5 Vermont site, the Pb concentration in the litter layer (Oi horizon) was 85% lower in 1996 than in
6 1979. In the Gaspe peninsula of Quebec, the decrease was only 50%.
7 Since drainage water Pb concentrations remain low, the Pb released from forest floor soils
8 in the past has been largely immobilized in mineral soils (Miller and Friedland, 1994; Johnson
9 et al., 1995a; Johnson and Petras, 1998; Watmough and Hutchinson, 2004; Johnson et al., 2004).
10 This is supported by evidence from Pb-isotope analyses. Gasoline-derived Pb has a 206Pb:207Pb
11 ratio that can be easily discriminated from Pb in the rocks from which soils are derived. Using
12 isotopic mixing models with gasoline-Pb and Pb in soil parent materials as end members,
13 a number of researchers have documented the accumulation of pollutant Pb in mineral soils
14 (Bindler et al., 1999; Kaste et al., 2003; Watmough and Hutchinson, 2004; Bacon and Hewitt,
15 2005; Steinnes and Friedland, 2005). In a hardwood stand on Camel's Hump Mountain in
16 Vermont, as much as 65% of the pollutant Pb deposited to the stand had moved into mineral
17 horizons by 2001 (Kaste et al., 2003). In a spruce-fir stand, containing a thicker organic forest
18 floor layer, penetration of pollutant Pb into the mineral soil was much lower.
19 This recent research has resulted in a reevaluation of the turnover time of Pb in forest
20 floor soils. The Camel's Hump data suggest that Pb resides in the forest floor of deciduous
21 stands for about 60 years and about 150 years in coniferous stands (Kaste et al., 2003). These
22 values are somewhat greater than those published previously by Miller and Friedland (1994),
23 who used a Pb budget approach. Extremely rapid turnover of Pb was observed in some
24 hardwood forest floor soils in south-central Ontario (Watmough et al., 2004). Their estimated
25 turnover times of 1.8 to 3.1 years are much lower than any other published values, which they
26 attribute to the mull-type forest floor at their sites. Mull-type forest floors are normally underlain
27 by organic-rich A horizons, capable of immobilizing Pb released from the forest floor. Indeed,
28 at the same site in Ontario, Watmough and Hutchinson (2004) found that 90% of the pollutant Pb
29 could be found in this A horizon.
30 The time period over which the accumulated Pb in soils may be released to drainage
31 waters remains unclear. If Pb moves as a pulse through the soil, there may be a point in the
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1 future at which problematic Pb concentrations occur. However, several authors have argued
2 against this hypothesis (Wang and Benoit, 1997; Kaste et al., 2003; Watmough et al., 2004),
3 contending that the strong linkage between Pb and DOM will result in a temporally dispersed
4 release of Pb in the form of Pb-DOM complexes. Thus, the greatest threat is likely to be in the
5 most highly contaminated areas surrounding point sources of Pb, where the amount of Pb
6 accumulated in the soil is high, and the death of vegetation has resulted in reduced soil organic
7 matter levels.
8
9 AX8.1.5.4 Summary
10 Atmospheric Pb pollution has resulted in the accumulation of Pb in terrestrial ecosystems
11 throughout the world. In the United States, pollutant Pb represents a significant fraction of the
12 total Pb burden in soils, even in sites remote from smelters and other industrial plants. However,
13 few significant effects of Pb pollution have been observed at sites that are not near point sources
14 of Pb. Evidence from precipitation collection and sediment analyses indicates that atmospheric
15 deposition of Pb has declined dramatically (>95%) at sites unaffected by point sources of Pb, and
16 there is little evidence that Pb accumulated in soils at these sites represents a threat to
17 groundwaters or surface water supplies.
18 The highest environmental risk for Pb in terrestrial ecosystems exists at sites within about
19 50 km of smelters and other Pb-emitting industrial sites. Assessing the risks specifically
20 associated with Pb is difficult, because these sites also experience elevated concentrations of
21 other metals and because of effects related to SC>2 emissions. The concentrations of Pb in soils,
22 vegetation, and fauna at these sites can be two to three orders of magnitude higher than in
23 reference areas (see Sections AX8.1.2. and AX8.1.5.2.). In the most extreme cases, near smelter
24 sites, the death of vegetation causes a near-complete collapse of the detrital food web, creating a
25 terrestrial ecosystem in which energy and nutrient flows are minimal. More commonly, stress in
26 soil microorganisms and detritivores can cause reductions in the rate of decomposition of detrital
27 organic matter. Although there is little evidence of significant bioaccumulation of Pb in natural
28 terrestrial ecosystems, reductions in microbial and detritivorous populations can affect the
29 success of their predators. Thus, at present, industrial point sources represent the greatest Pb-
30 related threat to the maintenance of sustainable, healthy, diverse, and high-functioning terrestrial
31 ecosystems in the United States.
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1 AX8.2 AQUATIC ECOSYSTEMS
2 AX8.2.1 Methodologies Used in Aquatic Ecosystem Research
3 As discussed in previous sections, aerial deposition is one source of Pb deposition to
4 aquatic systems. Consequently, to develop air quality criteria for Pb, consideration must be
5 given to not only the environmental fate of Pb, but also to the environmental effects of Pb in the
6 aquatic environment through consideration of laboratory toxicity studies and field evaluations.
7 Perhaps the most straightforward approach for evaluating the effects of Pb is to consider extant
8 criteria for Pb in aquatic ecosystems, i.e., water and sediment quality criteria. A key issue in
9 developing Pb water and sediment criteria that are broadly applicable to a range of water bodies
10 is properly accounting for Pb bioavailability and the range in species sensitivities. This section
11 summarizes how these criteria are derived, the types of toxicity studies considered, and key
12 factors that influence the bioavailability of Pb in surface water and sediment to aquatic life.
13 Because Pb in the aquatic environment is often associated with other metals (e.g., cadmium,
14 copper, zinc), the importance of considering the toxicity of metal mixtures is also discussed.
15 Finally, some issues related to background Pb concentrations are briefly addressed. It is beyond
16 the scope of this section to review all methodologies in aquatic system research, but good
17 reviews can be found in summary books, such as Rand et al. (1995).
18
19 AX8.2.1.1 Analytical Methods
20 Common analytical methods for measuring Pb in the aquatic environment are summarized
21 in Table AX8-2.1.1. For relevance to the ambient water quality criteria (AWQC) and sediment
22 quality criteria for Pb discussed below, minimum detection limits should be in the low parts per
23 billion (ppb) range for surface water and the low parts per million (ppm) range for sediment.
24 In addition to the methods presented in Table AX8-2.1.1, many of the methods discussed
25 in Section AX8.1.1 can be applied to suspended solids and sediments collected from aquatic
26 ecosystems. Just as in the terrestrial environment, the speciation of Pb and other trace metals in
27 natural freshwaters and seawater plays a crucial role in determining their reactivity, mobility,
28 bioavailability, and toxicity. Many of the same speciation techniques employed for the
29 speciation of Pb in terrestrial ecosystems (see Section AX8.1.1 and AX8.1.2) are applicable in
30 aquatic ecosystems.
31
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Table AX8-2.1.1. Common Analytical Methods for Measuring Lead in Water,
Sediment, and Tissue
Analysis Type Analytical Method
Direct-Aspiration (Flame) Atomic Absorption EPA SW-846 Method 7420a,
Spectroscopy (AAS) EPA Method 239. lb,
Standard Method 31 llc
Graphite Furnace Atomic Absorption Spectroscopy EPA SW-846 Method 7421a,
(GFAAS) EPA Method 239.2b,
Standard Method 3113C
Inductively Coupled Plasma EPA SW-846 Method 601 OBa,
(ICP) EPA Method 200.7b,
Standard Method 3120C
Inductively Coupled Plasma-Mass Spectrometry EPA SW-846 Method 6020a,
(ICP-MS) EPA Method 200.8b
a U.S. Environmental Protection Agency (1986c) Test Methods for Evaluating Solid Waste, Physical/Chemical
Methods (SW-846). Third Edition, September 1986; Final Updates I (7/92), IIA (8/93), II (9/94), IIB (1/95),
III (12/96), IIIA (4/98), IIIB (11/04).
b U.S. Environmental Protection Agency (1991) Methods for the Determination of Metals in Environmental
Samples. EPA/600/4-91-010. June 1991 (Supplement I, EPA/600/R-94-111, May 1994).
0 American Public Health Association (1995) Standard Methods for the Examination of Water and Wastewater,
19th Edition. American Public Health Association, American Water Works Association, Water Pollution
Control Federation.
1 There is now a better understanding of the potential effects of sampling, sample handling,
2 and sample preparation on aqueous-phase metal speciation. Thus, a need has arisen for dynamic
3 analytical techniques that are able to capture a metal's speciation, in-situ and in real time. Some
4 of these recently developed dynamic trace metal speciation techniques include:
5 • Diffusion gradients in thin-film gels (DGT)
6 • Gel integrated microelectrodes combined with voltammetric in situ profiling (GIME-
7 VIP)
8 • Stripping chronopotentiometry (SCP)
9 • Flow-through and hollow fiber permeation liquid membranes (FTPLM and FIFPLM)
10 • Donnan membrane technique (DMT)
11 • Competitive ligand-exchange/stripping voltammetry (CLE-SV)
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1 Various dynamic speciation techniques were compared in a study by Sigg et al. (2006)
2 using freshwaters collected in Switzerland. They found that techniques involving in-situ
3 measurement (GEVIE-VIP) or in-situ exposure (DOT, DMT, and HFPLM) appeared to the most
4 appropriate for avoiding Pb and other trace metal speciation artifacts associated sampling and
5 sample handling.
6
7 AX8.2.1.2 Ambient Water Quality Criteria: Development
8 The EPA's procedures for deriving AWQC are described in Stephan et al. (1985) and are
9 summarized here. With few exceptions, AWQC are derived based on data from aquatic toxicity
10 studies conducted in the laboratory. In general, both acute (short term) and chronic (long term)
11 AWQC are developed. Depending on the species, the toxicity studies considered for developing
12 acute criteria range in length from 48 to 96 hours. Acceptable endpoints for acute AWQC
13 development are mortality and/or immobilization, expressed as the median lethal concentration
14 (LCso) or median effect concentration (ECso). For each species, the geometric mean of the
15 acceptable LC50/EC50 data is calculated to determine the species mean acute value (SMAV).
16 For each genera, the geometric mean of the relevant SMAVs is then calculated to determine the
17 genus mean acute value (GMAV). The GMAVs are then ranked from high to low, and the final
18 acute value (FAV; the 5th percentile of the GMAVs, based on the four GMAVs surrounding the
19 5th percentile) is determined. Because the FAV is based on LCso/ECso values (which represent
20 unacceptably high levels of effect), the FAV is divided by two to estimate a low-effect level.
21 This value is then termed the acute criterion, or criterion maximum concentration (CMC). Based
22 on the most recent AWQC document for Pb (U.S. Environmental Protection Agency, 1985),
23 Table AX8-2.1.2 shows the freshwater SMAVs and GMAVs for Pb, and the resulting freshwater
24 CMC. Note that the freshwater AWQC are normalized for the hardness of the site water, as
25 discussed further below in Section AX8.2.1.3.
26 To develop chronic AWQC, acceptable chronic toxicity studies should encompass the full
27 life cycle of the test organism, although for fish, early life stage or partial life cycle toxicity
28 studies are considered acceptable. Acceptable endpoints include reproduction, growth and
29 development, and survival, with the effect levels expressed as the chronic value, which is the
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Table AX8-2.1.2. Development of Current Acute Freshwater Criteria for Lead
(U.S. Environmental Protection Agency, 1985)1
Rank
10
9
8
7
6
5
4
3
2
1
Species
Midge (Tanytarsus dissimilis)
Goldfish (Carassius auratus)
Guppy (Poecilia reticulatd)
Bluegill (Lepomis macrochirus)
Fathead minnow (Pimephales promelas)
Brook trout (Salvelinus fontinalis)
Rainbow trout (Oncorhynchus mykiss)
Snail (Aplexa hypnorum)
Cladoceran (Daphnia magnd)
Amphipod (Gammarus pseudolimnaeus)
GMAV
(^g/L)
235,900
101,100
66,140
52,310
25,440
4,820
2,448
1,040
447.8
142.6
FAV
CMC
SMAV
(^g/L)
235,900
101,100
66,140
52,310
25,440
4,820
2,448
1,040
447.8
142.6
= 67.54 jig/L
= 33.77 (ig/L
1 All values are normalized to a hardness of 50 mg/L (see Section AX8.2.1.3).
1 geometric mean of the no-observed-effect concentration (NOEC)1 and the lowest-observed-
2 effect concentration (LOEC)2. Although a chronic criterion could be calculated as the 5th
3 percentile of genus mean chronic values (GMCVs), sufficient chronic toxicity data are generally
4 lacking, as is the case for Pb. Consequently, an acute-chronic ratio (ACR) is typically applied to
5 the FAV to derive the chronic criterion. As the name implies, the ACR is the ratio of the acute
6 LCso to the chronic value, based on studies with the same species and in the same dilution water.
1 The NOEC is the highest concentration tested that did not result in statistically significant effects relative
to the control.
2 The LOEC is the lowest concentration tested that resulted in statistically significant effects relative to
the control.
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1 For Pb, the final ACR is 51.29, which results in a final chronic value (FCV) of 1.317 |ig/L (at a
2 hardness of 50 mg/L).
3 The U.S. EPA guidelines for developing AWQC (Stephan et al., 1985) are now more than
4 20 years old and thus are not reflective of scientific advances in aquatic toxicology and risk
5 assessment that have developed since the 1980s. For example, the toxicological importance of
6 dietary metals has been increasingly recognized and approaches for incorporating dietary metals
7 into regulatory criteria are being evaluated (Meyer et al., 2005). Other issues include
8 consideration of certain sublethal endpoints that are currently not directly incorporated into
9 AWQC development (e.g., endocrine toxicity, behavioral responses) and protection of threatened
10 and endangered (T&E) species (U.S. Environmental Protection Agency, 2003). In deriving
11 appropriate and scientifically defensible air quality criteria for Pb, it will be important that the
12 state-of-the-science for metals toxicity in aquatic systems be incorporated into the development
13 process.
14 Subsequent sections summarize some of the toxicity studies that meet the AWQC
15 development guidelines, with an emphasis on key studies published since the last Pb AWQC
16 were derived in 1984.
17
18 AX8.2.1.3 Ambient Water Quality Criteria: Unavailability Issues
19 In surface waters, the environmental fate of metal contaminants is mitigated through
20 adsorption, complexation, chelation, and other processes that affect bioavailability. The toxicity
21 of divalent cations tends to be highest in soft waters with low concentrations of dissolved organic
22 matter and suspended particles. In an acidic environment (pH <4), the ionic form of most metals
23 generally predominates and is considered to be the more toxic form. As the pH increases,
24 carbonate, oxide, hydroxide, and sulfide complexes of the metals tend to predominate, and tend
25 to be less toxic (Florence, 1977; Miller and Mackay, 1980). The portion of dissolved metal
26 available for uptake or bioaccumulation is influenced by modifying factors that "sequester" the
27 metal in an environmental matrix, thereby reducing the bioavailability of the metal at the sites of
28 action. Metals can become complexed (bound) to a ligand that can make metals either more
29 toxic (via transport mechanisms) or less toxic (by changing the metal's biological activity).
30 Metals that complex tightly to ligands generally are not readily bioavailable and, thus, are less
31 toxic to aquatic biota than their free-metal ion counterparts (Carlson et al., 1986; McCarthy,
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1 1989). There are many kinds of ligands, organic and inorganic, as well as natural and man-
2 made. Ligands found in natural surface waters and municipal and industrial effluent discharges
3 include glycine, ammonia, oxalate, humic or fulvic acids, hydroxide, carbonate, bicarbonate,
4 chloride, and hydrogen sulfide (Stumm and Morgan, 1970; Martin, 1986; Pagenkopf, 1986).
5 Recognizing the importance of calcium and magnesium ions (hardness) in modifying Pb
6 toxicity, the current freshwater AWQC for Pb are normalized based on the hardness of the site
7 water. The acute freshwater criteria, for example, are 34, 82, and 200 |ig/L at hardness levels of
8 50, 100, and 200 mg/L (as CaCOs). Although it has been known for some time that other water
9 quality parameters such as pH, dissolved organic carbon (DOC), and alkalinity affect the
10 bioavailability of metals to aquatic biota, it was the relatively recent development of the biotic
11 ligand model (BLM) that allowed AWQC to consider all of these factors. Paquin et al. (2002)
12 provided a thorough review of the factors influencing metal bioavailability and how research
13 over the last few decades has culminated in the development of the BLM.
14 By understanding the binding affinities of various natural ligands in surface waters and
15 how the freshwater fish gill interacts with free cations in the water, one can predict how metals
16 exert their toxic effects (Schwartz et al., 2004). Models developed prior to the BLM are the firee-
17 ion activity model (FLAM) and the gill surface interaction model (GSIM). The FIAM accounts
18 for the binding of free-metal ion and other metal complexes to the site of toxic action in an
19 organism; and it also considers competition between metal species and other cations (Paquin
20 et al., 2002). The GSIM is fundamentally similar to the FIAM in that it accounts for competition
21 between metal ions and hardness cations at the physiological active gill sites, but whereas the
22 FIAM is largely conceptual, the GSIM was used in interpreting toxicity test results for individual
23 metals and metal mixtures (Pagenkopf, 1983). The BLM was adapted from the GSIM and uses
24 the biotic ligand, rather than the fish gill as the site of toxic action (Di Toro et al., 2001; Paquin
25 et al., 2002). This approach, therefore, considers that the external fish gill surface contains
26 receptor sites for metal binding (Schwartz et al., 2004) and that acute toxicity is associated with
27 the binding of metals to defined sites (biotic ligands) on or within the organism (Paquin et al.,
28 2002). The model is predicated on the theory that mortality (or other toxic effects) occurs when
29 the concentration of metal bound to biotic ligand exceeds a threshold concentration (Di Toro
30 et al., 2001; Paquin et al., 2002). Direct uptake via the gills is thought to be the pathway for Pb
31 uptake in freshwater fish (Merlini and Pozzi, 1977; Hodson et al., 1978). Free metal cations "out
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1 compete" other cations and bind to the limited number of active receptor sites on the gill surface,
2 which may ultimately result in suffocation and/or disruption of ionoregulatory mechanisms in the
3 fish, leading to death (Di Toro et al., 2001; Paquin et al., 2002). Because the BLM uses the
4 biotic ligand (not the fish gill) as the site of action, the model can be applied to other aquatic
5 organisms, such as crustaceans, where the site of action is directly exposed to the aqueous
6 environment (Di Toro et al., 2001).
7 Although the BLM is currently being considered as a tool for regulating metals on a site-
8 specific basis, there are potential limitations in using the BLM to regulate metals in surface
9 waters that should be understood in developing air quality criteria for lead. For example, dietary
10 metals have also been shown to contribute to uptake by aquatic biota and, in some cases,
11 increased toxicity. Besser et al. (2005) observed that chronic (42-day) Pb toxicity to the
12 amphipod Hyalella azteca was greater from a combined aqueous and dietary exposure than from
13 a water-only exposure. The feasibility of incorporating dietary metals into BLMs is under
14 investigation. Furthermore, chronic exposures are typically of greatest regulatory concern, but
15 chronic BLMs to date have received limited attention (De Schamphelaere and Janssen, 2004).
16 There are also other ligands not accounted for in the BLM that require more research. Bianchini
17 and Bowles (2002) emphasized the importance of reduced sulfur as a metal ligand, limitations in
18 scientific knowledge on reduced sulfur, and provided recommendations for studies necessary to
19 incorporate sulfide ligands into the BLM.
20 To date, the EPA has incorporated the BLM into draft freshwater criteria for copper, but
21 the BLM is likely to be also included in the revised Pb criteria.
22
23 AX8.2.1.4 Sediment Quality Criteria: Development and Unavailability Issues
24 As with metals in surface waters, the environmental fate of metal contaminants in
25 sediments is moderated through various binding processes that reduce the concentration of free,
26 bioavailable metal. Sediments function as a sink for Pb, as with most metals. Lead compounds
27 such as Pb-carbonates, Pb-sulfates, and Pb-sulfides predominate in sediments (Prosi, 1989).
28 Total Pb has a higher retention time and a higher percentage is retained in sediments compared to
29 copper and zinc (Prosi, 1989). Lead is primarily accumulated in sediments as insoluble Pb
30 complexes adsorbed to suspended particulate matter. Naturally occurring Pb is bound in
31 sediments and has a low geochemical mobility (Prosi, 1989). Organic-sulfide and moderately
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1 reducible fractions are less mobile, whereas cation-exchangeable fractions and easily-reducible
2 fractions are more mobile and more readily bioavailable to biota (Prosi, 1989). Most Pb
3 transported in surface waters is in a particulate form, originating from the erosion of sediments in
4 rivers or produced in the water column (Prosi, 1989).
5 Sediment quality criteria have yet to be adopted by the EPA, but an equilibrium
6 partitioning procedure has recently been published (U.S. Environmental Protection Agency,
7 2005c). The EPA has selected an equilibrium partitioning approach because it explicitly
8 accounts for the bioavailability of metals. This approach is based on mixtures of cadmium,
9 copper, Pb, nickel, silver, and zinc. Equilibrium partitioning (EqP) theory predicts that metals
10 partition in sediment between acid-volatile sulfide, pore water, benthic organisms, and other
11 sediment phases such as organic carbon. When the sum of the molar concentrations of
12 simultaneously extracted metal (ZSEM) minus the molar concentration of AVS is less than zero,
13 it can accurately be predicted that sediments are not toxic because of these metals. Note that this
14 approach can be used to predict the lack of toxicity, but not the presence of toxicity. It is
15 important to emphasize that metals must be evaluated as a mixture using this approach.
16 If individual metals, or just two or three metals, are measured in sediment, ZSEM would be
17 misleadingly small and it may inaccurately appear that ZSEM / AVS is less than 1.0.
18 If ZSEM / AVS is normalized to the organic carbon fraction (i.e., (ZSEM / AVS)//OC),
19 mortality can be more reliably predicted by accounting for both the site-specific organic carbon
20 and AVS concentrations. When evaluating a metal mixture containing cadmium, copper, Pb,
21 nickel, silver, and zinc, the following predictions can be made (U.S. Environmental Protection
22 Agency, 2005c):
23 • A sediment with (SEM / AVS)//bc < 130 jimol/goc should pose low risk of adverse
24 biological effects due to these metals.
25 • A sediment with 130 jimol/goc < (SEM / AVS)//bc < 3000 jimol/goc may have adverse
26 biological effects due to these metals.
27 • In a sediment with (SEM / AVS)//bc > 3000 jimol/goc, adverse biological effects may
28 be expected.
29 A third approach is to measure pore water concentrations of cadmium, copper, Pb, nickel,
30 and zinc and then divide the concentrations by their respective FCVs. If the sum of these
31 quotients is <1.0, these metals are not expected to be toxic to benthic organisms.
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1 It should be noted that the AVS-SEM approach may not be relevant to benthic organisms
2 that ingest sediment particles. For example, Griscom et al. (2002) found that metals associated
3 with either reduced or oxidized sediment particles can be assimilated by deposit and suspension
4 feeding bivalve species due to the low pH and moderate reducing conditions in bivalve guts.
5 Lee et al. (2000) agree that metal concentrations in pore water may be mostly controlled by
6 equilibration with metal sulfides in sediments, but they argue that metal exposure by benthic
7 organisms is not necessarily controlled only by porewater. In addition, some studies suggest that
8 AVS-SEM measurements in the natural environment must be interpreted cautiously as AVS can
9 be quite variable with sediment depth and season (Van den Berg et al., 1998). Thus, although
10 the AVS-SEM approach for developing sediment quality criteria is being pursued by the U.S.
11 EPA, there is clearly not scientific consensus on this approach, at least not for all circumstances.
12 Many alternative approaches for developing sediment quality guidelines are based on
13 empirical correlations between metal concentrations in sediment to associated biological effects,
14 based on sediment toxicity tests (Long et al., 1995; Ingersoll et al., 1996; MacDonald et al.,
15 2000). However, these guidelines are based on total metal concentrations in sediment and do not
16 account for the bioavailability of metals between sediments. Sediment quality guidelines
17 proposed for Pb from these other sources are shown in Table AX8-2.1.3.
Table AX8-2.1.3. Recommended Sediment Quality Guidelines for Lead
Source
MacDonald et al. (2000)
Ingersoll etal. (1996)
Long etal. (1995)
Water Type
Freshwater
Freshwater
Saltwater
Guideline Type
TEC
PEC
ERL
ERM
ERL
ERM
Cone, (mg/kg dw)
35.8
128
55
99
46.7
218
TEC = Threshold effect concentration; PEC = Probable effect concentration; ERL = Effects range - low;
ERM = Effects range - median
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1 AX8.2.1.5 Metal Mixtures
2 As discussed above, the EPA's current approach for developing sediment criteria for Pb
3 and other metals is to consider the molar sum of the metal concentrations (ZSEM). Although a
4 similar approach has not been applied to AWQC, metal mixtures have been shown to be more
5 toxic than individual metals (Spehar and Fiandt, 1986; Enserink et al., 1991). Spehar and Fiandt
6 (1986) evaluated the acute and chronic toxicity of a metal mixture (arsenic, cadmium, chromium,
7 copper, mercury, and Pb) to fathead minnows (Pimephalespromelas) and a daphnid
8 (Ceriodaphnia dubid). In acute tests, the joint toxicity of these metals was observed to be more
9 than additive for fathead minnows and nearly strictly additive for daphnids. In chronic tests, the
10 joint toxicity of the metals was less than additive for fathead minnows and nearly strictly
11 additive for daphnids. One approach for considering the additive toxicity of Pb with other metals
12 is to use the concept of toxic units (TUs). Toxic units for each component of a metal mixture are
13 derived by dividing metal concentrations by their respective acute or chronic criterion. The TUs
14 for all the metals in the mixture are then summed. A ETU > 1.0 suggests the metal mixture is
15 toxic (note that this is the same approach as discussed above for developing metal sediment
16 criteria based on pore water concentrations). According to Norwood et al. (2003), the TU
17 approach is presently the most appropriate model for predicting effects of metal mixtures based
18 on the currently available toxicity data. However, it should also be emphasized that the TU
19 approach is most appropriate at a screening level, because the true toxicity of the mixture is
20 dependent on the relative amounts of each metal. The TU approach is also recommended with
21 mixtures containing less than six metals.
22 Lead and other metals often co-occur in sediments with other toxicants, such as organic
23 contaminants. Effects-based sediment quality guidelines (SQGs) have been developed over the
24 past 20 years to aid in the interpretation of the relationships between complex chemical
25 contamination and adverse biological effects (Long et al., 2006). Mean sediment quality
26 guideline quotients (mSQGQs) can be calculated by dividing the concentrations of chemicals in
27 sediments by their respective SQGs and then calculating the mean of the quotients for the
28 individual chemicals. Long et al. (2006) performed a critical review of this approach and found
29 that it reasonably predicts the incidence and magnitude of toxicity in laboratory tests and the
30 incidence of impairment to benthic communities increases incrementally with increasing
31 mSQGQs. However, the authors pointed out some of the limitations of this approach, such as a
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1 lack of agreement on the level of mSQGQs, masking of an individual chemical's effect due to
2 data aggregation, lack of SQGs for all chemicals of concern, and mSQGQs were not initially
3 derived as a regulatory standard or criterion, thus there is a reluctance to use them in
4 enforcement or remediation (Long et al., 2006).
5 For assessing Pb effects on aquatic ecosystems, it is not truly feasible to account for metal
6 mixtures, because these will obviously vary highly from site to site. However, the toxicity of
7 metal mixtures in surface water should be considered on a site-specific basis.
8
9 AX8.2.1.6 Background Lead
10 Because Pb is naturally occurring, it is found in all environmental compartments
11 including surface water, sediment, and aquatic biota. Background Pb concentrations are spatially
12 variable depending on geological features and local characteristics that influence Pb speciation
13 and mobility. In the European Union risk assessments for metals, an "added risk" approach has
14 been considered that assumes only the amount of metal added above background is relevant in a
15 toxicological evaluation. However, this approach ignores the possible contribution of
16 background metal levels to toxic effects, and background metal levels are regionally variable,
17 precluding the approach from being easily transferable between sites. In terms of deriving
18 environmental criteria for Pb, background levels should be considered on a site-specific basis if
19 there is sufficient information that Pb concentrations are naturally elevated. As discussed
20 previously, the use of radiogenic Pb isotopes is useful for source apportionment.
21
22 AX8.2.2 Distribution of Lead in Aquatic Ecosystems
23 Atmospheric Pb is delivered to aquatic ecosystems primarily through deposition (wet
24 and/or dry) or through erosional transport of soil particles (Baier and Healy, 1977; Dolske and
25 Sievering, 1979). A number of physical and chemical factors govern the fate and behavior of Pb
26 in aquatic systems. The EPA summarized some of these controlling factors in the 1986 Pb
27 AQCD (U.S. Environmental Protection Agency, 1986a). For example, the predominant form of
28 Pb in the environment is in the divalent (Pb2+) form and complexation with inorganic and
29 organic ligands is dependent on pH (Lovering, 1976; Rickard and Nriagu, 1978). A significant
30 portion of Pb in the aquatic environment exists in the undissolved form (i.e., bound to suspended
31 particulate matter). The ratio of Pb in suspended solids to Pb in filtrate varies from 4:1 in rural
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1 streams to 27:1 in urban streams (Getz et al., 1977). In still waters, Pb is removed through
2 sedimentation at a rate determined by temperature, pH, oxidation-reduction (redox) potential,
3 organic content, grain size, and chemical form of Pb in the water and biological activities (Jenne
4 and Luoma, 1977). Since the publication of the 1986 Pb AQCD (U.S. Environmental Protection
5 Agency, 1986a), knowledge of the properties of Pb in aquatic ecosystems has expanded. This
6 section will provide further detail on the chemical species and the environmental factors
7 affecting speciation of Pb in the aquatic environment. In addition, quantitative distributions of
8 Pb in water, sediment, and biological tissues will be presented for aquatic ecosystems throughout
9 the United States. Finally, recent studies discussing the tracing of Pb in aquatic systems will be
10 summarized.
11
12 AX8.2.2.1 Speciation of Lead in Aquatic Ecosystems
13 The speciation of Pb in the aquatic environment is controlled by many factors. The
14 primary form of Pb in aquatic environments is divalent (Pb2+), while Pb4+ exists only under
15 extreme oxidizing conditions (Rickard and Nriagu, 1978). Labile forms of Pb (e.g., Pb2+,
16 PbOH+, PbCOs) are a significant portion of the Pb inputs to aquatic systems from atmospheric
17 washout. Lead is typically present in acidic aquatic environments as PbSO4, PbCl4, ionic Pb,
18 cationic forms of Pb-hydroxide, and ordinary Pb-hydroxide (Pb(OH)2). In alkaline waters,
19 common species of Pb include anionic forms of Pb-carbonate (Pb(CC>3)) and Pb(OH)2.
20 Speciation models have been developed based on the chemical equilibrium model developed by
21 Tipping (1994) to assist in examining metal speciation. The EPA MINTEQA2 computer model
22 (http://www.epa.gov/ceampubl/mmedia/minteq/) is one such equilibrium speciation model that
23 can be used to calculate the equilibrium composition of dilute aqueous solutions in the laboratory
24 or in natural aqueous systems. The model is useful for calculating the equilibrium mass
25 distribution among dissolved species, adsorbed species, and multiple solid phases under a variety
26 of conditions, including a gas phase with constant partial pressures. In addition to chemical
27 equilibrium models, the speciation of metals is important from a toxicological perspective.
28 The BLM was developed to study the toxicity of metal ions in aquatic biota and was previously
29 described in Section AX8.2.1.3. Further detail on speciation models is not provided herein,
30 rather a general overview of major speciation principles are characterized in the following
31 sections.
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1 Acidity (pH)
2 Freshwater
3 Most of the Pb in aquatic environments is in the inorganic form (Sadiq, 1992). The
4 speciation of inorganic Pb in freshwater aquatic ecosystems is dependent upon pH and the
5 available complexing ligands. Solubility varies according to pH, temperature, and water
6 hardness (Weber, 1993). Lead rapidly loses solubility above pH 6.5 (Rickard and Nriagu, 1978)
7 and as water hardness increases. In freshwaters, Pb typically forms strong complexes with
8 inorganic OH and CO32 and weak complexes with Cl (Long and Angino, 1977; Bodek et al.,
9 1988). The primary form of Pb at low pH (<6.5) is predominantly Pb2+ and less abundant
10 inorganic forms include Pb(HCO)3, Pb(SO4)22~, PbCl, PbCO3, and Pb2(OH)2CO3
11 (Figure AX8-2.2.1). At higher pH (>7.5), Pb forms hydroxide complexes (PbOH+, Pb(OH)2,
12 Pb(OH)3 , Pb(OH)42 ).
13 Organic compounds in surface waters may originate from natural (e.g., humic or fulvic
14 acids) or anthropogenic sources (e.g., nitrilotriacetonitrile and ethylenediaminetetraacetic acid
15 [EDTA]) (U.S. Environmental Protection Agency, 1986b). The presence of organic complexes
16 has been shown to increase the rate of solution of Pb bound as Pb-sulfide (Lovering, 1976).
17 Soluble organic Pb compounds are present at pH values near 7 and may remain bound at pH
18 values as low as 3 (Lovering, 1976; Guy and Chakrabarti, 1976). At higher pH (7.4 to 9), Pb-
19 organic complexes are partially decomposed. Water hardness and pH were found to be
20 important in Pb-humic acid interactions (O' Shea and Mancy, 1978). An increase in pH
21 increased the concentration of exchangeable Pb complexes, while an increase in hardness tended
22 to decrease the humic acid-Pb interactions. Thus, the metals involved in water hardness
23 apparently inhibit the exchangeable interactions between metals and humic acids.
24
25 Marine Water
26 In marine systems, an increase in salinity increases complexing with chloride and
27 carbonate ions and reduces the amount of free Pb2+. In seawaters and estuaries at low pH, Pb is
28 primarily bound to chlorides (PbCl, PbCl2, PbCl37, PbCU2 ) and may also form inorganic
29 Pb(HCO)3, Pb(SO4)22~, or PbCO3. Elevated pH in saltwater environments results in the
30 formation of Pb hydroxides (PbOH+, Pb(OH)2, Pb(OH)3 , Pb(OH)42 ) (Figure AX8-2.2.2).
May 2006 AX8-118 DRAFT-DO NOT QUOTE OR CITE
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100
c
o
Pb(OH),"(aq)
Pb(CO3),
PbOH~
PH
Figure AX8-2.2.1. Distribution of aqueous lead species as a function of pH based on
a concentration of 1 ug Pb/L (U.S. Environmental Protection
Agency, 1999).
1 A recent examination of Pb species in seawater as a function of chloride concentration suggested
2 that the primary species were PbCl3~ > PbCO3 > PbCl2 > PbCl+ > and Pb(OH)+ (Fernando,
3 1995). Lead in freshwater and seawater systems is highly complexed with carbonate ligands
4 suggesting that Pb is likely to be highly available for sorption to suspended materials (Long and
5 Angino, 1977).
6 Current information suggests that inorganic Pb is the dominant form in seawater;
7 however, it has been shown that organically bound Pb complexes make up a large portion of the
8 total Pb (Capodaglio et al., 1990).
9
10 Sorption
11 Sorption processes (i.e., partitioning of dissolved Pb to suspended particulate matter or
12 sediments) appear to exert a dominant effect on the distribution of Pb in the environment
13 (U.S. Environmental Protection Agency, 1979). Sorption of Pb results in the enrichment of
14 bed sediments, particularly in environments with elevated organic matter content from
May 2006
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c
o
*3
2
•+-•
c
-------
1 (Gao et al., 2003). Changes in water chemistry (e.g., reduced pH or ionic composition) can
2 cause sediment Pb to become remobilized and potentially bioavailable to aquatic organisms
3 (Weber, 1993).
4
5 Biotramformation
6 Methylation may result in Pb remobilization and reintroduction into the aqueous
7 environment compartment and its subsequent release into the atmosphere (Syracuse Research
8 Corporation., 1999). However, methylation is not a significant environmental pathway
9 controlling the fate of Pb in the aquatic environment. The microbial methylation of Pb in aquatic
10 systems has been demonstrated experimentally, but evidence for natural occurrence is limited
11 (Beijer and Jernelov, 1984; DeJonghe and Adams, 1986). Reisinger et al. (1981) examined the
12 methylation of Pb in the presence of numerous bacteria known to alkylate metals and did not find
13 evidence of Pb methylation under any test condition. Tetramethyl-Pb may be formed by the
14 methylation of Pb-nitrate or Pb-chloride in sediments (Bodek et al., 1988). However,
15 tetramethyl-Pb is unstable and may degrade in aerobic environments after being released from
16 sediments (U.S. Environmental Protection Agency, 1986b). Methylated species of Pb may also
17 be formed by the decomposition of tetralkyl-Pb compounds (Radojevic and Harrison, 1987;
18 Rhue et al., 1992). Sadiq (1992) reviewed the methylation of Pb compounds and suggested that
19 chemical methylation of Pb is the dominant process and that biomethylation is of secondary
20 importance.
21
22 AX8.2.2.2 Spatial Distribution of Lead in Aquatic Ecosystems
23 The 1986 Pb AQCD (U.S. Environmental Protection Agency, 1986b) did not describe the
24 distribution and concentration of Pb throughout aquatic ecosystems of the United States.
25 Consequently, an analysis of readily available data on Pb concentrations was conducted to
26 determine the distribution of Pb in the aquatic environment. Data from the United States
27 Geological Survey (USGS) National Water-Quality Assessment (NAWQA) program were
28 queried and retrieved. NAWQA contains data on Pb concentrations in surface water, bed
29 sediment, and animal tissue for more than 50 river basins and aquifers throughout the country,
30 and it has been used by the EPA for describing national environmental concentrations for use in
31 developing AWQC. The authors recognize that the NAWQA program encountered analytical
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1 challenges with the chemical analysis of Pb in surface waters. The analytical methods available
2 during the NAWQA program were not as sensitive as methods currently applied today.
3 Therefore, analytical detection limits are elevated and a large portion of the data set contains
4 non-detected values. Nevertheless, this data provides a comprehensive overview of Pb
5 concentrations in U.S. surface waters that is supplemented with data from other relevant studies.
6 The following sections describe the estimated concentrations of Pb from NAWQA and other
7 research programs.
8 NAWQA data are collected during long-term, cyclical investigations wherein study units
9 undergo intensive sampling for 3 to 4 years, followed by low-intensity monitoring and
10 assessment of trends every 10 years. The NAWQA program's first cycle was initiated in 1991;
11 therefore, all available data are less than 15 years old. The second cycle began in 2001 and is
12 ongoing; data are currently available through 30 September 2003. The NAWQA program study
13 units were selected to represent a wide variety of environmental conditions and contaminant
14 sources; therefore, agricultural, urban, and natural areas were all included. Attention was also
15 given to selecting sites covering a wide variety of hydrologic and ecological resources.
16 NAWQA sampling protocols are designed to promote data consistency within and among
17 study units while minimizing local-scale spatial variability. Water-column sampling is
18 conducted via continuous monitoring, fixed-interval sampling, extreme-flow sampling, as well as
19 seasonal, high-frequency sampling in order to characterize spatial, temporal, and seasonal
20 variability as a function of hydrologic conditions and contaminant sources. Sediment and tissue
21 samples are collected during low-flow periods during the summer or fall to reduce seasonal
22 variability. Where possible, sediment grab samples are collected along a 100-m stream reach,
23 upstream of the location of the water-column sampling. Five to ten deposit!onal zones at various
24 depths, covering left bank, right bank, and center channel, are sampled to ensure a robust
25 representation of each site. Fine-grained samples from the surficial 2 to 3 cm of bed sediment at
26 each depositional zone are sampled and composited. Tissue samples are collected following a
27 National Target Taxa list and decision trees that help guide selection from that list to
28 accommodate local variability.
29 The NAWQA dataset was chosen over other readily available national databases (i.e. the
30 USEPA-maintained database for the STOrage and RETrieval [STORET] of chemical, physical,
31 and biological data), because the study design and methods used to assess the water quality of
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1 each study unit are rigorous and consistent, and, as such, these data may be presented with a high
2 level of confidence. This is in stark contrast to the STORET database, which essentially serves
3 as a depot for any organization wishing to share data they have generated. This lack of a
4 consistent methodology or QA/QC protocol has lead to the STORET data being highly qualified
5 and offered with only a mild level of confidence. Furthermore, because there is no standard for
6 site selection within STORET, the database may be biased toward contaminated sites. Finally,
7 and, perhaps most importantly, the majority of the available Pb data in STORET predate the use
8 of clean techniques for Pb quantification.
9 The authors recognize the existence of several local and regional datasets that may be of
10 quality equal to NAWQA; however, due to the national scope of this assessment, these datasets
11 were not included in the following statistical analyses. However, because the NAWQA database
12 does not cover lakes and the marine/estuarine environment, and we were unable to identify any
13 monitoring data of similar quality, local and regional datasets were used in these cases to provide
14 general information on environmental Pb concentrations.
15
16 Data Acquisition and Analysis
17 The following data were downloaded for the entire United States (all states) from the
18 NAWQA website (http://water.usgs.gov/nawqa/index.html): site information, dissolved Pb
19 concentration in surface water (|ig/L), total Pb concentration (|ig/g) in bed sediment (<63 |im)3,
20 and Pb concentration in animal tissue (|ig/g dw). Using the land use classification given for each
21 site, the data were divided into two groups: "natural" and "ambient" (Table AX8-2.2.1).
22 All samples were considered to fall within the ambient group (the combined contribution of
23 natural and anthropogenic sources), whereas the natural group comprised "forest," "rangeland,"
24 or "reference" samples only4. These groups follow those defined and recommended for use by
25 the EPA's Framework for Inorganic Metals Risk Assessment (U.S. Environmental Protection
26 Agency, 2004b). Finally, in addition to the natural/ambient classification, tissue samples were
27 further divided into "whole organism" and "liver" groups.
28
3 NAWQA sediment samples are sieved to <63 um to promote the collection of fine-grained surficial
sediments, which are natural accumulators of trace elements.
4 The authors acknowledge that while Pb samples collected from sites classified under these three land use
categories will most closely reflect natural background concentrations, atmospheric input of lead may be present.
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Table AX8-2.2.1. NAWQA Land Use Categories and Natural/Ambient
Classification
NAWQA Land Use Categories
Agricultural
Commercial/Industrial
Cropland
Forest
Mining
Mixed
NA
Orchard/Vineyard
Other/Mixed
Pasture
Rangeland
Reference
Residential
Urban
Classification
Ambient
Ambient
Ambient
Ambient/Natural
Ambient
Ambient
Ambient
Ambient
Ambient
Ambient
Ambient/Natural
Ambient/Natural
Ambient
Ambient
1 All data were compiled in spreadsheets wherein non-detect values were converted to one-
2 half the detection limit and the total number of samples, percentage of non-detect values (percent
3 censorship), minimum, maximum, median, mean, standard deviation, and cumulative density
4 functions were calculated for each endpoint for both the natural and ambient groups.
5 As discussed below, some datasets were highly censored; however, deletion of non-detect data
6 has been shown to increase the relative error in the mean to a greater extent than inclusion of
7 non-detects as 1A of the detection limit (Newman et al., 1989); therefore means and other
8 statistics were calculated using the latter method for this analysis. Finally, since all data were
9 geo-referenced, a geographic information system (GIS; ArcGIS) was used to generate maps,
10 conduct spatial queries and analyses, and calculate statistics.
11
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1 Lead Distributions Generated from the NA WQA Database
2 Natural versus Ambient Groups
3 There were four to eight times more ambient surface water (Table AX8-2.2.2) and bulk
4 sediment (Table AX8-2.2.3) samples in the compiled dataset than natural samples. This is most
5 likely a function of both the NAWQA program site selection process and the fact that sites
6 unaffected by human activities are extremely limited. The spatial distributions of natural and
7 ambient surface water/sediment sites were fairly comparable, with natural samples located in
8 almost all of the same areas as ambient samples except in the Midwest (Ohio, Illinois, Iowa, and
9 Michigan), where natural sites were not present (Figure AX8-2.2.3). This exception may be
10 because these areas are dominated by agricultural and urban areas. The same spatial
11 distributions were observed for the natural and ambient liver and whole organism tissue samples
12 (Figure AX8-2.2.4 and Figure AX8-2.2.5).
13
14
Table AX8-2.2.2. Summary Statistics of Ambient and Natural Levels of
Dissolved Lead in Surface Water
Surface Water Dissolved Pb (ug/L)
Statistic
% Censorship
N
Minimum
Maximum
Mean
Standard Deviation
95th Percentile
96th Percentile
97th Percentile
98th Percentile
99th Percentile
Natural
87.91
430
0.04
8.40
0.52
0.59
0.50
0.67
1.00
1.79
2.48
Ambient
85.66
3445
0.04
29.78
0.66
1.20
1.10
2.00
2.34
3.58
5.44
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Table AX8-2.2.3. Summary Statistics of Ambient and Natural Levels of Total Lead in
<63 jim Bulk Sediment
Statistic
% Censorship
N
Minimum
Maximum
Mean
Standard Deviation
Median
95th percentile
Bulk Sediment <63
Natural
1.16
258
0.50
12000
109.07
786.74
22.00
161.50
um Total Lead (ug/g)
Ambient
0.48
1466
0.50
12000
120.11
672.41
28.00
200.00
1 Surface Water
2 The total number of surface water Pb samples was 3,445; however these data were highly
3 censored with 85.66% of the ambient samples (2951/3445) and 87.91% of the natural samples
4 (378/430) below the detection limit5 (Table AX8-2.2.2). Consequently, the majority of the
5 variability between these two datasets fell between the 95th and 100th (maximum) percentiles,
6 as was shown by the frequency distributions of the two groups deviating only at the upper and
7 lower tails with most of the overlapping data falling at 0.50 |ig/L (one-half of the most common
8 detection limit, 1.0 |ig/L; Figure AX8-2.2.6). As expected, due to the definitions of the natural
9 and ambient groups, the 95th and 100th percentiles were consistently higher for the ambient
10 samples than the natural samples. Similarly, the mean ambient Pb concentration (0.66 |ig/L) was
11 higher than the mean natural Pb concentration (0.52 |ig/L).6
12
5 The NAWQA dataset contains multiple detection limits for Pb in surface water that have decreased over
time. While the majority of data were analyzed with a detection limit of 1.0 ug/L (before 2000/2001), the most
recent samples were analyzed with either a 0.5, 0.2, 0.16, or 0.08 ug/L detection limit (after 2000/2001), and some
older samples (N = 20) were analyzed with a detection limit of 2.0 ug/L.
6 The same pattern was observed upon calculating the mean Pb concentrations based on detect data only
(ambient mean = 1.66 ug/L, natural mean = 0.87 ug/L); however, as previously discussed, calculations included
non-detect data as 1A of the detection limit to reduce the relative error in the mean.
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• Natural Surface Water/Sediment Sites
o Ambient Surface Water/Sediment Sites
Figure AX8-2.2.3. Spatial distribution of natural and ambient surface water/sediment sites (Surface water: natural N = 430,
ambient N = 3445; Sediment: natural N = 258, ambient N = 1466).
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• Natural Whole Organism Sites
O Ambient Whole Organism Sites
Figure AX8-2.2.5. Spatial distribution of natural and ambient whole organism tissue sample sites (Natural N = 93,
Ambient N = 332).
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100
- 80
> 60
"+-
"= 40
O 20
Ambient Natural
0.0 0.1 1.0 10.0 100.0
Surface Water Dissolved Pb (ug/L)
Figure AX8-2.2.6. Frequency distribution of ambient and natural levels of surface water
dissolved lead (ug/L).
4
5
6
7
8
9
10
Due to the preponderance of non-detectable (ND) measurements, assessing national trends
in surface water-dissolved Pb concentrations was not possible. However, areas with elevated Pb
concentrations were identified by classifying the data with detectable Pb concentrations above
and below the 99th percentile. The 99th percentile (versus the 95th percentile) was chosen in
this instance to represent extreme conditions given the small window of variability in the dataset.
By convention, the 95th percentile was used in subsequent analyses of this type. Areas with high
surface water Pb concentrations were observed in Washington, Idaho, Utah, Colorado, Arkansas,
and Missouri (Figure AX8-2.2.7). The maximum measured Pb concentration was located in
Canyon Creek at Woodland Park, ID, a site classified as mining land use.
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Legend
Surface Water Dissolved Pb (M9/L)
o Non-detect
0 0.51 - 5.44 (<99th precentile)
5.45 - 29.78 (>99th percentile)
Figure AX8-2.2.7. Spatial distribution of dissolved lead in surface water (N = 3445).
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1 Because the NAWQA database does not cover lakes or the sea where atmospheric
2 deposition of Pb is highly likely, the primary literature was searched for studies using ultra-clean
3 sampling/analytical techniques to characterize Pb concentrations in these environments. Lead
4 concentrations in lakes and oceans were generally found to be much lower than those measured
5 in the lotic waters assessed by NAWQA. Surface water concentrations of dissolved Pb measured
6 in Hall Lake, Washington in 1990 ranged from 2.1 - 1015.3 ng/L (Balistrieri et al., 1994).
7 Nriagu et al. (1996) found that the average surface water dissolved Pb concentrations measured
8 in the Great Lakes (Superior, Erie, and Ontario) between 1991 and 1993 were 3.2, 6.0, and
9 9.9 ng/L, respectively. Lead concentrations ranged from 3.2 - 11 ng/L across all three lakes.
10 Similarly, 101 surface water total Pb concentrations measured at the HOT station ALOHA
11 between 1998 and 2002 ranged from 25 - 57 pmol/kg (5-11 ng/kg; (Boyle et al., 2005). Based
12 on the fact that Pb is predominately found in the dissolved form in the open ocean (<90%;
13 Schaule and Patterson, 1981), dissolved Pb concentrations measured at these locations would
14 likely have been even lower than the total Pb concentrations reported.
15
16 Sediment
17 There were approximately one-half of the number of surface water data available for
18 sediments (N = 1466). In contrast to the surface water data, however, very few sediment data
19 were below the detection limit (7/1466 ambient ND, 3/258 natural ND; Table AX8-2.2.3).
20 As expected, the mean ambient Pb concentration was higher than the mean natural Pb
21 concentration (120.11 and 109.07 |ig/g, respectively). Similarly, the median ambient Pb
22 concentration was higher than the median natural Pb concentration (28.00 and 22.00 |ig/g,
23 respectively) and the ambient 95th percentile was higher than the natural 95th percentile
24 (200.00 and 161.50 jig/g, respectively). While the natural and ambient surface water Pb
25 distributions differed only at the extremes, the natural sediment Pb percentiles were consistently
26 lower than the ambient percentiles throughout the distributions (Figure AX8-2.2.8). Unlike the
27 surface water dataset, because the sediment dataset was not heavily censored, assessing national
28 trends in sediment Pb concentrations was possible. The data were mapped and categorized into
29 the four quartiles of the frequency distribution (Figure AX8-2.2.9). The following observations
30 were made:
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100
1 10 100 1000 10000 100000
Bulk Sediment <63um Total Pb (M9/g)
Figure AX8-2.2.8. Frequency distribution of ambient and natural levels of bulk sediment
<63 jim total Pb (ug/g).
1 • Sediment Pb concentrations generally increased from west to east (the majority of
2 sites along East Coast had Pb concentrations in the fourth quartile of the sediment Pb
3 concentration frequency distribution).
4 • Several "hot spots" of concentrated sites with elevated sediment Pb concentrations
5 were apparent in various western states.
6 • Sediment Pb concentrations were generally lowest in the midwestern states
7 (the majority of sites in North Dakota, Nebraska, Minnesota, and Iowa had Pb
8 concentrations in the first or second quartile of sediment Pb concentration
9 frequency distribution).
10 As was seen with surface water Pb concentrations, the highest measured sediment Pb
11 concentrations were found in Idaho, Utah, and Colorado. Not surprisingly, of the top 10
12 sediment Pb concentrations recorded, 7 were measured at sites classified as mining land use.
13
14 Tissue
15 As was true for the surface water data, there were a high number of tissue samples below
16 the detection limit (47/93 natural whole organism ND, 130/332 ambient whole organism ND,
17 74/83 natural liver ND, 398/559 ambient liver ND; Table AX8-2.2.4). In general, more
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Bulk Sediment <63|j Total Lead (|jg/g)
0 0.50- 18.00 (1st Quartile)
0 18.01 - 28.00 (2nd Quartile)
• 28.01 -49.00 (3rd Quartile)
• 49.01 -12000.00 (4th Quartile)
Figure AX8-2.2.9. Spatial distribution of total lead in bulk sediment <63 um (N = 1466).
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Table AX8-2.2.4. Summary Statistics of Ambient and Natural Levels of Lead
in Whole Organism and Liver Tissues
Tissue Pb (jig/g dry weight)
Statistic
% Censorship
N
Minimum
Maximum
Mean
Standard Deviation
Median
95th percentile
Whole
Natural
50.54
93
0.08
22.60
0.95
2.53
0.11
1.26
Organism
Ambient
39.16
332
0.08
22.60
1.03
1.74
0.15
1.06
Natural
89.16
83
0.01
3.37
0.28
0.54
0.35
2.50
Liver
Ambient
71.20
559
0.01
12.69
0.36
0.96
0.59
3.24
1 non-censored data were available for whole organism samples than liver samples, and for
2 ambient sites than natural sites. As expected, for whole organism samples, the 95th percentile Pb
3 concentration measured at ambient sites was higher than that measured at natural sites (3.24 and
4 2.50 |ig/g, respectively); however, Pb liver concentration 95th percentiles for ambient and
5 natural samples were very similar, with the natural 95th percentile actually higher than the
6 ambient 95th percentile (1.26 and 1.06 |ig/g, respectively). In addition, as expected, the median
7 and mean Pb liver concentrations of ambient samples (0.15 and 0.36 |ig/g, respectively) were
8 higher than the median and mean Pb liver concentrations of natural samples (0.11 and 0.28 |ig/g,
9 respectively). The same pattern was observed in the whole organism median and mean Pb
10 concentrations (ambient: median = 0.59, mean = 1.03; natural: median = 0.35, mean =
11 0.95 |ig/g). In addition, the frequency distributions of the liver and whole organism Pb
12 concentrations followed the same trends, with the natural percentiles consistently lower than the
13 ambient percentiles throughout the distributions (Figure AX8-2.2.10 and Figure AX8-2.2.11).
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100
* Ambient D Natural
0.01
0.1 1 10
Liver Pb (M9/9 dry weight)
100
Figure AX8-2.2.10. Frequency distribution of ambient and natural levels of lead in liver
tissue (ug/g dry weight).
100
* Ambient Natural
0.01 0.1 1 10 100
Whole Organism Pb (pg/g dry weight)
Figure AX8-2.2.11. Frequency distribution of ambient and natural levels of lead in whole
organism tissue (ug/g dry weight).
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1 These whole organism results were compared with findings from the 1984 U.S. Fish and
2 Wildlife Service (USFWS) National Contaminant Biomonitoring Program (NCBP) (Schmitt and
3 Brumbaugh, 1990). As part of this program, 321 composite samples of 3 to 5 whole, adult fish
4 of a single species were collected from 109 river and Great Lake stations throughout the country.
5 Samples were analyzed for Pb concentrations (|ig/g ww) and the geometric mean, maximum, and
6 85th percentile were calculated. Upon comparing these summary statistics with the equivalent
7 NAWQA ambient group value (NCBP stations were representative of both natural and
8 anthropogenically influenced conditions), a very strong agreement between the two analyses was
9 observed for each endpoint (Table AX8-2.2.5). For example, NCBP and NAWQA geometric
10 mean Pb concentrations were nearly identical (0.55 and 0.54 |ig/g dw, respectively) and the 85th
11 percentiles only differed by 0.5 |ig Pb/g dw (NCBP, 1.10 and NAWQA, 1.60). The authors
12 acknowledge that a high degree of censorship is present in both of these datasets and no firm
13 conclusions can be drawn by comparing these means. The objective of this exercise was limited
14 to showing how the NAWQA data compare to other national datasets.
Table AX8-2.2.5. Comparison of NCBP and NAWQA Ambient Lead Levels
in Whole Organism Tissues
Whole Organism Lead Concentration (jig/g dry weight)
Statistic NCBP1 NAWQA
Geometric Mean 0.55 0.54
Maximum 24.40 22.60
85th Percentile 1.10 1.60
1 To convert between wet and dry weight, wet weight values were multiplied by a factor of five.
15 As was the case with surface water data, the high amount of non-detectable measurements
16 did not allow for a national assessment of spatial trends in Pb tissue concentrations. Instead,
17 areas with high Pb tissue concentrations were identified by classifying the data above and below
18 the 95th percentile. Similar to surface water and sediments, tissue concentrations were found to
19 be elevated in Washington, Idaho, Utah, Colorado, Arkansas, and Missouri; however, several of
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1 the highest measured Pb concentrations were also found in study units in the southwestern and
2 southeastern states (Figure AX8-2.2.12 and Figure AX8-2.2.13). As expected, the majority of
3 the samples with elevated Pb concentrations were taken from sites classified as urban,
4 commercial/industrial, or mining.
5
6 AX8.2.2.3 Tracing the Fate and Transport of Lead in Aquatic Ecosystems
7 The following section presents a generalized framework for the fate and transport of Pb in
8 aquatic systems (Figure AX8-2.2.14). The primary source of Pb in natural systems is
9 atmospheric deposition (Rickard and Nriagu, 1978; U.S. Environmental Protection Agency,
10 1986a). Estimated median global atmospheric emission for anthropogenic and natural sources
11 are 332 x 106 kg/year and 12 x 106 kg/year, respectively (summarized by Giusti et al., 1993).
12 Inorganic and metallic Pb compounds are nonvolatile and will partition to airborne particulates
13 or water vapors (Syracuse Research Corporation., 1999). Dispersion and deposition of Pb is
14 dependent on the particle size (U.S. Environmental Protection Agency, 1986a; Syracuse
15 Research Corporation., 1999). More soluble forms of Pb will be removed from the atmosphere
16 by washout in rain.
17 In addition to atmospheric deposition, Pb may enter aquatic ecosystems through industrial
18 or municipal wastewater effluents, storm water runoff, erosion, or direct point source inputs
19 (e.g., Pb shot or accidental spills). Once in the aquatic environment, Pb will partition between
20 the various compartments of the system (e.g., dissolved phase, solid phase, biota). The
21 movement of Pb between dissolved and particulate forms is governed by factors such as pH,
22 sorption, and biotransformation (see Section AX8.2.2.1). Lead bound to organic matter will
23 settle to the bottom sediment layer, be assimilated by aquatic organisms, or be resuspended in the
24 water column. The uptake, accumulation, and toxicity of Pb in aquatic organisms from water
25 and sediments are influenced by various environmental factors (e.g., pH, organic matter,
26 temperature, hardness, bioavailability). These factors are further described in Section
27 AX8.2.3.4). The remainder of this section discusses some methods for describing the
28 distribution of atmospheric Pb in the aquatic environment.
29
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Liver Lead (|jg/g dry weight)
O Non-detect
0 0.10 -1.06 (<95th precentile)
1.06-12.69 (>95th precentile)
- „ - ' Qrto >V
'3~W??®i. '^$fe&i^
r, (Cn/"%. ; /+H1 '- ~Ji«l.)-:dU.
A
Figure AX8-2.2.12. Spatial distribution of lead in liver tissues (N = 559).
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Whole organism Lead (\iglg dry weight)
O Non-detect
0 0.20 - 3.24 (<95th precentile)
3.24 - 22.60 (>95th precentile)
Figure AX8-2.2.13. Spatial distribution of lead in whole organism tissues (N = 332).
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Atmospheric ~"
Precipitation/Deposition
Figure AX8-2.2.14. Lead cycle in an aquatic ecosystem.
1 Sediment Core Dating and Source Tracing
2 In addition to directly measuring Pb concentrations in various aquatic compartments (see
3 Section AX8.2.3.3), it is useful to study the vertical distribution of Pb. Sediment profiling and
4 core dating is a method used to determine the extent of accumulation of atmospheric Pb and
5 provide information on potential anthropogenic sources. Sediment concentration profiles are
6 typically coupled with Pb isotopic analysis. The isotope fingerprinting method utilizes
7 measurements of the abundance of common Pb isotopes (i.e., 204Pb, 206Pb, 207Pb, 208Pb) to
8 distinguish between natural Pb over geologic time and potential anthropogenic sources. Details
9 of this method were described in Section AX8.1.2. The concentration of isotope 204Pb has
10 remained constant throughout time, while the other isotope species can be linked to various
11 anthropogenic Pb sources. Typically, the ratios or signatures of isotopes (e.g., 207Pb:206Pb) are
12 compared between environmental samples to indicate similarities or differences in the site being
13 investigated and the potential known sources.
14 Generally, Pb concentrations in sediment vary with depth. For example, Chow et al.
15 (1973) examined sediment Pb profiles in southern California. Lead concentrations were
16 increased in the shallower sediment depths and comparatively decreased at greater depths. These
17 changes in sediment vertical concentration were attributed to higher anthropogenic Pb fluxes
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1 from municipal sewage, storm runoff, and atmospheric deposition. Similar experiments
2 conducted throughout the United States have also suggested an increase in Pb concentrations in
3 the upper sediment layer concomitant with increases in anthropogenic inputs (Bloom and
4 Crecelius, 1987; Case et al., 1989; Ritson et al., 1999; Chillrud et al., 2003).
5 Sediment Pb concentration profiles and isotope analysis have also been used to identify
6 specific anthropogenic sources. For example, Flegal et al. (1987) used isotopic ratios to trace
7 sources of Pb in mussels from Monterey Bay, CA to a specific slag deposit. Several
8 investigators have examined isotopic tracers to determine potential regional sources of Pb in
9 eastern North America and the Great Lakes (Flegal et al., 1989b; Graney et al., 1995; Blais,
10 1996). Water samples from Lake Erie and Lake Ontario were collected and analyzed. Lead
11 isotope ratios (206Pb:207Pb) from the lakes were compared to known ratios for Pb aerosols derived
12 from industrial sources in Canada and the United States and found to correlate positively. This
13 indicated that a majority of Pb in the lakes was derived from those industrial sources (Flegal
14 et al., 1989b). Lead isotopes in sediment cores from Quebec and Ontario, Canada were also used
15 to distinguish between the amount of Pb deposited from local Canadian sources (28.4 to 61.7%)
16 and U.S. sources (38.3 to 71.6%) (Blais, 1996). Examination of Pb isotopes in sediment and
17 suspended sediment in the St. Lawrence River were used to identify potential anthropogenic Pb
18 sources from Canada (Gobeil et al., 1995, 2005). Graney et al. (1995) used Pb isotope
19 measurements to describe the differing historic sources of Pb in Lake Erie, Ontario and in
20 Michigan. Temporal changes in Pb isotopic ratios were found to correspond to sources such as
21 regional deforestation from 1860 through 1890, coal combustion and or smelting through 1930,
22 and the influence of leaded gasoline consumption from 1930 to 1980.
23 The historic record of atmospheric Pb pollution has been studied to understand the natural
24 background Pb concentration and the effects of Pb accumulation on ecosystems (Bindler et al.,
25 1999; Renberg et al., 2000, 2002; Brannvall et al., 2001a,b). The most extensive work in this
26 area has been conducted at pristine locations in Sweden (Bindler et al., 1999). In this study, soil,
27 sediment, and tree rings were sampled for Pb concentrations and isotopic analyses were
28 conducted on the soil samples. From this record, historic Pb concentrations and Pb accumulation
29 rates were estimated. Present day concentrations in the forest soils ranged from 40 to 100 mg/kg,
30 while a natural background concentration was estimated at <1 mg/kg. The authors were able to
31 model Pb concentrations for the past 6,000 years and also to project Pb concentrations for the
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1 next 400 years, given an assumed atmospheric deposition rate of 1 mg Pb nT2/year. Models
2 such as this are useful tools in determining the critical limits of metals in soils or sediments
3 (Bindler et al., 1999; Renberg et al., 2002).
4 Lead source association may also be assessed through retrospective measurements.
5 Squire et al. (2002) used a time-series approach to evaluate the change in Pb in San Francisco
6 Bay, CA from 1989 to 1999. This approach involved the use of detailed linear regression models
7 and long-term monitoring data to determine changes in Pb concentrations and to identify events
8 corresponding to those changes. Sediment and water samples were collected throughout the bay
9 and combined with data on effluent discharges, urban runoff, atmospheric deposition, and river
10 discharges. The authors identified a 40% decline of Pb in the southern portion of the bay but
11 found no change in the northern reach. The decline was attributed to a reduction in wastewater
12 source loadings over the previous decade.
13
14 AX8.2.2.4 Summary
15 Lead is widely distributed in aquatic ecosystems, predominantly originating from
16 atmospheric deposition or point source contribution. The fate and behavior of Pb in aquatic
17 systems is regulated by physical and chemical factors such as pH, salinity, sediment sorption,
18 transformation, and uptake by aquatic biota. In the United States, Pb concentrations in surface
19 waters, sediments, and fish tissues range from 0.04 to 30 |ig/L, 0.5 to 12,000 mg/kg, and 0.08 to
20 23 mg/kg, respectively. Atmospheric sources are generally decreasing, as the United States has
21 removed Pb from gasoline and other products. However, elevated Pb concentrations remain at
22 sites associated with mining wastes or wastewater effluents. Since the 1986 Pb AQCD, much
23 has been learned about the processes affecting Pb fate and transport. Detailed analyses are
24 currently available (i.e., Pb isotope dating) to allow for constructing the history of Pb
25 accumulation and identifying specific Pb contaminant sources. Continued source control along
26 with examination of the physical and chemical properties will further allow for the reduction of
27 Pb concentrations throughout the United States.
28
29 AX8.2.3 Aquatic Species Response/Mode of Action
30 Recent advancements in understanding the responses of aquatic biota to Pb exposure are
31 highlighted in this section. A summary of the conclusions on the review of aquatic responses to
32 Pb from the appropriate sections of the 1986 Pb AQCD, Volume II (U.S. Environmental
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1 Protection Agency, 1986a) and the subsequent conclusions and recommendations contained in
2 the EPA staff review of that document (U.S. Environmental Protection Agency, 1990) are also
3 provided. In addition, this section summarizes research subsequent to the 1986 Pb AQCD on Pb
4 uptake into aquatic biota, effects of Pb speciation on uptake, resistance mechanisms to Pb
5 toxicity, physiological effects of Pb, factors that affect responses to Pb, and factors associated
6 with global climate change. Areas of research that are not addressed here include literature
7 related to exposure to Pb shot or pellets and studies that examine human health-related endpoints
8 (e.g., hypertension), which are described in other sections of this document.
9
10 AX8.2.3.1 Lead Uptake
11 Lead is nutritionally nonessential and non-beneficial and is toxic to living organisms in all
12 of its forms (Eisler, 2000). Lead can bioaccumulate in the tissues of aquatic organisms through
13 ingestion of food and water and adsorption from water (Vazquez et al., 1999; Vink, 2002) and
14 subsequently lead to adverse effects (see Section AX8.2.5). Recent research has suggested that
15 due to the low solubility of Pb in water, dietary Pb (i.e., lead adsorbed to sediment, particulate
16 matter, and food) may contribute substantially to exposure and toxicity in aquatic biota (Besser
17 et al., 2005). Besser et al. (2004) exposed the amphipod Hyalella azteca to concentrations of Pb
18 to evaluate the influence of waterborne and dietary Pb exposure on acute and chronic toxicity.
19 The authors found that acute toxicity was unaffected by dietary exposure but that dietary Pb
20 exposure did contribute to chronic toxic effects (i.e., survival, growth, reproduction) in H. azteca.
21 Field studies in areas affected by metal contamination (i.e., Clark Fork River, MO; Coeur
22 d'Alene, ID) (Woodward et al., 1994, 1995; Farag et al., 1994) have also demonstrated the
23 effects of dietary metals on rainbow trout. However, there has been a debate on the importance
24 of dietary exposure, as few controlled laboratory studies have been able to replicate the effects
25 observed in the field studies (Hodson et al., 1978; Mount, 1994; Erikson, 2001). This may be
26 due to differences in the availability of Pb from the dietary sources used in laboratory studies,
27 differences in speciation, and/or nutritional characteristics of the Pb dosed diets. In many field
28 and laboratory studies, dietary exposure is rarily considered, but food provided to biota in these
29 studies adsorb metals from water. Therefore, both dietary and waterborne exposure are
30 occurring and both may be considered to play roles in eliciting the measured effects.
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1 Lead concentrations in the tissues of aquatic organisms are generally higher in algae and
2 benthic organisms and lower in higher trophic-level consumers (Eisler, 2000). Thus, trophic
3 transfer of Pb through food chains is not expected (Eisler, 2000). Metals are not metabolized;
4 therefore, they are good integrative indicators of exposure in aquatic biota (Luoma and Rainbow,
5 2005). Metal uptake is complex, being influenced by geochemistry, route of exposure (diet and
6 adsorption), depuration, and growth (Luoma and Rainbow, 2005). This section discusses the
7 factors affecting uptake of Pb by aquatic biota and the state of current research in this area.
8 As described in Section AX8.2.2.1, the solubility of Pb in water varies with pH,
9 temperature, and ion concentration (water hardness) (Weber, 1993). Lead becomes soluble and
10 bioavailable under conditions of low pH, organic carbon content, suspended sediment
11 concentrations, and ionic concentrations (i.e., low Cd, Ca, Fe, Mn, Zn) (Eisler, 2000). Lead
12 rapidly loses solubility above pH 6.5 (Rickard and Nriagu, 1978) and precipitates out as Pb(OH)+
13 and PbHCC>3+ into bed sediments. However, at reduced pH levels or ionic concentrations,
14 sediment Pb can remobilize and potentially become bioavailable to aquatic organisms (Weber,
15 1993).
16 The most bioavailable inorganic form of Pb is divalent Pb (Pb2+), which tends to be more
17 readily assimilated by organisms than complexed forms (Erten-Unal et al., 1998). On the other
18 hand, the low solubility of Pb salts restricts movement across cell membranes, resulting in less
19 accumulation of Pb in fish in comparison to other metals (e.g., Hg, Cu) (Baatrup, 1991).
20 The accumulation of Pb in aquatic organisms is, therefore, influenced by water pH, with
21 lower pHs favoring bioavailability and accumulation. For example, fish accumulated Pb at a
22 greater rate in acidic lakes (pH = 4.9 to 5.4) than in more neutral lakes (pH = 5.8 to 6.8) (Stripp
23 et al., 1990). Merlini and Pozzi (1977) found that pumpkinseed sunfish exposed to Pb at pH 6.0
24 accumulated three-times as much Pb as fish kept at pH 7.5. However, Albers and Camardese
25 (1993a,b) examined the effects of pH on Pb uptake in aquatic plants and invertebrates in acidic
26 (pH -5.0) and nonacidic (pH -6.5) constructed wetlands, ponds, and small lakes in Maine and
27 Maryland. Their results suggested that low pH had little effect on the accumulation of metals by
28 aquatic plants and insects and on the concentration of metals in the waters of these aquatic
29 systems (Albers and Camardese, 1993a,b).
30 Three geochemical factors that influence metal bioaccumulation in aquatic organisms
31 include speciation, particulate metal form, and metal form in the tissues of prey items (Luoma
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1 and Rainbow, 2005). Lead is typically present in acidic aquatic environments as PbSO/t, PbCU,
2 ionic Pb, cationic forms of Pb-hydroxide, and ordinary hydroxide Pb(OH)2. In alkaline waters,
3 common species of Pb include anionic forms of Pb-carbonate (Pb(CO3)) and Pb(OH)2. Labile
4 forms of Pb (e.g., Pb2+, PbOH+, PbCOs) are a significant portion of the Pb inputs to aquatic
5 systems from atmospheric washout. Particulate-bound forms are more often linked to urban
6 runoff and mining effluents (Eisler, 2000). Little research has been done to link the complex
7 concepts of chemical speciation and bioavailability in natural systems (Vink, 2002). The
8 relationship between the geochemistry of the underlying sediment and the impact of temporal
9 changes (e.g., seasonal temperatures) to metal speciation are particularly not well studied (Vink,
10 2002; Hassler et al., 2004).
11 Generally speaking, aquatic organisms exhibit three Pb accumulation strategies:
12 (1) accumulation of significant Pb concentrations with a low rate of loss, (2) excretion of Pb
13 roughly in balance with availability of metal in the environment, and (3) weak net accumulation
14 due to very low metal uptake rate and no significant excretion (Rainbow, 1996). Species that
15 accumulate nonessential metals such as Pb and that have low rates of loss must partition it
16 internally in such a way that it is sparingly available metabolically. Otherwise, it may cause
17 adverse toxicological effects (Rainbow, 1996). Aquatic organisms that exhibit this type of
18 physiological response have been recommended for use both as environmental indicators of
19 heavy metal pollution (Borgmann et al., 1993; Castro et al., 1996; Carter and Porter, 1997) and,
20 in the case of macrophytes, as phytoremediators, because they accumulate heavy metals rapidly
21 from surface water and sediment (Gavrilenko and Zolotukhina, 1989; Simoes Gon9alves et al.,
22 1991; Carter and Porter, 1997).
23 Uptake experiments with aquatic plants and invertebrates (e.g., macrophytes,
24 chironomids, crayfish) have shown steady increases in Pb uptake with increasing Pb
25 concentration in solution (Knowlton et al., 1983; Timmermans et al., 1992). In crayfish, the
26 process of molting can cause a reduction in body Pb concentrations, as Pb incorporated into the
27 crayfish shell is eliminated (Knowlton et al., 1983). Vazquez et al. (1999) reported on the uptake
28 of Pb from solution to the extracellular and intracellular compartments of 3 species of aquatic
29 bryophytes. Relative to the 6 metals tested, Pb was found to accumulate to the largest degree in
30 the extracellular compartments of all 3 bryophytes. The extracellular metals were defined as
31 those that are incorporated into the cell wall or are found on the outer surface of the plasma
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1 membrane (i.e., adsorbed) (Vazquez et al., 1999). Intracellular metals were defined as metals
2 introduced into the cell through a metabolically controlled process.
3 Arai et al. (2002) examined the effect of growth on the uptake and elimination of trace
4 metals in the abalone Haliotos. They reported that older abalones had generally lower whole
5 body concentrations of heavy metals than did younger, rapidly growing individuals. During the
6 rapid growth of juveniles, the organism is less able to distinguish between essential (e.g., Zn),
7 and nonessential metals (e.g., Pb). Once they reach maturity, they develop the ability to
8 differentiate these metals. Li et al. (2004) reported a similar response in zebra fish embryo-
9 larvae. Li et al. (2004) suggested that mature physiological systems are not developed in the
10 embryo-larvae to handle elevated concentrations of metals. Therefore, metals are transported
11 into the body by facilitated diffusion. Both the zebra fish and juvenile abalone demonstrate a
12 rapid accumulation strategy followed by a low rate of loss as described above. There are
13 insufficient data available to determine whether this phenomenon is true for other aquatic
14 organisms.
15 Growth rates are generally thought to be an important consideration in the comparison of
16 Pb levels in individuals of the same species. The larger the individual the more the metal content
17 is diluted by body tissue (Rainbow, 1996).
18 Once Pb is absorbed, it may sequester into varying parts of the organism. Calcium
19 appears to have an important influence on Pb transfer. For example, Pb uptake and retention in
20 the skin and skeleton of coho salmon was reduced when dietary Ca was increased (Varanasi and
21 Gmur, 1978). Organic Pb compounds tend to accumulate in lipids, and are taken up and
22 accumulated in fish more readily than inorganic Pb compounds (Pattee and Pain, 2003).
23 Given the complexities of metal uptake in natural systems, a model incorporating some of
24 the factors mentioned above is desirable. The EPA's Environmental Research Laboratory in
25 Duluth, Minnesota developed a thermodynamic equilibrium model, MINTEQ that predicts
26 aqueous speciation, adsorption, precipitation, and/or dissolution of solids for a defined set of
27 environmental conditions (MacDonald et al., 2002; Playle, 2004). Although not specifically
28 designed to model uptake, MINTEQ provides an indication of what forms of the metal are likely
29 to be encountered by aquatic organisms by estimating the formation of metal ions, complexation
30 of metals, and the general bioavailability of metals from environmental parameters. More
31 recently, a mechanistic model centered on biodynamics has been proposed by Luoma and
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1 Rainbow (2005) as a method of tying together geochemical influences, biological differences,
2 and differences among metals to model metal bioaccumulation. The biodynamic model would
3 be useful in determining the potential adverse effects on aquatic biota, which species are most
4 useful as indicators of metal effects, and how ecosystems may change when contaminated by
5 metals.
6 Two prominent models examine trace metal bioavailability and its link to effects (Hassler
7 et al., 2004). These include the free ion activity model (FIAM) and the biotic ligand model
8 (BLM). Specific information on these models is provided in Section AX8.2.1. Generally, FIAM
9 explores the activity of free ions in solution. The FIAM has been used to examine cationic
10 binding to sensitive sites in algae and takes into account dissolved organic matter in
11 complexation reactions (Niyogi and Wood, 2004). The BLM explores the activity of free ions at
12 biologically reactive sites (i.e., fish gill tissue). Both of these models can increase our awareness
13 of the processes governing the movement of Pb into aquatic biota. They provide insight into the
14 speciation of Pb under certain environmental conditions (e.g., pH, DOC, hardness) and are
15 important in helping understand how Pb and other metals move, accumulate, and cause effects in
16 aquatic organisms. To date, there has been no BLM model of Pb, although research has been
17 conducted on a Pb-gill binding model for rainbow trout (MacDonald et al., 2002; Niyogi and
18 Wood, 2003, 2004). Both the BLM and FIAM models have limitations including difficulty with
19 predictions in the presence of competing ions (e.g., Ca2+) and other factors (e.g., temperature)
20 that can affect membrane permeability of metals (Hassler et al., 2004).
21
22 Bioconcentration Factors (BCF)
23 BCFs for Pb are reported for various aquatic plants in Table AX8-2.3.1. The green alga
24 Cladophora glomerata is reported as having the highest BCF (Keeney et al., 1976). Duckweed
25 (Lemna minor) exhibited high BCF values ranging from 840 to 3560 depending on the method of
26 measurement (Rahmani and Sternberg, 1999). Duckweed that was either previously exposed or
27 not exposed to Pb was exposed to a single dose of Pb-nitrate at 5000 jig/L for 21 days.
28 Duckweed that was previously exposed to Pb removed 70 to 80% of the Pb from the water, while
29 the previously unexposed duckweed removed 85 to 90%. Both plant groups were effective at
30 removing Pb from the water at sublethal levels.
31
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Table AX8-2.3.1. Bioconcentration Factors for Aquatic Plants
BCF
840 to 2700
(measured digestion)
Species
Duckweed
(Lemna minor)
Test Conditions
21 days, Pb-nitrate
Reference
Rahmani and Sternberg
(1999)
1150 to 3560
(measured solution)
16,000 to 20,000
Duckweed
Alga (Cladophora
glomerata)
21 days, Pb-nitrate Rahmani and Sternberg
(1999)
not specified
Keeneyetal. (1976)
4
5
6
BCFs for Pb are reported for various invertebrates in Table AX8-2.3.2. BCFs for
freshwater snails were 738 for a 28-day exposure (Spehar et al., 1978) and 1,700 for a 120-day
exposure (Borgmann et al., 1978). Other reported values for invertebrates included a BCF of
1930 for the scud during a 4-day exposure (MacLean et al., 1996), and BCFs of 499 and 1120 for
the caddis fly and stonefly, respectively, in 28-day exposures (Spehar et al., 1978). In a 28-day
exposure, midge larvae were reported with a BCF of 3670 (Timmermans et al., 1992).
BCF
Table AX8-2.3.2. Bioconcentration Factors for Aquatic Invertebrates
Species
Test Conditions
Reference
738
1700
499
1120
1930
3670
Snail (Physa Integra)
Snail (Lymnaea palustrls)
Caddis fly (Brachycentrus sp.)
Stonefly (Pteronarcys dorsata)
Scud (Hyalella azteca)
28 days, Pb-nitrate Spehar et al. (1978)
120 days, Pb-nitrate Borgmann et al. (1978)
28 days, Pb-nitrate Spehar et al. (1978)
28 days, Pb-nitrate Spehar et al. (1978)
4 days, Pb-chloride MacLean et al. (1996)
Midge larvae (Chironomus rlpanus) 28 days
Timmermans et al.
(1992)
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1 BCFs for freshwater fish were 42 and 45 for brook trout and bluegill, respectively
2 (Holcombe et al., 1976; Atchison et al., 1977). Although no BCFs have been reported for
3 amphibians, Pb-nitrate was reported to accumulate mainly in the ventral skin and in the kidneys
4 of frogs (Vogiatzis and Loumbourdis, 1999).
5 Bioconcentration factors and bioaccumulation factors (BAFs) are not necessarily the best
6 predictors of tissue concentration levels given exposure concentration levels (Kapustka et al.,
7 2004). The role of homeostatic mechanisms is a major consideration in tissue concentrations
8 found in exposed biota. Similarly, measuring BCFs and BAFs in organisms may not accurately
9 reflect how metals are treated within the organisms (e.g., partitioning to specific organelles,
10 sequestering to organ tissues). Therefore, they are not recommended for use in conducting metal
11 risk assessments (Kapustka et al., 2004).
12
13 AX8.2.3.2 Resistance Mechanisms
14 Detoxification Mechanisms
15 Detoxification includes the biological processes by which the toxic qualities, or the
16 probability and/or severity of harmful effects, of a poison or toxin are reduced by the organism.
17 In the case of heavy metals, this process frequently involves the sequestration of the metal,
18 rendering it metabolically inactive. Recent research into heavy metal detoxification in aquatic
19 biota has focused on several physiological and biochemical mechanisms for detoxifying Pb.
20 This section examines these mechanisms and the ability of plants, protists, invertebrates, and fish
21 to mitigate Pb toxicity.
22
23 Plants and Protists
24 Deng et al. (2004) studied the uptake and translocation of Pb in wetland plant species
25 surviving in contaminated sites. They found that all plants tended to sequester significantly
26 larger amounts of Pb in their roots than in their shoots. Deng et al. (2004) calculated a
27 translocation factor (TF), the amount of Pb found in the shoots divided by the amount of Pb
28 found in the root system, and found that TFs ranged from 0.02 to 0.80. Concentrations of Pb in
29 shoots were maintained at low levels and varied within a narrow range. Deng et al. (2004)
30 observed that plants grown in Pb-contaminated sites usually contained higher concentrations
31 than the 27 mg/kg toxicity threshold established for plants by Beckett and Davis (1977). Some
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1 of the wetland plants examined by Deng et al. (2004) also accumulated high concentrations of
2 metals in shoot tissues; however, these metals were assumed to be detoxified (metabolically
3 unavailable), as no toxic response to these elevated concentrations was observed. Deng et al.
4 (2004) suggested that this ability is likely related to discrete internal metal detoxification
5 tolerance mechanisms.
6 Phytochelatins are thiol-containing intracellular metal-binding polypeptides that are
7 produced by plants and protists in response to excessive uptake of heavy metals (Zenk, 1996).
8 Phytochelatins are synthesized by the enzyme phytochelatin synthase that is activated by the
9 presence of metal ions and uses glutathione as a substrate. When phytochelatins are synthesized
10 in sufficient amounts to chelate the metal ion, the enzyme is deactivated (Morelli and Scarano,
11 2001).
12 Morelli and Scarano (2001) studied phytochelatin synthesis and stability in the marine
13 diatom Phaeodactylum tricornutum in the presence of Pb. They found that when metal exposure
14 was alleviated, significant cellular Pb-phytochelatin complex content was lost. Their findings
15 support a hypothesis of vacuolarization proposed for higher plants (Zenk, 1996), in which metal -
16 phytochelatin complexes are actively transported from the cytosol to the vacuole, where they
17 undergo rapid turnover. Zenk (1996) suggested that the complex dissociates, and the metal-free
18 peptide is subsequently degraded. Morelli and Scarano (2001) proposed concomitant occurrence
19 of phytochelatin synthesis and release during metal exposure, as a coincident detoxification
20 mechanism in P. tricornutum.
21
22 Aquatic Invertebrates
23 Like plants and protists, aquatic animals detoxify Pb by preventing it from being
24 metabolically available, though their mechanisms for doing so vary. Invertebrates use
25 lysosomal-vacuolar systems to sequester and process Pb within glandular cells (Giamberini and
26 Pihan, 1996). They also accumulate Pb as deposits on and within skeletal tissue (Knowlton
27 et al., 1983; Anderson et al., 1997; Boisson et al., 2002), and some can efficiently excrete Pb
28 (Vogt and Quinitio, 1994; Prasuna et al., 1996).
29 Boisson et al. (2002) used radiotracers to evaluate the transfer of Pb into the food pathway
30 of the starfish Asterias rubens as well as its distribution and retention in various body
31 compartments. Boisson et al. (2002) monitored Pb elimination after a single feeding of Pb-
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1 contaminated molluscs and found that Pb was sequestered and retained in the skeleton of the
2 starfish, preventing it from being metabolically available in other tissues. Elimination (as
3 percent retention in the skeleton) was found to follow an exponential time course. Elimination
4 was rapid at first, but slowed after 1 week, and eventually stabilized, implying an infinite
5 biological half-life for firmly bound Pb. Results of radiotracer tracking suggest that Pb migrates
6 within the body wall from the organic matrix to the calcified skeleton. From there, the metal is
7 either absorbed directly or adsorbed on newly produced ossicles (small calcareous skeletal
8 structures), where it is efficiently retained as mineral deposition and is not metabolically active
9 (Boisson et al., 2002).
10 AbdAllah and Moustafa (2002) studied the Pb storage capability of organs in the marine
11 snail Nerita saxtilis. Enlarged electron-dense vesicles and many granules were observed in
12 digestive cells of these snails and are suggested to be the site of storage of detoxified metals.
13 N. saxtilis were found to be capable of concentrating Pb up to 50 times that of surrounding
14 marine water without exhibiting signs of histopathologic changes. This ability has been
15 attributed to chelation with various biochemical compounds, such as thionine (forming
16 metallothionine) (Rainbow, 1996), or complexation with carbonate, forming lipofuchsin
17 (AbdAllah and Moustafa, 2002). Granules observed in lysosomal residual bodies were presumed
18 to be the result of Pb accumulation. The presence of large vacuoles and residual bodies were
19 indicative of the fragmentation phase of digestion, suggesting that Pb was also processed
20 chemically in the digestive cells.
21 The podocyte cells of the pericardial gland of bivalves are involved in the ultrafiltration of
22 the hemolymph (Giamberini and Pihan, 1996). A microanalytical study of the podocytes in
23 Dreissenapolymorpha exposed to Pb revealed lysosomal-vacuolar storage/processing similar to
24 that in the digestive cells of Nerita saxtilis. The lysosome is thought to be the target organelle
25 for trace metal accumulation in various organs of bivalves (Giamberini and Pihan, 1996).
26 Epithelial secretion is the principal detoxification mechanism of the tiger prawn Penaeus
27 monodon. Vogt and Quinitio (1994) found that Pb granules tended to accumulate in the
28 epithelial cells of the antennal gland (the organ of excretion) of juveniles exposed for 5 and 10
29 days to waterborne Pb. The metal is deposited in vacuoles belonging to the lysosomal system.
30 Continued deposition leads to the formation of electron-dense granules. Mature granules are
31 released from the cells by apocrine secretion into the lumen of the gland, and presumably
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1 excreted through the nephridopore (i.e., the opening of the antennal gland). Apocrine secretion
2 is predominant, so that as granules form, they are kept at low levels. Excretion was also found to
3 be a primary and efficient detoxification mechanism in the shrimp Chrissia halyi (Prasuna et al.,
4 1996).
5 Crayfish exposed to Pb have been shown to concentrate the metal in their exoskeleton and
6 exuvia through adsorption processes. More than 80% of Pb found in exposed crayfish has been
7 found in exoskeletons (Knowlton et al., 1983; Anderson et al., 1997). Following exposure,
8 clearance is most dramatic from the exoskeleton. The result of a 3-week Pb-clearance study with
9 red swamp crayfish Procambarus clarkia, following a 7-week exposure to 150 jig Pb/L, showed
10 an 87% clearance from the exoskeleton due, in part, to molting. Other organs or tissues that take
11 up significant amounts of Pb include the gills, hepatopancreas, muscle, and hemolymph, in
12 decreasing order. These parts cleared >50% of accumulated Pb over the 3-week clearance
13 period, with the exception of the hepatopancreas. The hepatopancreas is the organ of metal
14 storage and detoxification, although the molecular mechanisms of metal balance in crayfish have
15 yet to be extensively investigated (Anderson et al., 1997).
16
17 Fish
18 Most fish use mucus as a first line of defense against heavy metals (Coello and Khan,
19 1996). In fish, some epithelia are covered with extracellular mucus secreted from specialized
20 cells. Mucus contains glycoproteins, and composition varies among species. Mucosal
21 glycoproteins chelate Pb, and settle, removing the metal from the water column. Fish may
22 secrete large amounts of mucus when they come into contact with potential chemical and
23 biochemical threats. Coello and Khan (1996) investigated the role of externally added fish
24 mucus and scales in accumulating Pb from water, and the relationship of these with the toxicity
25 of Pb in fmgerlings of green sunfish, goldfish and largemouth bass. The authors compared trials
26 in which fish scales from black sea bass (Centropristis striatd) and flounder (Pseudopleuronectes
27 americanus) and mucus from largemouth bass were added to green sunfish, goldfish, and
28 largemouth bass test systems and to reference test systems. On exposure to Pb, fish immediately
29 started secreting mucus from epidermal cells in various parts of the body. Metallic Pb stimulated
30 filamentous secretion, mostly from the ventrolateral areas of the gills, while Pb-nitrate stimulated
31 diffuse molecular mucus secretion from all over the body. The addition of largemouth bass
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1 mucus significantly increased the LTso (the time to kill 50%) for green sunfish and goldfish
2 exposed to 250 mg/L of Pb-nitrate. In contrast, Tao et al. (2000) found that mucus reduced the
3 overall bioavailability of Pb to fish but that the reduction was insignificant. Coello and Khan
4 (1996) found that scales were more significant in reducing LTso than mucous. Fish scales can
5 accumulate high concentrations of metals, including Pb, through chelation with keratin. Scales
6 were shown to buffer the pH of Pb-nitrate in solution and remove Pb from water after which they
7 settled out of the water column. Addition of scales to test water made all species (green sunfish,
8 goldfish, and largemouth bass) more tolerant of Pb.
9
10 Summary of Detoxi fiction Processes
11 Mechanisms of detoxification vary among aquatic biota and include processes such as
12 translocation, excretion, chelation, adsorption, vacuolar storage, and deposition. Lead
13 detoxification has not been studied extensively in aquatic organisms, but existing results indicate
14 the following:
15 • Protists and plants produce intracellular polypeptides that form complexes with Pb (Zenk,
16 1996; Morelli and Scarano, 2001).
17 • Macrophytes and wetland plants that thrive in Pb-contaminated regions have developed
18 translocation strategies for tolerance and detoxification (Knowlton et al., 1983; Deng et al.,
19 2004).
20 • Some starfish (asteroids) sequester the metal via mineral deposition into the exoskeleton
21 (Boisson et al., 2002).
22 • Species of mollusc employ lysosomal-vacuolar systems that store and chemically process Pb
23 in the cells of their digestive and pericardial glands (Giamberini and Pihan, 1996; AbdAllah
24 and Moustafa, 2002).
25 • Decapods can efficiently excrete Pb (Vogt and Quinitio, 1994; Giamberini and Pihan, 1996)
26 and sequester metal through adsorption to the exoskeleton (Knowlton et al., 1983).
27 • Fish scales and mucous chelate Pb in the water column, and potentially reduce visceral
28 exposure.
29
30 Avoidance Response
31 Avoidance is the evasion of a perceived threat. Recent research into heavy metal
32 avoidance in aquatic organisms has looked at dose-response relationships as well as the effects of
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1 coincident environmental factors. Preference/avoidance response to Pb has not been extensively
2 studied in aquatic organisms. In particular, data for aquatic invertebrates is lacking.
3 Using recent literature, this section examines preference-avoidance responses of
4 invertebrates and fish to Pb and some other environmental gradients.
5
6 Aquatic Invertebrates
1 Only one study was identified on avoidance response in aquatic invertebrates. Lefcort
8 et al. (2004) studied the avoidance behavior of the aquatic pulmonate snail Physella columbiana
9 from a pond that had been polluted with heavy metals for over 120 years. In a Y-maze test, first
10 generation P. columbiana from the contaminated site avoided Pb at 9283 |ig/L (p < 0.05) and
11 moved toward Pb at 6255 |ig/L (p < 0.05). It is thought that attraction to Pb at certain elevated
12 concentrations is related to Pb neuron-stimulating properties (Lefcort et al., 2004). These results
13 are consistent with those from similar studies. Control snails from reference sites, and first and
14 second-generation snails from contaminated sites were capable of detecting and avoiding heavy
15 metals, although the first generation was better than the second generation, and the second was
16 better than the controls at doing so. This suggests that detection and avoidance of Pb is both
17 genetic and environmentally based for P. columbiana. Lefcort et al. (2004) observed heightened
18 sensitivity to, and avoidance of, heavy metals by the snails when metals where present in
19 combination.
20
21 Aquatic Vertebrates
22 Steele et al. (1989) studied the preference-avoidance response of bullfrog (Rana
23 catesbeiana) to plumes of Pb-contaminated water following 144-h exposure to 0 to 1000 jig
24 Pb/L. In this laboratory experiment, tadpoles were exposed to an influx of 1000 jig Pb/L at five
25 different infusion rates (i.e., volumes per unit time into the test system). Experiments were
26 videotaped and location data from the tank were used to assess response. No significant
27 differences were seen in preference-avoidance responses to Pb in either nonexposed or
28 previously exposed animals. In a similar subsequent study, Steele et al. (1991) studied
29 preference-avoidance response to Pb in American toad (Bufo americanus) using the same
30 exposure range (0 to 1000 jig Pb/L). B. americanus did not significantly avoid Pb, and
31 behavioral stress responses were not observed. The results do not indicate whether the tadpoles
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1 were capable of perceiving the contaminant. Lack of avoidance may indicate insufficient
2 perception or the lack of physiological stress (Steele et al., 1991).
3 The olfactory system in fish is involved in their forming avoidance response to heavy
4 metals (Brown et al., 1982; Svecevicius, 1991). It is generally thought that behavioral avoidance
5 of contaminants may be a cause of reduced fish populations in some water bodies, because of
6 disturbances in migration and distribution patterns (Svecevicius, 2001). Unfortunately,
7 avoidance of Pb by fish has not been studied as extensively as for other heavy metals
8 (Woodward etal., 1995).
9 Woodward et al. (1995) studied metal mixture avoidance response in brown trout
10 (Salmo trutta), as well as the added effects of acidification. A 1-fold (Ix) mixture contained 1.1
11 Hg/L Cd, 12 |ig/L Cu, 55 |ig/L Zn, and 3.2 |ig/L Pb (all metals were in the form of chlorides).
12 Avoidance was quantified as time spent in test water, trip time to test water, and number of trips.
13 Brown trout avoided the Ix mixture as well as the 0.5x, 2x, 4x, and lOx mixtures, but not the
14 O.lx mixture. Reduced avoidance was observed at higher concentrations (4x and lOx). The
15 authors proposed that the reduced avoidance response was due to impaired perception due to
16 injury. These responses are typical of other fish species to individual metals of similar
17 concentrations (Woodward et al., 1995). This study does not conclusively indicate which of the
18 metals in the mixture may be causing the avoidance response. However, given the neurotoxic
19 effects of Pb, impaired perception is a likely response of Pb-exposed fish.
20 When test water was reduced in pH from 8 to 7, 6 to 5, brown trout avoidance increased,
21 but with no significant difference between metal treatments and controls. However, in the Ix
22 metal mixture treatment, brown trout made fewer trips into the test water chamber at the lower
23 pHs (Woodward et al., 1995). This response may be related to an increased abundance of Pb
24 cations at lower pH values in the test system.
25 Scherer and McNicol (1998) investigated the preference-avoidance responses of lake
26 whitefish (Coregonus clupeaformis) to overlapping gradients of light and Pb. Whitefish were
27 found to prefer shade in untreated water. Lead concentrations under illumination ranged from 0
28 to 1000 |ig/L, and from 0 to 54,000 |ig/L in the shade. Under uniform illumination, Pb was
29 avoided at concentrations above 10 |ig/L, but avoidance behavior lacked a dose-dependent
30 increase over concentrations ranging from 10 to 1000 jig Pb/L. Avoidance in shaded areas was
31 strongly suppressed, and whitefish only avoided Pb at concentrations at or above 32,000 |ig/L.
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1 Summary of Avoidance Response
2 In summary, of those aquatic organisms studied, some are quite adept at avoiding Pb in
3 aquatic systems, while others seem incapable of detecting its presence. Snails have been shown
4 to be sensitive to Pb and to avoid it at high concentrations. Conversely, anuran (frog and toad)
5 species lack an avoidance response to the metal. Fish avoidance of many chemical toxicants has
6 been well established, and it is a dominant sublethal response in polluted waters (Svecevicius,
7 2001). However, no studies have been located specifically examining avoidance behavior for Pb
8 in fish. Environmental gradients, such as light and pH, can alter preference-avoidance responses.
9
10 AX8.2.3.3 Physiological Effects of Lead
11 This section presents a review of the physiological effects and functional growth
12 responses associated with the exposure of aquatic biota to Pb. Physiological effects of Pb on
13 aquatic biota can occur at the biochemical, cellular, and tissue levels of organization and include
14 inhibition of heme formation, adverse effects to blood chemistry, and decreases in enzyme
15 levels. Functional growth responses resulting from Pb exposure include changes in growth
16 patterns, gill binding affinities, and absorption rates.
17
18 Biochemical Effects
19 Lead was observed to have a gender-selective effect on brain endocannabinoid (eCB)
20 (e.g., 2-arachidonylglycerol [2-AG] and jV-arachidonylethanolamine [AEA]) levels in fathead
21 minnow Pimephalespromelas (Rademacher et al., 2005). Cannabinoids, such as eCB, influence
22 locomotor activity in organisms. Increased levels of cannabinoids have been shown to stimulate
23 locomotor activity and decreased levels slow locomotor activity (Safiudo-Pefia et al., 2000).
24 Male and female fathead minnows were exposed to 0 and 1000 |ig/L of Pb. Female minnows in
25 the control group contained significantly higher levels of AEA and 2-AG compared to males. At
26 a concentration of 1000 jig Pb/L, this pattern reversed, with males showing significantly higher
27 levels of AEA in the brain than females (Rademacher et al., 2005). After 14-days exposure to
28 the 1000 jig Pb/L treatment, significantly higher levels of 2-AG were found in male fathead
29 minnows, but no effect on 2-AG levels in females was observed (Rademacher et al., 2005).
30 Lead acetate slightly inhibited 7-ethoxyresorufin-o-deethylase (7-EROD) activity in
31 Gammaruspulex exposed for up to 96 h to a single toxicant concentration (ECso) (Kutlu and
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1 Susuz, 2004). The exact concentration used in the study was not reported. The EROD enzyme
2 is required to catalyze the conjugation and detoxification of toxic molecules and has been
3 proposed as a biomarker for contaminant exposure. The authors believe more detailed studies
4 are required to confirm EROD as a biomarker for Pb exposure. The enzyme group alanine
5 transferases (ALT) has been suggested as a bioindicator/biomarker of Pb stress (Blasco and
6 Puppo, 1999). A negative correlation was observed between Pb accumulation and ALT
7 concentrations in the gills and soft body of Ruditapesphilippinarum exposed to 350 to 700 |ig/L
8 of Pb for 7 days (Blasco and Puppo, 1999).
9 Studies have identified ALAD in fish and amphibians as a useful indicator of Pb exposure
10 (Gill et al., 1991; Nakagawa et al., 1995a,b). ALAD catalyzes the formation of hemoglobin and
11 early steps in the synthesis of protoporphyrin (Gill et al., 1991; Nakagawa et al., 1995b). The
12 absence of an inhibitory effect on this enzyme following exposure to cadmium, copper, zinc, and
13 mercury suggests that this enzyme reacts specifically to Pb (Johansson-Sjobeck and Larsson,
14 1979; Gill et al., 1991). A 0% decrease in ALAD activity was reported in common carp
15 (Cyprinus carpio) exposed to a Pb concentration of 10 jig/L for 20 days (Nakagawa et al.,
16 1995b). The recovery of ALAD activity after exposure to Pb has also been examined in carp
17 (Nakagawa et al., 1995a). After 2-week exposure to 200 jig Pb/L, ALAD activity decreased to
18 approximately 25% of value reported for controls (Nakagawa et al., 1995a). Fish removed from
19 the test concentration after 2 weeks and placed in a Pb-free environment recovered slightly, but
20 ALAD activity was only 50% of the controls even after 4 weeks (Nakagawa et al., 1995a).
21 Vogiatzis and Loumbourdis (1999) exposed the frog (Rana ridibundd) to a Pb concentration of
22 14,000 |ig/L over 30 days and a 90% decrease in ALAD activity was observed in the frogs.
23
24 Blood Chemistry
25 Numerous studies have examined the effects of Pb exposure on blood chemistry in aquatic
26 biota. These studies have primarily used fish in acute and chronic exposures to Pb
27 concentrations ranging from 100 to 10,000 |ig/L. Decreased erythrocyte, hemoglobin, and
28 hemocrit levels were observed in rosy barb (Barbuspuntius) during an 8-week exposure to
29 126 |ig/L of Pb-nitrate (Gill et al., 1991).
30 No difference was found in red blood cell counts and blood hemoglobin in yellow eels
31 (Anguilla anguilla) exposed to 0 and 300 |ig/L of Pb for 30 days (Santos and Hall, 1990). The
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1 number of white blood cells, in the form of lymphocytes, increased in the exposed eels. The
2 authors concluded this demonstrates the lasting action of Pb as a toxicant on the immune system
3 (Santos and Hall, 1990). Significant decreases in red blood cell counts and volume was reported
4 in blue tilapia (Oreochromis aureus) exposed to Pb-chloride at a concentration of 10,000 |ig/L
5 for 1 week (Allen, 1993).
6 Blood components, such as plasma glucose, total plasma protein, and total plasma
7 cholesterol, were unaffected in yellow eels exposed to 300 |ig/L of Pb for 30 days (Santos and
8 Hall, 1990). Effects on plasma chemistry were observed in Oreochromis mossambicus exposed
9 to 0, 18,000, 24,000, and 33,000 |ig/L of Pb (Ruparelia et al., 1989). Significant decreases in
10 plasma glucose (hypoglycemic levels) were reported at concentrations of 24,000 and 33,000 //g
11 Pb/L after 14 and 21 days of exposure, and at 18,000 jig Pb/L after 21 days of exposure
12 (Ruparelia et al., 1989). Plasma cholesterol levels dropped significantly in comparison to
13 controls after 14 days of exposure to 33,000 jig Pb/L and in all test concentrations after 21 days
14 of exposure (Ruparelia et al., 1989). Similarly, concentrations of blood serum protein, albumin,
15 and globulin were identified as bioindicators of Pb stress in carp (Cyprinus carpio) exposed to
16 Pb-nitrates at concentrations of 800 and 8000 |ig Pb/L (Gopal et al., 1997).
17
18 Tissues
19 In fish, the gills serve as an active site for ion uptake. Recent studies have examined the
20 competition between cations for binding sites at the fish gill (e.g., Ca2+, Mg2+, Na+, H+, Pb2+)
21 (MacDonald et al., 2002; Rogers and Wood, 2003, 2004). Studies suggest that Pb2+ is an
22 antagonist of Ca2+ uptake (Rogers and Wood, 2003, 2004). MacDonald et al. (2002) proposed a
23 gill-Pb binding model that assumes Pb2+ has a > 100 times greater affinity for binding sites at the
24 fish gill than other cations. More toxicity studies are required to quantify critical Pb burdens that
25 could be used as indicators of Pb toxicity (Niyogi and Wood, 2003).
26
27 Growth Responses
28 A negative linear relationship was observed in the marine gastropod abalone (Haliotis)
29 between shell length and muscle Pb concentrations (Arai et al., 2002). Abalones were collected
30 from two sites along the Japanese coast. Haliotis discus hannai were collected from along the
31 coast at Onagawa; Haliotis discus were collected from along the coast at Amatsu Kominato. The
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1 authors did not report significant differences between the two sampling sites. From samples
2 collected at Onagawa, Pb concentrations of 0.03 and 0.01 jig/g were associated with abalone
3 shell lengths of 7.7 cm (3 years old) and 12.3 cm (6 years old), respectively. From samples
4 collected at Amatsu Kominato, Pb concentrations of 0.09 and 0.01 jig/g were associated with
5 abalone shell lengths of 3.9 cm (0 years old) and 15.3 cm (8 years old), respectively (Arai et al.,
6 2002). The authors theorized that young abalones, experiencing rapid growth, do not
7 discriminate between the uptake of essential and nonessential metals. However, as abalones
8 grow larger and their rate of growth decreases, they increasingly favor the uptake of essential
9 metals over nonessential metals. This is demonstrated by the relatively consistent concentrations
10 of Cu, Mn, and Zn that were reported for the abalone samples (Arai et al., 2002).
11
12 Other Physiological Effects
13 Increased levels of Pb in water were found to increase fish production of mucus: excess
14 mucus coagulates were observed over the entire body of fishes. Buildup was particularly high
15 around the gills, and in the worst cases, interfered with respiration and resulted in death by
16 anoxia (Aronson, 1971; National Research Council of Canada., 1973).
17
18 AX8.2.3.4 Factors That Modify Organism Response to Lead
19 A great deal of research has been undertaken recently to better understand the factors that
20 modify aquatic organism response to Pb. The driving force behind this research is the
21 development of the BLM approach to AWQC development. A discussion of research on the
22 many factors that can modify aquatic organism response to Pb is provided in this section.
23
24 Organism Age and Size Influence on Lead Uptake and Response
25 It is generally accepted that Pb accumulation in living organisms is controlled, in part, by
26 metabolic rates (Farkas et al., 2003). Metabolic rates are, in-turn, controlled by the physiological
27 conditions of an organism, including such factors as size, age, point in reproductive cycle,
28 nutrition, and overall health. Of these physiological conditions, size and age are the most
29 commonly investigated in relation to heavy metal uptake. This section reviews recent research
30 focusing on relationships between body size, age, and Pb accumulation in aquatic invertebrates
31 and fish.
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1 Invertebrates
2 MacLean et al. (1996) investigated bioaccumulation kinetics and toxicity of Pb in the
3 amphipod Hyalella azteca. Their results indicated that body size did not greatly influence Pb
4 accumulation in H. azteca exposed to 50 or 100 |ig/L of PbCb for 4 days. Canli and Furness
5 (1993) found similar results in the Norway lobster Nephrops norvegicus exposed to 100 |ig/L of
6 Pb(NC>3)2 for 30 days. No significant sex- or size-related differences were found in
7 concentrations of Pb in the tissue. The highest tissue burden was found in the carapaces (42%).
8 Several studies have determined that Pb can bind to the exoskeleton of invertebrates and
9 sometimes dominate the total Pb accumulated (Knowlton et al., 1983). This adsorption of Pb to
10 the outer surface of invertebrates can result in strong negative relationships for whole-body Pb
11 concentration as a function of body mass (i.e., concentrations decrease rapidly with increased
12 body size and then stabilize) (MacLean et al., 1996).
13 Drava et al. (2004) investigated Pb concentrations in the muscle of red shrimp Aristeus
14 antennatus from the northwest Mediterranean. Lead concentrations ranged from 0.04 to
15 0.31 |ig/g dw. No significant relationships between size and Pb concentration in A. antennatus
16 were found, and concentrations were not related to reproductive status.
17 Arai et al. (2002) analyzed abalones (Haliotis) at various life stages from coastal regions
18 of Japan. They investigated growth effects on the uptake and elimination of Pb. Results
19 indicated a significant negative linear relationship between age, shell length and Pb
20 concentrations in muscle tissue. The relationship was consistent despite habitat variations in Pb
21 concentrations between the study sites, suggesting that Pb concentrations changed with growth in
22 the muscle tissue of test specimens and implying that abalone can mitigate Pb exposure as they
23 age.
24
25 Fish
26 Douben (1989) investigated the effects of body size and age on Pb body burden in the
27 stone loach (Noemacheilus barbatulus L.). Fish were caught during two consecutive springs
28 from three Derbyshire rivers. Results indicated that Pb burden increased slightly with age.
29 Similarly, Kock et al. (1996) found that concentrations of Pb in the liver and kidneys of Arctic
30 char (Salvelinus alpinus) taken from oligotrophic alpine lakes were positively correlated with
31 age. It has been suggested that fish are not able to eliminate Pb completely, and that this leads to
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1 a stepwise accumulation from year to year (Kock et al., 1996). In contrast, Farkas et al. (2003)
2 found a negative relationship between Pb concentrations and muscle and gill Pb concentrations
3 in the freshwater fish Abramis brama. Fish were taken from a low-contaminated site and
4 contained between 0.44 and 3.24 jig/g Pb dw. Negative correlations between metal
5 concentration and fish size in low-contaminated waters likely results from variations in feeding
6 rates associated with developmental stages. This hypothesis is consistent with the fact that in
7 low-contaminated waters, feeding is the main route of uptake and feeding rates decrease with
8 development in fish (Farkas et al., 2003).
9 In summary, relationships between age, size, and Pb body burden in aquatic invertebrates
10 and fish are interspecifically variable and depend on many environment-related variables (e.g.,
11 exposure) (Farkas et al., 2003).
12
13 Genetics
14 There are few studies documenting the effects of Pb on organismal and population
15 genetics, although rapid advances in biotechnology have prompted recent research in this area
16 (Beaty et al., 1998). There are two principal effects that sublethal exposure to a contaminant can
17 have on the genetics of an organism and/or population: (1) a contaminant may influence
18 selection by selecting for certain phenotypes that enable populations to better cope with the
19 chemical; or (2) a contaminant can be genotoxic, meaning it can produce alterations in nucleic
20 acids at sublethal exposure concentrations, resulting in changes in hereditary characteristics or
21 DNA inactivation (Shugart, 1995). Laboratory studies have shown that exposure to Pb2+ at
22 10 mg/mL in blood produces chromosomal aberrations (i.e., deviations in the normal structure or
23 number of chromosomes) in some organisms (Cestari et al., 2004). Effects of genotoxicity and
24 toxin-induced selection do not preclude one another, and may act together on exposed
25 populations. This section reviews Pb genotoxicity and the effects of Pb-induced selection in
26 aquatic populations.
27
28 Selection
29 Evidence for genetic selection in the natural environment has been observed in some
30 aquatic populations exposed to metals (Rand et al., 1995; Beaty et al., 1998; Duan et al., 2000;
31 Kim et al., 2003). Because tolerant individuals have a selective advantage over vulnerable
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1 individuals in polluted environments, the frequency of tolerance genes will increase in exposed
2 populations over time (Beaty et al., 1998). Several studies have shown that heavy metals can
3 alter population gene pools in aquatic invertebrates. These changes have resulted in decreased
4 genetic diversity and are thought to be a potential source of population instability (Duan et al.,
5 2000; Kim et al., 2003).
6 Kim et al. (2003) investigated genetic differences and population structuring in the
7 gastropod Littorina brevicula from heavy-metal polluted and unpolluted environments.
8 Organisms from polluted sites contained a mean of 1.76 jig Pb/g, while organisms from
9 unpolluted sites contained 0.33 jig Pb/g. They found significant differences in haplotypes
10 between the test groups and allelic diversity was significantly lower among L. brevicula from
11 polluted regions. In contrast, Yap et al. (2004) performed a similar experiment with the green-
12 lipped mussel Perna viridis; they found that mussels from contaminated sites containing between
13 4 and 10 jig Pb/g, as well as other heavy metals, exhibited a higher percentage of polymorphic
14 loci and excess heterozygosity compared to those from uncontaminated sites. The higher level
15 of genetic diversity was attributed to greater environmental heterogeneity (i.e., variation due to
16 pollution gradients) in contaminated sites (Yap et al., 2004).
17 Duan et al. (2000) investigated amphipod (Hyalella azteca) selective mortality and
18 genetic structure following acute exposure to Pb (5.47 mg/L Pb(NC>2)2) as well as exposure to
19 other heavy metals. They found that genetic differentiation consistently increased among
20 survivors from the original population, supporting the hypothesis that heavy metals, including
21 Pb, have the potential to alter the gene pools of aquatic organisms.
22
23 Genotoxicity
24 Low-level (50 |ig/L) Pb exposure in water over 4 weeks resulted in DNA strand breakage
25 in the freshwater mussel Anodonta grandis (Black et al., 1996), although higher concentrations
26 (up to 5000 |ig/L) did not result in significant breakage by the end of the study period. These
27 results suggest that a threshold effect for DNA damage and repair exists, where DNA repair only
28 occurs once a certain body exposure level has been reached. More recently, Cestari et al. (2004)
29 observed similar results in neotropical fish (Hoplias malabaricus) that were fed Pb-contaminated
30 food over 18, 41, and 64 days. Lead body burdens in H. malabaricus were approximately 21 jig
31 Pb 2+/g. Results indicated that exposure to Pb significantly increased the frequency of
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1 chromosomal aberrations and DNA damage in kidney cell cultures, although when assessed at
2 the end of the longer exposure periods, aberrations were less common.
3
4 Environmental Biological Factors
5 Environmental factors that are biological in origin can alter the availability, uptake and
6 toxicity of Pb to aquatic organisms. These factors can be grouped into living and non-living
7 constituents. For example, living organisms may sequester Pb from the water column, reducing
8 the availability and toxicity of the metal in the water column to other biota, thus reducing
9 potential toxic effects in other organisms. Non-living organic material (e.g., components of
10 sioughed-off scales, mucus, carcasses, and other decomposing, humic material) can similarly
11 combine with Pb from the water column, rendering it unavailable. This section will review the
12 literature on biological environmental factors and their influence on the bioavailability, uptake,
13 and toxicity of Pb.
14 Van Hattum et al. (1996) studied the influence of abiotic variables, including DOC on Pb
15 concentrations in freshwater isopods (Proasellus meridianus and Asellus aquaticus). They found
16 that BCFs were significantly negatively correlated with DOC concentrations. Thus, as DOC
17 concentrations increased, BCFs decreased in P. meridianus and A. aquaticus, indicating that
18 DOC acts to inhibit the availability of Pb to these isopods.
19 Kruatrachue et al. (2002) investigated the combined effects of Pb and humic acid on total
20 chlorophyll content, growth rate, multiplication rate, and Pb uptake of common duckweed.
21 When humic acid was added to the Pb-nitrate test solutions (50, 100, and 200 mg Pb(NO3)2/ L),
22 toxicity of Pb to duckweed was decreased. The addition of humic acid to the Pb-nitrate solution
23 increased the pH. The authors suggested that there was a proton dissociation from the carboxyl
24 group in the humic acid that complexed with Pb, resulting in a decrease in free Pb ions available
25 to the plant.
26 Schwartz et al. (2004) collected natural organic matter (NOM) from several aquatic sites
27 across Canada and investigated the effects of NOM on Pb toxicity in rainbow trout
28 (Oncorhynchus my kiss). They also looked at toxicity effects as they related to the optical
29 properties of the various NOM samples. The results showed that NOM in test water almost
30 always increased LTso and that optically dark NOM tended to decrease Pb toxicity more than did
31 optically light NOM in rainbow trout.
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1 In summary, non-living constituents of biological origin in the environment have been
2 shown to reduce Pb availability and, therefore, toxicity in some aquatic organisms. It is
3 generally thought that this occurs through complexation or chelation processes that take place in
4 the water column.
5
6 Physicochemical Environmental Factors
1 This section reviews the literature on physicochemical environmental factors and their
8 influence on the bioavailability, uptake, and toxicity of Pb in aquatic organisms. These factors
9 are discussed with regard to their influence individually and in combination.
10 Studies generally agree that as pH increases, the toxicity of Pb decreases (Home and
11 Dunson, 1995b; MacDonald et al., 2002). As pH decreases, Pb becomes more soluble and more
12 readily bioavailable to aquatic organisms (Weber, 1993). Significantly lower survival, decreased
13 hatching success, slower development, and increased egg mass and larval mortality were
14 observed in Jefferson salamanders (Ambystomajeffersonianum) and wood frogs (Rana sylvaticd)
15 exposed to Pb at a pH of 4.5 versus a pH of 5.5 (Home and Dunson, 1995b). Contradictory
16 results have been reported for invertebrates. Over a 96-h exposure period, mortality increased
17 with decreasing pH for the bivalve Pisidium casertanum, while pH-independent mortality was
18 reported for gastropods and Crustacea under similar exposure conditions (Mackie, 1989).
19 Cladocerans (C. dubid) and amphipods (H. azteca) were also more sensitive to Pb toxicity at pH
20 6 to 6.5 than at higher pH levels (Schubauer-Berigan et al., 1993). Lead was 100 times more
21 toxic to the amphipod Hyalella azteca at a pH range of 5.0 to 6.0 (Mackie, 1989) than at a pH
22 range of 7.0 to 8.5 (Schubauer-Berigan et al., 1993). Lead was also more toxic to fathead
23 minnows at lower pH levels (Schubauer-Berigan et al., 1993).
24 The influence of pH on Pb accumulation has also been observed in sediments.
25 Accumulation of Pb by the isopod Asellus communis was enhanced at low pH, after a 20-day
26 exposure to Pb-contaminated sediments (Lewis and Mclntosh, 1986). In A aquaticus,
27 temperature increases were found to be more important than increased pH in influencing Pb
28 accumulation (Van Hattum et al., 1996). Increased water temperature was also found to reduce
29 Pb uptake fluxes in green microalga (Chlorella kesslerif) (Hassler et al., 2004). Lead and zinc
30 body concentrations in Asellus sp. were found to vary markedly with seasonal temperature
31 changes, with greater concentrations present in spring and summer (Van Hattum et al., 1996).
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1 Acute and chronic toxicity of Pb increases with decreasing water hardness, as Pb becomes
2 more soluble and bioavailable to aquatic organisms (Home and Dunson, 1995a; Borgmann et al.,
3 2005). There is some evidence that water hardness and pH work together to increase or decrease
4 the toxicity of Pb. Jefferson salamanders exposed to Pb for 28 days at low pH and low water
5 hardness experienced 50% mortality, while exposure to Pb at high pH and high water hardness
6 resulted in 91.7% survival (Home and Dunson, 1995a). Exposure to Pb at high pH and low
7 water hardness or low pH and high water hardness resulted in 75 and 41.7% survival,
8 respectively (Home and Dunson, 1995a). Similar results were reported for Jefferson
9 salamanders during a 7-day exposure and wood frogs during 7- and 28-day exposures (Home
10 and Dunson, 1995c). In some cases, water hardness and pH in the absence of Pb have been
11 shown to affect survival adversely. Mean acute survival of wood frogs and Jefferson
12 salamanders exposed to low pH and low water hardness, in the absence of Pb, was 83.3 and
13 91.7%, respectively. Mean chronic survival of wood frogs and Jefferson salamanders exposed to
14 low pH and low water hardness, in the absence of Pb, was 79.2 and 41.7%, respectively (Home
15 and Dunson, 1995c).
16 High Ca2+ concentrations have been shown to protect against the toxic effects of Pb
17 (Sayer et al., 1989; MacDonald et al., 2002; Hassler et al., 2004; Rogers and Wood, 2004).
18 Calcium affects the permeability and integrity of cell membranes and intracellular contents
19 (Sayer et al., 1989). As Ca2+ concentrations decrease, the passive flux of ions (e.g., Pb) and
20 water increases. At the lowest waterborne Ca2+ concentration (150 |imol/L), Pb accumulation in
21 juvenile rainbow trout (Oncorhynchus mykiss) branchials significantly increased as Pb
22 concentration in water increased (Rogers and Wood, 2004). At higher Ca2+ concentrations, Pb
23 accumulation did not significantly increase with Pb concentration in water. This result
24 demonstrates the protective effects of waterborne Ca2+ and supports the suggestion that the Ca2+
25 component of water hardness determines the toxicity of Pb to fish (Rogers and Wood, 2004).
26 Rogers and Wood (2004) reported that the uptake of Ca2+ and Pb2+ involves competitive
27 inhibition of apical entry at lanthanum-sensitive Ca2+ channels and interference with the function
28 of the ATP-driven baso-lateral Ca2+ pump. High mortality was reported in brown trout (Salmo
29 trutta) fry exposed to Pb at a waterborne Ca2+ concentration of 20 |imol/L, while negligible
30 mortality was reported at the same Pb concentration but at a waterborne Ca2+ concentration of
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1 200 jimol/L (Sayer et al., 1989). Adverse effects to mineral uptake and skeletal development
2 were observed in the latter test group (Sayer et al., 1989).
3 The bioavailability of Pb and other metals that can be simultaneously extracted in
4 sediments may be modified through the role of acid volatile sulfide (AVS) under anoxic
5 conditions (Tessier and Campbell, 1987; Di Toro et al., 1992; Casas and Crecelius, 1994). The
6 term AVS (iron sulfide is an example) refers to the fraction of the sediment that consists of a
7 reactive pool of solid-phase sulfide. This phase is available to bind divalent metals that then
8 become unavailable for uptake by aquatic biota. The models proposed by Di Toro et al. (1992)
9 and Casas and Crecelius (1994) predict that when the molar ratio of simultaneously extractable
10 metals (SEM) to AVS in sediments is less than one, the metals will not be bioavailable due to
11 complexation with available sulfide.
12 Salinity is an important modifying factor to metal toxicity. Verslycke et al. (2003)
13 exposed the estuarine mysid Neomysis integer to individual metals, including Pb, and metal
14 mixtures under changing salinity. At a salinity of 5%, the reported LCso for Pb was 1140 |ig/L
15 (95% CL = 840, 1440 |ig/L). At an increased salinity of 25%o, the toxicity of Pb was
16 substantially reduced (LC50 = 4274 |ig/L [95% CL = 3540, 5710 |ig/L]) (Verslycke et al., 2003).
17 The reduction in toxicity was attributed to increased complexation of Pb2+ with Cl ions.
18
19 Nutritional Factors
20 The relationship between nutrition and Pb toxicity has not been thoroughly investigated in
21 aquatic organisms. In fact, algae species are the only aquatic organisms to have been studied
22 fairly frequently. Although nutrients have been found to have an impact on Pb toxicity, the
23 mechanisms involved are poorly understood. It is unclear whether the relationship between
24 nutrients and toxicity comprises organismal nutrition (the process by which a living organism
25 assimilates food and uses it for growth and for replacement of tissues), or whether nutrients have
26 interacted directly with Pb, inhibiting its metabolic interaction in the organism. This section
27 reviews the little information that has been gathered from studies documenting apparent Pb-
28 nutrition associations in aquatic organisms.
29 Jampani (1988) looked at the impact of various nutrients (i.e., sodium acetate, citric acid,
30 sodium carbonate, nitrogen, and phosphates) on reducing growth inhibition in blue-green algae
31 (Synechococcus aeruginosus) exposed to 200 mg Pb/L. Exposure to this Pb treatment
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1 concentration caused 100% mortality in algae. Results indicated that additional nitrogen,
2 phosphates, and some carbon sources, including sodium acetate, citric acid and sodium
3 carbonate, all protected the algae from Pb toxicity. Algae that had been starved prior to the
4 experiment were found to be significantly more sensitive to Pb exposure. Glucose was the only
5 nutrient tested that did not have a significant impact on Pb toxicity in S. aeruginosus. In a
6 similar study by Rao and Reddy (1985) on Scenedesmus incrassatulus, nitrogen, phosphate and
7 carbon sources (including glucose), all had protective effects, and reduced Pb toxicity at 300 and
8 400 mg Pb/L. Both studies proposed similar hypotheses regarding nutrient-Pb mechanisms that
9 led to reduced toxicity. One hypothesis was that the nutrients were able to reverse toxic effects.
10 The second hypothesis was that the nutrients interacted directly with Pb, in some way
11 sequestering the metal so as to inhibit its metabolic interaction with the organism (Rao and
12 Reddy, 1985; Jampani, 1988).
13 Rai and Raizada (1989) investigated the effects of Pb on nitrate and ammonium uptake as
14 well as carbon dioxide and nitrogen fixation in Nostoc muscorum over a 96-h period. Test
15 specimens were exposed to 10, 20, and 30 mg Pb/L. At 20 mg Pb/L, nitrate uptake was inhibited
16 by 64% after 24 h and by 39% after 96 h. Ammonium uptake was inhibited, and similarly,
17 inhibition decreased from 72% inhibition after 24 h to 26% inhibition after 96 h of exposure.
18 Carbon dioxide fixation and nitrogenase activity followed similar patterns, and results indicated
19 that Pb exposure can affect the uptake of some nutrients in N. muscorum.
20 Adam and Abdel-Basset (1990) studied the effect of Pb on metabolic processes of
21 Scenedesmus obliquus. They found that nitrogenase activity was inhibited by Pb nitrate, but
22 enhanced by Pb-acetate. As photosynthetic products and respiratory substrates, carbohydrate
23 and lipid levels were altered by Pb. Above 30 mg/L of Pb-nitrate, both macronutrients were
24 reduced. However, Pb-acetate was found to increase carbohydrate levels. Results suggest that
25 Pb can have an effect on macronutrients in S. obliquus and that effects may vary depending on
26 the chemical species.
27 Simoes Gon9alves et al. (1991) studied the impact of light, nutrients, air flux, and Pb, in
28 various combinations, on growth inhibition in the green algae Selenastrum capricornutum.
29 Results indicated that at lower Pb concentrations (<0.207 mg/L) and increased nutrient
30 concentrations, algae release more exudates that form inert complexes with Pb anions in the
31 water. This suggests that S. capricornutum can use exudates as a protection and that this
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1 protective mechanism depends on nutrient supply. These results are consistent with those of
2 Capelo et al. (1993), who investigated uptake of nitrogen and phosphorus in the algae
3 Selenastrum capricornutum over time in the absence and presence of 0.207 mg Pb/L. They
4 found that the presence of Pb had no significant influence on the assimilation of nitrogen and
5 phosphorus. However, they did find that in the presence of Pb, algae released higher
6 concentrations of organics with Pb-chelating groups.
7 Amiard et al. (1994) investigated the impact on soft tissue Pb concentrations of various
8 feeding regimes on oysters (Crassostrea gigas) during their spat rearing. They fed test groups of
9 C. gigas different amounts of Skeletonema costatum and additional natural phytoplankton grown
10 in test solutions. Results showed that size and food intake both negatively correlated with metal
11 concentrations in soft tissue. The authors hypothesized that this relationship was due in part to a
12 diluting effect of the food.
13 In summary, nutrients affect Pb toxicity in those aquatic organisms that have been studied.
14 Some nutrients seem to be capable of reducing toxicity, though the mechanisms have not been
15 well established. Exposure to Pb has not been shown to reduce nutrient uptake ability, though it
16 has been demonstrated that Pb exposure may lead to increased production and loss of organic
17 material (e.g., mucus and other complex organic ligands) (Capelo et al., 1993).
18
19 Interactions with Other Pollutants
20 Most of the scientific literature reviewed in this section considered how Pb and other
21 elements combine to affect uptake and exert toxicity. Research on the interactions of Pb with
22 complexing ligands and other physical and biological factors was more thoroughly discussed in
23 Section AX8.2.3.4. Predicting the response of organisms to mixtures of chemicals is difficult
24 (Norwood et al., 2003). For example, at low zinc concentrations, (2:1 Pb:Zn ratio) a synergistic
25 effect was observed in the frog, Bufo arenarum (Herkovits and Perez-Coll, 1991). At high
26 concentrations of zinc, an antagonistic effect was observed as Pb toxicity was reduced. This
27 demonstrates the complexity of metal mixture interactions as different metal concentrations,
28 environmental conditions (e.g., temperature, pH), and other factors can cause marked changes in
29 the effects observed (Norwood et al., 2003). In describing Pb interactions with other elements,
30 interaction types are classified here as antagonistic, synergistic, and additive. Each of these will
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1 be discussed below with specific reference to known Pb-metal interactions and implications on
2 Pb uptake and toxicity.
3
4 Antagonistic Interactions
5 When two or more metals compete for the same binding sites or interfere with transport
6 through cell walls or membranes, the interaction is termed less than strictly additive or
7 antagonistic. Antagonistic interactions can reduce metal bioavailability when metals are present
8 in combination, and may lead to reduced potential for toxicity (Hassler et al., 2004). A number
9 of elements act in an antagonistic fashion with Pb. For example, Pb is a well-known antagonist
10 to Ca2+ (Niyogi and Wood, 2004; Hassler et al., 2004), which is an essential element, required
11 for a number of physiological processes in most organisms. Lead ions have an atomic structure
12 similar to Ca2+ and can be transported either actively or passively across cell membranes in place
13 of Ca2+. An example of this interaction was reported by Behra (1993a,b) where Pb was shown to
14 activate calmodulin reactions in rainbow trout (O. mykiss) and sea mussel (Mytilus sp.) tissues in
15 the absence of calcium. Calmodulin (CaM) is a major intracellular calcium receptor and
16 regulates the activities of numerous enzymes and cellular processes. Allen (1994) reported that
17 Pb can replace calcium in body structures (e.g., bones, shells); replace zinc in ALAD, which is
18 required for heme biosynthesis; and react with sulfhydryl groups, causing conformation protein
19 distortion and scission of nucleic acids (Herkovits and Perez-Coll, 1991). Lead is also a known
20 antagonist to Mg2+, Na+, and Cl~ regulation in fish (Ahern and Morris, 1998; Rogers and Wood,
21 2003, 2004; Niyogi and Wood, 2004). Li et al. (2004) reported on the interaction of Pb2+ with
22 Cd2+ in the context of adsorption from solution by Phanerochaete chrysosporium, a filamentous
23 fungus. The authors found that cadmium uptake decreased with increasing concentration of Pb
24 ions with Pb2+ outcompeting Cd2+ for binding sites.
25
26 Synergistic Interactions
27 Synergism occurs when the interaction of two or more metals causes an effect that is
28 greater than the effect observed from the individual metals themselves (Hagopian-Schlekat et al.,
29 2001) or, put another way, a greater than the strictly additive effect of the individual metals in a
30 mixture (Playle, 2004). Synergism is likely the result of increased bioavailability of one or more
31 of the metal ions due to the presence of other metals (Hassler et al., 2004). Hassler et al. (2004)
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1 reported that in the presence of copper (Cu2+), there was a significantly higher rate of
2 internalization of Pb in the green algae Chlorella kesserii. It was suggested that Cu2+ may have
3 affected organism physiology through the disruption of cell membrane integrity. This would
4 allow increased cation (i.e., Pb2+) permeability and, therefore, substantially increase
5 internalization of Pb. Hagopian-Schlekat et al. (2001) examined the impact of individual metals
6 and complex metal mixtures containing Cd, Cu, Ni, Zn, and Pb to the estuarine copepod
7 Amphiascus tenuiremis. The copepods were exposed to metal-spiked sediment and pore water.
8 The mixed metal sediment toxicity tests demonstrated greater than additive toxicity to
9 A. tenuiremis. It was postulated that the synergism observed was due to two or more metals
10 affecting the same biological function. Herkovits and Perez-Coll (1991) exposed Bufo arenarum
11 larvae to various Pb and zinc concentrations in solution. At low zinc concentrations, (2:1 Pb:Zn
12 ratio), a synergistic toxic effect was observed in the frog larvae relative to the effects observed
13 from exposure to the individual metals and at higher zinc concentrations. Enhanced Pb toxicity
14 was attributed to the interference of Pb with cellular activities due to binding with sulfhydryl
15 polypeptides and nucleic acid phosphates (Herkovits and Perez-Coll, 1991). Allen (1994)
16 reported on the accumulation of numerous metals and ions into specific tissues of the tilapia
17 Oreochromis aureus. Tilapia exposed to low concentrations of Pb and mercury (both at
18 0.05 mg/L) had significantly higher concentrations of Pb in internal organs than those fish
19 exposed to Pb alone. Similarly, low concentrations of cadmium with low concentrations of Pb
20 caused increased uptake of Pb in certain organs (e.g., liver, brain, and caudal muscle).
21
22 Additive Interactions
23 The combined effects of two or more metals may result in additivity when the observed
24 effects are greater than that observed with individual metals but equivalent to a summation of the
25 effects from multiple metals. Lead has been shown to complex with Cl in aquatic systems. For
26 example, Verslycke et al. (2003) exposed the estuarine mysid Neomysis integer to six different
27 metals, including Pb, and a combined metal mixture under changing salinity conditions. At a
28 salinity of 5%, the reported LC50 for Pb was 1140 |ig/L (840, 1440 |ig/L). At an increased
29 salinity of 25%o, the toxicity of Pb was substantially reduced (LC50 = 4274 |ig/L [3540,
30 5710 //g/L]) (Verslycke et al., 2003). This reduction in toxicity was attributed to the increased
31 concentration of Cl ion due to increased salinity, in that it complexed with divalent Pb in the
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1 test system. Exposure ofN. integer to Pb in combination with the other five metals (Hg, Cd, Cu,
2 Zn, Ni) resulted in roughly strictly additive toxicity (Verslycke et al., 2003).
3 Long et al. (2006) performed a critical review of the uses of mean sediment quality
4 guideline quotients (mSQGQs) in assessing the toxic effects of contaminant mixtures (metals and
5 organics) in sediments. This approach has been used in numerous surveys and studies since
6 1994. mSQGQs are useful to risk assessors but their inherent limitations and underlying
7 assumptions must be fully understood (see Section AX8.2.1.5).
8
9 Summary of Interactions With Other Pollutants
10 Norwood et al. (2003) reported that in a review and reinterpretation of published data on
11 the interactions of metals in binary mixtures (n = 15 studies), antagonistic (6) and additive
12 interactions (6) were the most common for Pb. The complexity of the interactions and possible
13 modifying factors makes determining the impact of even binary metal mixtures to aquatic biota
14 difficult (Norwood et al., 2003; Playle, 2004). The two most commonly reported Pb-element
15 interactions are between Pb and calcium and between Pb and zinc. Both calcium and zinc are
16 essential elements in organisms and the interaction of Pb with these ions can lead to adverse
17 effects both by increased Pb uptake and by a decrease in Ca and Zn required for normal
18 metabolic functions.
19
20 AX8.2.3.5 Factors Associated with Global Climate Change
21 It is highly unlikely that Pb has any influence on generation of ground-level ozone,
22 depletion of stratospheric ozone, global warming, or other indicators of global climate change.
23 Lead compounds have relatively short residence times in the atmosphere, making it unlikely that
24 they will reach the stratosphere, and they do not absorb infrared radiation, making them unlikely
25 to contribute to stratospheric ozone depletion or global warming. Also, these compounds are
26 unlikely to have a significant interaction with ground-level nitrogen oxides or volatile organic
27 compounds, thus precluding generation of ground-level ozone.
28 Approached from another viewpoint, climate change can have a major impact on the
29 fate/behavior of Pb in the environment and, therefore, can subsequently alter organism or
30 ecosystem responses. For example, changes in temperature regime (Q10 rule), changes in
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1 precipitation quantity and quality (e.g., acidic deposition) may influence fate, transport, uptake,
2 and bioavailability of Pb (Syracuse Research Corporation., 1999).
3
4 AX8.2.3.6 Summary
5 There have been a number of advancements in the understanding of Pb behavior in the
6 environment and its impact on aquatic organisms since 1986. In particular, greater knowledge of
7 factors that influence Pb accumulation in aquatic organisms, mechanisms of detoxification and
8 avoidance of Pb, and greater understanding of the interactions of Pb in aquatic systems.
9 Recently, the development of the Biotic Ligand Model (BLM) and its exploration of the activity
10 of free metal ions at biologically reactive sites (i.e., fish gill tissue) have been a large contributor
11 to the understanding of metal speciation and movement into and effects to aquatic biota. To
12 date, there has been no BLM model of Pb although research has been conducted on a Pb-gill
13 binding model for rainbow trout. Further research in support of BLM model development for Pb
14 is recommended to further our understanding of these issues.
15
16 AX8.2.4 Exposure/Response of Aquatic Species
17 This section outlines and highlights the critical recent advancements in the understanding
18 of the toxicity of Pb to aquatic biota. The section begins with a review of the major findings and
19 conclusions from the 1986 Pb AQCD (U.S. Environmental Protection Agency, 1986a). The
20 following sections summarize the research conducted since 1986 on determining the
21 concentrations of Pb that cause the effects discussed in Section 8.2.3. Effects levels are
22 discussed at three primary trophic levels: primary producers, consumers, and decomposers.
23 Issues related to indirect effects (e.g., effects on predator/prey interactions, habitat alteration,) are
24 not to be addressed.
25
26 AX8.2.4.1 Summary of Conclusions From the Previous Criteria Document
27 The 1986 Pb AQCD (U.S. Environmental Protection Agency, 1986a) reviewed data in the
28 context of the sublethal effects of lead exposure. The document focused on describing the types
29 and ranges of lead exposures in ecosystems likely to adversely impact domestic animals. As
30 such, the criteria document did not provide a comprehensive analysis of the effects of lead to
31 most aquatic primary producers, consumers, and decomposers. For the aquatic environment,
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1 general reviews of the effects of lead to algae, aquatic vertebrates, and invertebrates were
2 undertaken. A summary of these reviews is provided below.
3
4 Algae
5 The 1986 Pb AQCD (U.S. Environmental Protection Agency, 1986a) reported that some
6 algal species (e.g., Scenedesmus sp.) were found to exhibit physiological changes when exposed
7 to high lead or organolead concentrations in situ. The observed changes included increased
8 numbers of vacuoles, deformations in cell organelles, and increased autolytic activity. Increased
9 vacuolization was assumed to be a tolerance mechanism by which lead was immobilized within
10 cell vacuoles.
11
12 Aquatic Vertebrates
13 The 1986 Pb AQCD (U.S. Environmental Protection Agency, 1986a) reported that
14 hematological and neurological responses were the most commonly reported effects in aquatic
15 vertebrates. These effects include red blood cell destruction and inhibition of the enzyme
16 ALAD, required for hemoglobin synthesis. At high lead concentrations, neurological responses
17 included neuromuscular distortion, anorexia, muscle tremor, and spinal curvature (e.g., lordosis).
18 The lowest reported exposure concentration causing either hematological or neurological effects
19 was 8 |ig/L (U.S. Environmental Protection Agency, 1986a).
20
21 Aquatic Invertebrates
22 Numerous studies were cited on the effects of lead to aquatic invertebrates in the 1986 Pb
23 AQCD (U.S. Environmental Protection Agency, 1986a). In general, lead concentrations in
24 aquatic invertebrates were found to be correlated closely with concentrations in water rather than
25 food. Freshwater snails were found to accumulate lead in soft tissue, often in granular bodies of
26 precipitated lead. Mortality and reproductive effects were reported to begin at 19 jig Pb/L for the
27 freshwater snail Lymneapalutris and 27 jig Pb/L for Daphnia sp.
28 The review of the NAAQS for Pb (U.S. Environmental Protection Agency, 1990) made
29 only one recommendation reported in the sections of the 1986 Pb AQCD dealing with effects to
30 aquatic biota. This was the need to consider the impact of water hardness on Pb bioavailability
31 and toxicity, to be consistent with the recommendations of the AWQC for the protection of
32 aquatic life (U.S. Environmental Protection Agency, 1985).
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1
2 AX8.2.4.2 Recent Studies on Effects of Lead on Primary Producers
3 Using literature published since the 1986 Pb AQCD (U.S. Environmental Protection
4 Agency, 1986a), this section examines the toxicity of Pb (individually and in metal mixtures) to
5 algal and aquatic plant growth, its effects on metabolic processes (e.g., nutrient uptake), and its
6 impact on primary productivity in natural systems.
7
8 Toxicity of Lead to Algae
9 The toxicity of Pb to algal growth has been investigated for a number of species including
10 Chlorella vulgaris, Closterium acerosum, Pediastrum simplex., Scenedesmus quadricauda,
11 Scenedesmu obliquus, Syneschoccus aeruginosus, and Nostoc muscorum (Jampani, 1988; Rai
12 and Raizada, 1989; Adam and Abdel-Basset, 1990; Fargasova, 1993; Bilgrami and Kumar,
13 1997). Study durations ranged from 7 to 20 days and Pb-nitrate was the most commonly used
14 form of Pb. Effects to algal growth {Chlorella vulgaris, Closterium acerosum, Pediastrum
15 simplex, Scenedesmus quadricauda), ranging from minimal to complete inhibition, have been
16 reported at Pb concentrations between 100 and 200,000 |ig/L (Jampani, 1988; Bilgrami and
17 Kumar, 1997). Most studies report the percent inhibition in test groups compared to controls
18 rather than calculating the LOEC, NOEC, or ECso values. Clinical signs of Pb toxicity include
19 the deformation and disintegration of algae cells and a shortened exponential growth phase
20 (Jampani, 1988; Fargasova, 1993). Other effects of Pb block the pathways that lead to pigment
21 synthesis, thus affecting photosynthesis, the cell cycle and division, and ultimately result in cell
22 death (Jampani, 1988).
23 From the studies reviewed, Closterium acerosum is the most sensitive alga species tested
24 (Bilgrami and Kumar, 1997). Exposure of these algae to 1000 and 10,000 jig/L as lead nitrate
25 for 6 days resulted in cell growth that was 52.6 and 17.4%, respectively, of controls (Bilgrami
26 and Kumar, 1997). Chlorella vulgaris, Pediastrum simplex, and Scenedesmus quadricauda were
27 also exposed to Pb-nitrate in this study. Compared to controls, cell growth at 1000 and 10,000
28 |ig Pb-nitrate/L was 65.3 and 48.7%, 64.5 and 42.7%, and 77.6 and 63.2%, respectively
29 (Bilgrami and Kumar, 1997). Scenedesmus quadricauda exhibited a similar magnitude of effects
30 when exposed to lead (Pb2+) for 20 days at 0, 5500, 11,000, 16,500, 22,000, 27,500, and 33,000
31 |ig/L (Fargasova, 1993). This study reported an ECso for growth inhibition at 13,180 |ig/L (95%
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1 CI: 10,190, 14,620). Decreased cell number, but increased cell size, was observed in
2 Selenastrum capricornutum1 exposed to lead (Pb2+) at 207.2 |ig/L and a Q/V (flux of air [Q]
3 divided by volume of the culture [V]) of 4.7 x 10~3 sec ~* for 9 days (Simoes Gon9alves et al.,
4 1991). The Q/V is a measure of culture growth where an increase in the Q/V ratio indicates
5 growth. The pigment concentration per cell decreased with exposure to Pb, so while the algae
6 cells were larger, they were less healthy (Simoes Gon9alves et al., 1991). Growth rates were not
7 reported, making comparison with other studies difficult.
8 High Pb concentrations were required to elicit effects in Nostoc muscorum and
9 Scenedesmus aeruginosus (Jampani, 1988; Rai andRaizada, 1989). Following 15-day
10 exposures, test groups exposed to 10,000, 20,000, and 30,000 jig Pb/L experienced growth rates
11 that were 90.5, 76.9, and 66.7% of the controls (Rai and Raizada, 1989). Synechococcus
12 aeruginosus experienced little inhibition of growth from exposure to Pb-nitrate up to a
13 concentration of 82,000 |ig/L (Jampani, 1988). At a test concentration of 100,000 |ig/L,
14 complete inhibition of growth was observed, and at a concentration of 200,000 |ig/L, algae failed
15 to establish a single colony (Jampani, 1988). Scenedesmus obliquus are quite tolerant to the
16 effects of Pb-nitrate and Pb-acetate on growth. Algae exposed to Pb-nitrate or Pb-acetate up to
17 180,000 |ig/L had higher cell numbers than controls (Adam and Abdel-Basset, 1990). Exposure
18 to the highest concentration of 300,000 jig/L Pb-nitrate or Pb-acetate resulted in cell numbers
19 that were 81 and 90% of the controls, respectively (Adam and Abdel-Basset, 1990).
20 Lead in combination with other metals (e.g., Pb and Cd, Pb and Ni, etc.) is generally less
21 toxic than exposure to Pb alone (Rai and Raizada, 1989). Nostoc muscorum exposed to
22 chromium and Pb in combination demonstrated better growth than when exposed to either of the
23 metals alone (Rai and Raizada, 1989). Antagonistic interaction was observed in the exposure of
24 Nostoc muscorum to Pb and nickel in combination (Rai and Raizada, 1989). When applied
25 separately, these metals demonstrated different levels of toxicity; however, in combination, they
26 exerted similar effects (Rai and Raizada, 1989). More information on toxic interactions of Pb
27 with other metals is provided in Section AX8.2.3.5.
28
The species name Selenastrum capricornutum has been changed to Pseudokirchneriella subcapitata. The
former species name is used in this report.
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1 Aquatic Plants
2 The toxicity of Pb to aquatic plant growth has been studied using Spirodelapolyrhiza,
3 Azollapinnata, and Lemna gibba (Gaur et al., 1994; Gupta and Chandra, 1994; Miranda and
4 Ilangovan, 1996). Test durations ranged from 4 to 25 days and test concentrations ranged
5 between 49.7 and 500,000 |ig Pb/L (Gaur et al., 1994; Miranda and Ilangovan, 1996). Research
6 on aquatic plants has focused on the effects of Pb on aquatic plant growth and chlorophyll and
7 protein content.
8 Of the species reviewed here, the effects of Pb on aquatic plant growth are most
9 pronounced in Azollapinnata (Gaur et al., 1994). An ECso of 1100 |ig/L was reported for A.
10 pinnata exposed to Pb-nitrate for 4 days. S. polyrhiza exposed to Pb-nitrate under the same test
11 conditions had a reported ECso for growth of 3730 |ig/L (Gaur et al., 1994). Lemna gibba was
12 shown to be the least sensitive plant species to Pb: significant growth inhibition was reported at
13 concentrations of 200,000 |ig/L or greater after 25 days of exposure to concentrations of 30,000,
14 50,000, 100,000, 200,000, 300,000, or 500,000 |ig/L (Miranda and Ilangovan, 1996). The
15 maximum growth rate for L. gibba was observed at 10 days of exposure. After this point, the
16 growth rate declined in controls and test concentrations (Miranda and Ilangovan, 1996). Clinical
17 signs of Pb toxicity include yellowing and disintegration of fronds, reduced frond size, and
18 chlorosis (Gaur et al., 1994; Miranda and Ilangovan, 1996). Toxicity results suggest that effects
19 to growth from Pb exposure occur in a dose-dependent manner (Gaur et al., 1994).
20
21 Effects of Lead on Metabolic Processes
22 Algal and aquatic plant metabolic processes are variously affected by exposure to Pb, both
23 singularly and in combination with other metals. Lead adversely affects the metabolic processes
24 of nitrate uptake, nitrogen fixation, ammonium uptake, and carbon fixation at concentrations of
25 20,000 jig Pb/L or greater (Rai and Raizada, 1989). Lead in combination with nickel has an
26 antagonistic effect on nitrogen fixation and ammonium uptake, but a synergistic effect on nitrate
27 uptake and carbon fixation (Rai and Raizada, 1989). Lead in combination with chromium has an
28 antagonistic effect on nitrate uptake, but it has a synergistic effect on nitrogen fixation,
29 ammonium uptake, and carbon fixation (Rai and Raizada, 1989).
30 Lead effects on nitrate uptake in Nostoc muscorum (jig NOs/jig Chi a) were greatest after
31 24 h, when exposure to 20,000 |ig/L reduced nitrate uptake by 64.3% compared to controls.
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1 Nitrate uptake reported after 48, 72, and 96 h was reduced by 30.0, 37.5, and 38.9%,
2 respectively, compared to controls (Rai and Raizada, 1989). Lead in combination with
3 chromium, both at a test concentration of 20,000 |ig/L, demonstrated antagonistic effects on
4 nitrate uptake. Compared to controls, nitrate uptake was reduced by 52.4, 30, 25, and 22.2% at
5 24, 48, 72 and 96 h, respectively (Rai and Raizada, 1989). The greatest effect on uptake
6 occurred at 24 h when, compared to controls, a 52.4% reduction was reported in the test
7 concentration. Lead and nickel in combination at test concentrations of 20,000 and 1000 |ig/L,
8 respectively, resulted in a greater reduction of nitrate uptake than Pb alone at 48, 72, and 96 h
9 (Rai and Raizada, 1989).
10 After 24, 48, and 72 h of Pb exposure at 20,000 |ig/L, nitrogenase activity (nmol C2H4/ jig
11 protein/hr) in Nostoc muscorum was reduced by 39.3, 61.8, and 14.1%, respectively, compared
12 to controls (Rai and Raizada, 1989). A concentration of 207.2 |ig Pb/L had little effect on
13 nitrogen or phosphorus assimilation in Selenastrum capricornutum over 7 days (Capelo et al.,
14 1993). An antagonistic effect on nitrogenase activity was generally reported for Nostoc
15 muscorum exposed to Pb in combination with nickel at 20,000 and 1,000 |ig/L, respectively (Rai
16 and Raizada, 1989). Compared to controls, nitrogenase activity was reduced by 42.9, 32.7, and
17 13.6% at 24, 48, and 72 h, respectively (Rai and Raizada, 1989). Lead and chromium, both
18 administered at a concentration of 20,000 |ig/L, had a synergistic impact on nitrogenase activity
19 in Nostoc muscorum. Nitrogenase activity in the test group was reduced by 60.7, 60, and 50%
20 compared to the controls at 24, 48, and 72 h, respectively (Rai and Raizada, 1989).
21 Lead-induced inhibition of ammonium uptake (jig NIL; uptake/jig Chi a) was greatest in
22 Nostoc muscorum after 48 h of exposure to 20,000 |ig/L of lead. Compared to controls, the Pb
23 test concentration 20,000 |ig/L reduced ammonium uptake by 72, 82, 61, and 26 % at 24, 48, 72,
24 and 96 h, respectively (Rai and Raizada, 1989). Lead in combination with nickel at
25 concentrations of 20,000 and 1,000 |ig/L, respectively, demonstrated an antagonistic effect on
26 ammonium uptake. Compared to controls, ammonium uptake in the test group was reduced by
27 44.9, 54.1, 23.3, and 4% at 24, 48, 72, and 96 h, respectively (Rai and Raizada, 1989). Lead in
28 combination with chromium, both at concentrations of 20,000 |ig/L, demonstrated a synergistic
29 interaction with 24, 48, 72, and 96 h uptake rates reduced by 87.2, 88.5, 72.5, and 50 %,
30 respectively, compared to controls (Rai and Raizada, 1989).
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1 Nostoc muscorum exposed to 20,000 jig Pb/L experienced the greatest reduction in carbon
2 fixation at 0.5 h of exposure: 62% compared to controls. Inhibition of carbon fixation in the test
3 group was less pronounced after 1 and 2 h of exposure: 29 and 13% of controls (Rai and
4 Raizada, 1989). Lead in combination with nickel or chromium had synergistic effects to carbon
5 fixation. Lead and nickel concentrations of 20,000 and 1000 |ig/L, respectively, resulted in 0.5,
6 1, and 2 h carbon fixation rates reduced by 93, 92, and 91%, respectively, compared to controls
7 (Rai and Raizada, 1989). Lead with chromium at concentrations of 20,000 |ig/L resulted in 0.5,
8 1, and 2 h carbon fixation rates reduced by 65, 58, and 50%, respectively, compared to controls.
9 Nutrients such as nitrogen, phosphate, sodium acetate, sodium carbonate, and citric acid
10 have been shown to protect against the toxic effects of Pb to algae (Jampani, 1988). Nitrogen
11 compounds (ammonium chloride, potassium nitrate, sodium nitrate, sodium nitrite) protected
12 Synechococcus aeruginosus from a lethal Pb-nitrate dose of 200,000 |ig/L (Jampani, 1988). Two
13 phosphates (K2HPO4 and Na2HPC>4) were found to improve Synechococcus aeruginosus survival
14 from 0 to 72% at 200,000 |ig/L of Pb-nitrate (Jampani, 1988).
15 Compared to controls, protein content was reduced by 54.2 and 51.9% in aquatic plants
16 Vallisneria spiralis and Hydrilla verticillata, respectively, exposed to Pb for 7 days at 20,720
17 |ig/L (Gupta and Chandra, 1994). Decreased soluble protein content has been observed in
18 Scenedesmus obliquus exposed to Pb-nitrate or Pb-acetate at concentrations greater than 30,000
19 Hg/L, and in L. gibba at concentrations greater than 200,000 |ig/L (Adam and Abdel-Basset,
20 1990; Miranda and Ilangovan, 1996). Lemna gibba also showed increased loss of soluble starch
21 at concentrations >200,000 |ig/L (Miranda and Ilangovan, 1996). Under the conditions
22 described previously (Gupta and Chandra, 1994), ECso values for chlorophyll content were
23 14,504 and 18,648 |ig/L for Vallisneria spiralis and Hydrilla verticillata, respectively (Gupta
24 and Chandra, 1994). Effects to chlorophyll a content have been observed in Scenedesmus
25 obliquus at Pb-nitrate and Pb-acetate concentrations >30,000 jig/L (Adam and Abdel-Basset,
26 1990).
27
28 Summary of Toxic Effects Observed in Single-Species Bioassays
29 Algae and aquatic plants have a wide range in sensitivity to the effects of Pb in water.
30 Both groups of primary producers experience ECso values for growth inhibition between
31 approximately 1000 and >100,000 |ig/L (Jampani, 1988; Gaur et al., 1994; Bilgrami and Kumar,
May 2006 AX8-179 DRAFT-DO NOT QUOTE OR CITE
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1 1997). The most sensitive primary producers reported in the literature for effects to growth were
2 Closterium acersoum and Azollapinnata (Gaur et al., 1994; Bilgrami and Kumar, 1997). The
3 least sensitive primary producers reported in the literature for effects to growth were
4 Synechococcus aeruginosus and Lemna gibba (Jampani, 1988; Miranda and Ilangovan, 1996).
5 Exposure to Pb in combination with other metals is generally less toxic to growth than exposure
6 to lead alone. Studies have shown that lead adversely affects the metabolic processes of nitrate
7 uptake, nitrogen fixation, ammonium uptake, and carbon fixation (Rai and Raizada, 1989). Lead
8 in combination with nickel or chromium produced synergistic effects for nitrate uptake,
9 nitrogenase activities, ammonium uptake, and carbon fixation (Rai and Raizada, 1989).
10
11 Leads Effects on Primary Productivity
12 Lead nitrate and Pb-acetate have been shown to have adverse effects on the primary
13 productivity of aquatic plants in two water bodies in India (Jayaraj et al., 1992). One of the two
14 water bodies was a freshwater tank that receives wastewater and supports a rich population of
15 hyacinths, and the other was a wastewater stabilization pond. Water quality characteristics in the
16 freshwater tank were pH = 7.5, dissolved oxygen = 6 mg/L, and water hardness (CaCOs) =
17 100 mg/L. Water quality characteristics in the wastewater pond were pH = 8.1, dissolved
18 oxygen = 6.2 mg/L, and water hardness (CaCOs) = 160 mg/L (Jayaraj et al., 1992). Lead nitrate
19 concentrations of 500, 5000, 10,000, 25,000, and 50,000 |ig/L were combined with appropriate
20 water samples in light and dark bottles and suspended in each of the water bodies for 4 h. The
21 concentrations of Pb-acetate (5000, 10,000, 25,000, 50,000, and 100,000 |ig/L) were applied in
22 the same manner. The ECso values were determined based on the concentration required to
23 inhibit gross productivity (GP) and net productivity (NP) by 50% (Jayaraj et al., 1992). The
24 results demonstrated that Pb-nitrate was more toxic to primary production than Pb-acetate.
25 In the freshwater tank, Pb-nitrate EC50 values for GP and NP were 25,100 and 6310 |ig/L,
26 respectively, compared to Pb-acetate ECso values of 50,100 and 28,200 jig/L for GP and NP,
27 respectively (Jayaraj et al., 1992). In the stabilization pond, Pb-nitrate ECso values for GP and
28 NP were 31,600 and 28,200 |ig/L, respectively, compared to Pb-acetate EC50 values of 79,400
29 and 316 |ig/L for GP and NP, respectively (Jayaraj et al., 1992). The higher toxicity reported in
30 the freshwater tank was attributed to differences in species composition and diversity. The
31 freshwater tank was dominated by water hyacinths that decreased the photic zone available for
May 2006 AX8-180 DRAFT-DO NOT QUOTE OR CITE
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1 photosynthesis and consumed a great deal of available nutrients. The stabilization pond had a
2 rich nutrient budget, resulting in improved alga growth and species diversity (Jayaraj et al.,
3 1992).
4
5 AX8.2.4.3 Recent Studies on Effects of Lead on Consumers
6 This section focuses on the effects of Pb to aquatic biota including invertebrates, fish, and
7 other biota with an aquatic life stage (e.g., amphibians). It is not intended to be a comprehensive
8 review of all research conducted. Rather, the intent is to illustrate the concentrations and effects
9 of Pb on freshwater and marine aquatic species. Eisler (2000) provides an overview of much of
10 the recent available literature on the toxicity of Pb to fish and aquatic invertebrates. An
11 extensive literature search was conducted using numerous electronic bibliographic and database
12 services (e.g., DIALOG, EPA ECOTOX) and limited temporally from 1986 to present. This
13 temporal limit was due to the availability of the EPA water quality criteria report for the
14 protection of aquatic life, released in 1986 (U.S. Environmental Protection Agency, 1986b).
15 Based on the results of the literature search and recent reviews of the toxicity of Pb (Eisler,
16 2000), numerous studies have been published on the toxicity of Pb to aquatic consumers.
17 Hardness, pH, temperature, and other factors are important considerations when characterizing
18 the acute and chronic toxicity of lead (Besser et al., 2005) (Section AX8.2.3.5). However, many
19 of the studies reviewed did not report critical information on control mortality, water quality
20 parameters, or statistical methods, making comparing effects between studies difficult. Studies
21 reporting only physiological responses to Pb exposure (e.g., reduction of ALAD) are not
22 discussed here, as this topic was covered more completely in Section AX8.2.3.4. This section
23 provides a review of toxicity studies conducted with invertebrates, fish, and other aquatic
24 organisms.
25
26 Invertebrates
27 Exposure of invertebrates to Pb can lead to adverse effects on reproduction, growth,
28 survival, and metabolism (Eisler, 2000). The following presents information on the toxicity of
29 Pb to invertebrates in fresh and marine waters.
30
May 2006 AX8-181 DRAFT-DO NOT QUOTE OR CITE
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1 Freshwater Invertebrates
2 Acute and chronic Pb toxicity data for freshwater invertebrates are summarized in Table
3 AX8-2.4.1. As described in Section AX8.2.3.5, water hardness is a critical factor governing the
4 solubility, bioavailability, and ultimately the toxicity of Pb. The acute and chronic toxicity of Pb
5 increases with decreasing water hardness as Pb becomes more soluble and bioavailable to
6 aquatic organisms. For example, Borgmann et al. (2005) examined the toxicity of 63 metals,
7 including Pb, to Hyalella azteca at two levels of water hardness (soft water hardness, 18 mg
8 CaCOs/L; hard water, 124 mg CaCOs/L). Lead was 23 times more acutely toxic to H. azteca in
9 soft water than hard water. Besser et al. (2005) found that acute toxicity to H. azteca was also
10 modified by water hardness.
11 At a mean pH of 7.97 in soft water (hardness (CaCOs) = 71 mg/L) mortality was >50%
12 for//, azteca at a dissolved Pb concentration of 151 |ig/L. The LOEC for survival in hard water
13 (hardness (CaCO3) = 275 mg/L) at pH 8.27 was 192 |ig/L as dissolved Pb and 466 |ig/L as total
14 Pb. Both waterborne and dietary Pb were found to contribute to reduced survival of//, azteca
15 (Besser et al., 2005).
16 Exposure duration may also play an important role in Pb toxicity in some species.
17 For example, Kraak et al. (1994) reported that filtration in the freshwater mussel Dreissena
18 polymorpha was adversely affected at significantly lower Pb concentrations over 10 weeks of
19 exposure than was the case after 48 h of exposure.
20 The influence of pH on lead toxicity in freshwater invertebrates varies between
21 invertebrate species. Over a 96-h exposure period, mortality increased with decreasing pH in the
22 bivalve Pisidium casertanum, while pH-independent mortality was reported for gastropod and
23 crustacean species under similar exposure conditions (Mackie, 1989). Cladocerans
24 (Ceriodaphnia dubia), amphipods (//. azteca), and mayflies (Leptophlebia marginata) were also
25 more sensitive to Pb toxicity at lower pH levels (Schubauer-Berigan et al., 1993; Gerhardt,
26 1994). Lead was 100 times more toxic to the amphipod, Hyalella azteca, at a pH range of 5.0 to
27 6.0 (Mackie, 1989) than at a pH range of 7.0 to 8.5 (Schubauer-Berigan et al., 1993).
28 The physiology of an aquatic organism at certain life stages may be important when
29 determining the toxicity of metals to test organisms. For example, Bodar et al. (1989) exposed
30 early life stages ofDaphnia magna to concentrations of Pb(NC>3)2. The test medium had a pH
31 of 8.3 + 0.2, water hardness (CaCO3) of 150 mg/L, and temperature of 20 + 1 °C. Lead
May 2006 AX8-182 DRAFT-DO NOT QUOTE OR CITE
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Table AX8-2.4.1. Effects of Lead to Freshwater and Marine Invertebrates
to
o
o
ON
X
oo
1
oo
OJ
o
^
Prj
H
6
o
21
o
^^
H
O
H
W
O
^
HH
H
h-l
Species
Freshwater
Cladoceran
(Ceriodaphnia dubia)
Worm
(Lumbriculus variegatus)
Amphipod
(Hyalella azteca)
Amphipod
(Hyalella azteca)
Amphipod
(Hyalella azteca)
Mayfly
(Leptophlebia marginata)
Mayfly
(Leptophlebia marginata)
Chemical
lead
chloride
lead
chloride
lead
chloride
lead
chloride
lead
chloride
lead
chloride
lead
chloride
Endpoint:
Cone. Oig/L)*
LC50:
280
>2,700
>2,700
LC50:
>8,000
>8,000
>8,000
LC50:
<90
>5,400
>5,400
LC50:
27(20.1-36.4)
LC50:
60 (53.6-67.3)
LC50:
1090
(400-133200)
LC50:
5000
Duration of
Exposure Water Chemistry
48 h pH:
6-6.5
7-7.5
8-8.5
Hardness: 280-300 mg/L CaCO3
96 h pH:
6-6.5
7-7.5
8-8.5
Hardness: 280-300 mg/L CaCO3
96 h pH:
6-6.5
7-7.5
8-8.5
Hardness: 280-300 mg/L CaCO3
8 days Hardness
130 mg/L
pH 7.8-8.6
8 days Hardness
130 mg/L
pH 7.8-8.6
96 h pH:
4.5
96 h pH
7.0
Test Type - Effect
static-survival
static-survival
static-survival
renewal, 1 -week-
old amphipods
renewal, 10- to 16-
week old
amphipods
acute - survival
acute - survival
Reference
Schubauer-Berigan
etal. (1993)
Schubauer-Berigan
etal. (1993)
Schubauer-Berigan
etal. (1993)
MacLean et al.
(1996)
MacLean et al.
(1996)
Gerhardt (1994)
Gerhardt (1994)
-------
Table AX8-2.4.1 (cont'd). Effects of Lead to Freshwater and Marine Invertebrates
^<:
to
o
o
ON
w i
X
oo
1
oo
o
m
H
6
O
0
— ]
o
o
H
W
o
o
1 — I
H
W
Species
Amphipod
(Hyalella azteca)
Bivalve
(Pisidium compressum)
Bivalve
(Pisidium casertanum)
Gastropod
(Amnicola limosa)
Mussel
(Dreissena polymorpha)
Mussel
(Dreissena polymorpha)
Amphipod
(Hyalella azteca)
Amphipod
(Hyalella azteca)
Chemical
lead
nitrate
lead
nitrate
lead
nitrate
lead
nitrate
lead
nitrate
lead
nitrate
lead
nitrate
lead
nitrate
Endpoint:
Cone. Qig/L)*
LC50:
10
21
18
LC50:
38,000
21,300
11,400
LC50:
23,600
23,500
56,000
LC50:
10,300
20,600
9,500
EC50:
370
91
LT50:
358
LC50:
4.8(3.3-7.1)
LC50:
113(101-126)
Duration of
Exposure Water Chemistry
96 h pH:
5.0
5.5
6.0
96 h pH:
3.5
4.0
4.5
96 h pH:
3.5
4.0
4.5
96 h pH:
3.5
4.0
4.5
48 h pH = 7.9;
10 wks CaCO3/L;
72 days pH = 7.9;
CaCO3/L;
Hardness = 150 mg
Temp = 15 °C
Hardness = 150 mg
Temp = 15 °C
7 days pH = 7.37 - 8.27
Hardness
DOC = 0.
7 days pH = 8.21
Hardness
DOC=1.
= 18 mg CaCO3/L
28mg/L
-8.46
= 124 mg CaCO3/L;
Img/L
Test Type - Effect Reference
acute-survival Mackie (1989)
acute- survival Mackie (1989)
acute- survival Mackie (1989)
acute- survival Mackie (1989)
renewal - filtration Kraak et al. (1994)
renewal - filtration Kraak et al. (1994)
renewal - survival Borgmann et al.
(2005)
renewal - survival Borgmann et al.
(2005)
-------
Table AX8-2.4.1 (cont'd). Effects of Lead to Freshwater and Marine Invertebrates
to
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o
X
oo
i
oo
o
l>
H
6
o
1 — J
z,
0
H
O
o
w
o
o
HH
H
W
Species
Mayfly
(Leptophlebia marginata)
Cladoceran
(D. magna)
Cladoceran
(D. magna)
Cladoceran
D. magna)
Amphipod
(Hyalella azteca)
Tubificid worm
(Tubifex tubifex)
Marine
Copepod
(Amphiascus tenuiremis)
Bivalve
(Mytilus galloprovincialis)
Chemical
lead
chloride
lead
nitrate
lead
chloride
lead
chloride
lead
lead
nitrate
lead
lead
nitrate
* - Brackets after effect concentration are
Endpoint: Duration of
Cone. (^ig/L)* Exposure
LC50: 96 h
1090 (95% C.I. =
133.2)
>5000
LC50: 48 h
0.45
NOEC: 12 to 21 d
260
NOEC: 10 d
270
LOEC: 96 h
(Dissolved Pb)
192
(Total Pb) 466
EC50: 24 h
237(183-316) 48 h
142 (107-184)
LC50: sediment 96 h
2462 ug
metal/dry
sediment
EC50 : 221
(58.9-346.3)
LOEC : 50
95% confidence intervals.
Water Chemistry
pH = 4.5-6.5;
DOC -21.6 mgCr1;
Cond = 7.0 ,uS cm"1
pH = 8.3± 0.2
Hardness (CaCO3) = 150 mg/L
Temp= 20 °C
Not specified
Not specified
pH = 8.27
Hardness (CaCO3) = 275 mg/L
Temp = 21.1 °C
pH = 7.5-7.7
Hardness = 245 mg/L
Temp = 29.5-31 °C
pH = 7.7± 0.1
Dissolved O2 -6.3 ± 0.3 mg/L
Salinity - 32 ppt
artificial seawater
Test Type - Effect
renewal - survival
static -
embryogenesis
renewal -
reproduction
renewal - growth
flow through -
survival
static -
immobilization
embryogenesis
Reference
Gerhardt (1994)
Bodaretal. (1989)
Enserink et al.
(1991)
Enserink et al.
(1991)
Besser et al. (2005)
Khangarot(1991)
Hagopian-Schlekat
etal. (2001)
Beiras and
Albentosa (2003)
-------
1 concentrations of < 100 mg/L had no impact on Daphnia egg development. The authors
2 suggested that this may due to the Daphnia egg structure, which consists of two layers: the inner
3 vitelline layer and outer chlorion layer. The chlorion layer in other species (e.g., rainbow trout)
4 is known to adsorb metals, thereby, preventing ionic injury to the developing embryo.
5 Exposures to sediment-associated Pb can be toxic to sediment-dwelling organisms.
6 In freshwater sediments, 48-h exposure of water fleas (Daphnia magnd) to 7000 mg Pb/kg dw
7 significantly reduced mobility, while exposure to 13,400 mg Pb/kg dw for 24 h produced the
8 same effect (Dave, 1992a,b). Longer-term (i.e., 14-day) exposure of midges (Chironomus
9 tentans) to sediments containing 31,900 mg Pb/kg dw resulted in 100% mortality.
10
11 Marine Invertebrates
12 In estuarine environments, salinity is an important modifying factor to Pb toxicity.
13 Verslycke et al. (2003) exposed the estuarine mysid Neomysis integer to individual metals,
14 including Pb, and metal mixtures under changing salinity. Water temperature (20+1 °C) and
15 salinity were reported, although no other water quality parameters were available (e.g., pH, water
16 hardness). At a salinity of 5%o, the reported LC50 for Pb was 1140 |ig/L (95% CI: 840,
17 1440 |ig/L). At an increased salinity of 25%o, the toxicity of lead was substantially reduced
18 (LC50 = 4274 |ig/L [3540, 5710 |ig/L]) (Verslycke et al., 2003).
19 Sensitivity to Pb can also vary between genders in some aquatic organisms. For example,
20 Hagopian-Schlekat et al. (2001) examined the toxicity of Pb-chloride in sediment and sediment
21 pore water to female and male estuarine copepods Amphiascus tenuiremis. The reported LCso
22 for total lead was 2462 mg Pb/kg dw (95% CI: 2097, 2891 mg Pb/kg dw). Gender effects were
23 observed in that male copepods were more sensitive (p = 0.038) to Pb than females as
24 determined by generalized linear model analysis.
25 Beiras and Albentosa (2003) examined the inhibition of embryo development in
26 commercial bivalves Ruditapes decussatus and Mytilus galloprovincialis after exposure to
27 concentrations of Pb(NOs)2 in seawater. No water chemistry parameters other than temperature
28 were reported (test conducted at 20 °C). An ECso range for R. decussatus was reported as 156 to
29 312 |ig/L, as insufficient data were available to calculate the actual ECso. The lowest observable
30 effect concentration (LOEC) was 156 |ig/L. ForM galloprovincialis., the ECso was 221 |ig/L
31 (95% CI: 58.9, 346.3) while the LOEC was reported as 50 |ig/L.
May 2006 AX8-186 DRAFT-DO NOT QUOTE OR CITE
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1 Fish
2 The general symptoms of Pb toxicity in fish include production of excess mucus, lordosis,
3 anemia, darkening of the dorsal tail region, degeneration of the caudal fin, destruction of spinal
4 neurons, ALAD inhibition, growth inhibition, renal pathology, reproductive effects, growth
5 inhibition, and mortality (Eisler, 2000). Toxicity in fish has been closely correlated with
6 duration of Pb exposure and uptake (Eisler, 2000). The following presents information on the
7 toxicity of Pb to fish in fresh and marine waters. Table AX8-2.4.2 summarizes the effects of Pb
8 on freshwater and marine fish.
9
10 Freshwater Fish
11 Many of the toxicity modifying factors described above and in Section AX8.2.3.5 (e.g.,
12 pH, DOC) for invertebrates are also important modifying factors for Pb toxicity to fish species.
13 The effects of pH on Pb bioavailability and subsequent toxicity have been well studied (Sayer
14 et al., 1989; Spry and Wiener, 1991; Schubauer-Berigan et al., 1993; Stouthart et al., 1994;
15 MacDonald et al., 2002; Rogers and Wood, 2003). Schubauer-Berigan et al. (1993) exposed
16 fathead minnow to Pb-chloride over 96 hours. The reported LCso ranged from 810 to >5400
17 Hg/L at pH 6 to 6.5 and pH 7 to 8.5, respectively.
18 Water hardness also has a strong influence on the effects of lead to fish. Chronic
19 exposure of rainbow trout fry to Pb in soft water resulted in spinal deformities at 71 to 146 |ig/L
20 after 2 months of exposure (Sauter et al., 1976) or 13.2 to 27 |ig/L (Davies and Everhart, 1973;
21 Davies et al., 1976), after 19 months of exposure. When exposed to Pb in hard water, only 0 and
22 10% of the trout (Oncorhynchus mykiss) developed spinal deformities at measured Pb
23 concentrations of 190 and 380 |ig/L, respectively. In soft water, 44 and 97% of the trout
24 developed spinal deformities at concentrations of 31 and 62 |ig/L, respectively (Davies et al.,
25 1976). The maximum acceptable toxicant concentration (MATC) for rainbow trout fry in soft
26 water was 4.1 to 7.6 |ig/L (Davies et al., 1976), while the MATC for brook trout was 58 to
27 119 |ig/L (Holcombe et al., 1976). Histological reproductive abnormalities were noted in mature
28 male rainbow trout at 10 |ig/L Pb-nitrate (Ruby et al., 1993).
29 Schwartz et al. (2004) examined the influence of NOM on Pb toxicity to rainbow trout
30 exposed for 96 h in a static system. The pH of the exposure system ranged between 6.5 and 7.0,
31 temperature was maintained between 9 and 11 °C, and Pb was added as PbCb. NOM from a
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Table AX8-2.4.2. Effects of Pb to Freshwater and Marine Fish
to
o
o
o^
X
5,400
>5,400
lead Reproductive effects:
nitrate 10
lead Reproductive Effects:
acetate 500
lead LC50: 1000
nitrate (800 - 1400)
not LC50:
reported 6.5 cm fish- 1030
3. 5 cm fish -300
Duration of
Exposure Water Chemistry
96 h pH:
6-6.5
7-7.5
8-8.5
Hardness: 280-300
mg/L CaCO3
12 days Hardness 128 mg/L
CaC03
29 days pH:
7.5-8.5;
Hardness 130 mg/L
CaCO3;
22-25 °C
(Pb 95% soluble)
96 h pH:
7.9-8.0
DOC = 3 mg/L
Hardness (CaCO3) =
140 mg/L
96 h pH:
7.1
Temperature- 15 C
Oxygen sat. 6.4 mg/L
Comments
static, measured
Decreased
spermatocyte
development
Fewer viable eggs
produced, testicular
damage
Flow through - Survival
static -renewal -
Survival
Reference
Schubauer-Berigan
etal. (1993)
Ruby etal. (1993)
Weber (1993)
Rogers and Wood
(2003)
Alam and Maughan
(1995)
-------
1 number of U.S. rivers and lakes was then added to the test system, and the LT50 was reported.
2 NOM was found to reduce the toxic effects of Pb to rainbow trout.
3 Fish size is an important variable in determining the adverse effects of Pb. Alam and
4 Maughan (1995) exposed two different sizes of common carp (Cyprinus carpio) to Pb
5 concentrations to observed effects on carp mortality. Water chemistry parameters were reported
6 (pH = 7.1; temperature = 20 °C). Smaller fish (3.5 cm) were found to be more sensitive to Pb
7 than were larger fish (6.5 cm). The reported LCsoS were 0.44 mg/L and 1.03 mg/L, respectively.
8
9 Marine Fish
10 There were no studies available that examined the toxicity of Pb to marine fish species for
11 the time period examined (1986 to present). However, Eisler (2000) reviewed available research
12 on Pb toxicity to marine species and reported studies done prior to 1986. Acute toxicity values
13 ranged from 50 |ig/L to 300,000 |ig/L in plaice (Pleuronectesplatessd) exposed to organic and
14 inorganic forms of Pb (Eisler, 2000). Organolead compounds (e.g., tetramethyl Pb, tetraethyl Pb,
15 triethyl Pb, diethyl Pb) were generally more toxic to plaice than inorganic Pb (Maddock and
16 Taylor, 1980).
17
18 Other A quatic Biota
19 A paucity of data exist on the effects of Pb to growth, reproduction, and survival of
20 aquatic stages of frogs and turtles. Rice et al. (1999) exposed frog larvae (Rana catesbeiand) to
21 780 |ig Pb/L and two oxygen concentrations (3.5 or 7.85 mg/L) for 7 days (Table AX8-2.4.3).
22 Exposure conditions included water hardness of 233 to 244 mg CaCOs/L, pH from 7.85 to 7.9,
23 and temperature at 23 °C. Frog larvae were found to display little to no activity in the low
24 oxygen and high Pb treatment. Hypoxia-like behavior was exhibited in larvae exposed to both
25 low and high oxygen concentrations and high Pb. Therefore, larvae of R. catesbeiana showed
26 sensitivity to Pb and responded with hypoxia-like behavior. Additionally, the larvae in the Pb
27 treatment were found to have lost body mass relative to controls and the other treatments. Rice
28 et al. (1999) suggested that the decrease in mass likely indicated the beginning of a period of
29 reduced growth rate. Larvae exposed for longer periods (>4 weeks) were smaller and
30 metamorphosed later compared to unexposed individuals.
31
May 2006 AX8-189 DRAFT-DO NOT QUOTE OR CITE
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Table AX8-2.4.3. Nonlethal Effects in Amphibians
to
o
o
ON
Species
Frogs
(Rana ridibunda)
Frogs
(Bufo arenamm)
Frogs
Endpoint:
Chemical Concentration
lead nitrate Biochemical effects:
14,000 ug/L
Mortality:
16 mg Pb2+/L
Hypoxia-like behavior:
Duration of
Exposure
30 days
5 days
7 days
Water Chemistry
not specified
not specified
O2 = 3. 5-7.85 mg/L
Comments
Hepatic ALAD decreased by
90%
Effects reported include erratic
swimming, loss of equilibrium
Larvae used
Reference
Vogiatzis and
Loumbourdis (1999)
Herkovits and Perez-Coll
(1991)
Puce etal. (1999)
X
oo
VO
o
(Rana catesbeiana)
780 ug/L
Turtle Hatchlings lead acetate NOEL: 100 ug/g
(Trachemys scripta) (Survival and
behavior)
4 weeks
pH = 7.85-7.9
Temp = 23 °C
CaCO3 = 233-244
mg/L
N/A
Exposure via single injection Burger et al. (1998)
H
6
o
o
H
O
O
H
W
O
O
HH
H
W
-------
1 Herkovits and Perez-Coll (1991) examined Pb toxicity to amphibian larvae (Bufo
2 arenarum). Larvae (n = 50) were exposed for up to 120 h at two Pb concentrations, 8 mg Pb2+/L
3 and 16 mg Pb2+/L. Relative to controls, the 8 mg Pb2+/L treatment group exhibited 40%
4 mortality and the 16 mg Pb2+/L group 60% mortality after 120 h (p < 0.05). The authors reported
5 behavioral effects, erratic swimming, and loss of equilibrium during the tests, symptoms that are
6 consistent with the action of Pb on the central and peripheral nervous systems (Rice et al., 1999).
7 Behavior (i.e., righting, body turnover, seeking cover), growth, and survival of hatchling
8 slider turtles (Trachemys scriptd) exposed to Pb-acetate were investigated in one study (Burger
9 et al., 1998). In the first part of the study, 6-month-old hatchlings received single Pb-acetate
10 injections at 50 or 100 jig/g body weight (bw). In the second part of the study, 3-week-old
11 turtles were injected once with doses of 250, 1000 or 2500 jig/g bw. There were no differences
12 in survival, growth, or behavior for hatchlings in the first study, however, several effects were
13 reported from the second part of the study at doses in the range of 250 to 2,500 jig/g bw. As the
14 dose increased, so did the plastron length (i.e., ventral section of the shell), carapace length, and
15 weight. The highest dose group had the lowest survival rate with an LD50 of 500 jig/g bw.
16 Behavioral effects included slower times of righting behavior and seeking cover. The authors
17 suggested a NOEL of 100 jig/g bw for slider turtles for survival and behavior.
18
19 AX8.2.4.4 Recent Studies on Effects of Lead on Decomposers
20 In this section, decomposers are defined as being bacteria and other microorganisms.
21 Many invertebrates are also potentially considered decomposers, but the effects of Pb to
22 invertebrates have been described in previous sections. There were no toxicity studies located on
23 the effects of Pb to aquatic decomposers in the time period of interest.
24
25 AX8.2.4.5 Summary
26 Lead in all its forms is known to cause adverse effects in aquatic organisms (Eisler, 2000).
27 Effects to algal growth have been observed at b concentrations ranging from 100 to
28 200,000 |ig/L. Clinical signs of Pb toxicity in plants include the deformation and disintegration
29 of algae cells and a shortened exponential growth phase. Other effects of Pb include a blocking
30 of the pathways that lead to pigment synthesis, thus affecting photosynthesis, cell cycle and
31 division, and ultimately resulting in death. The toxicity of Pb to macrophyte growth has been
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1 studied using Spirodelapolyrhiza, Azollapinnata, and Lemna gibba. Test durations ranged from
2 4 to 25 days and test concentrations ranged between 49.7 and 500,000 |ig/L.
3 Waterborne Pb is highly toxic to aquatic organisms, with toxicity varying with the species
4 and life stage tested, duration of exposure, form of Pb tested, and water quality characteristics.
5 Among the species tested, aquatic invertebrates, such as amphipods and water fleas, were the
6 most sensitive to the effects of Pb, with adverse effects being reported at concentrations ranging
7 from 0.45 to 8000 |ig/L. Freshwater fish demonstrated adverse effects at concentrations ranging
8 from 10 to >5400 |ig/L, depending generally upon water quality parameters. Amphibians tend to
9 be relatively Pb tolerant; however, they may exhibit decreased enzyme activity (e.g., ALAD
10 reduction) and changes in behavior (e.g., hypoxia response behavior). Lead tends to be more
11 toxic with longer-term exposures.
12
13 AX8.2.5 Effects of Lead on Natural Aquatic Ecosystems
14 Introduction
15 This section discusses the effects of Pb on natural aquatic ecosystems. Such effects
16 include changes in species composition and richness, ecosystem function, and energy flow due to
17 Pb stress. The format of this section generally follows a conceptual framework for discussing
18 the effects of a stressor such as Pb on an ecosystem. This conceptual framework was developed
19 by the EPA Science Advisory Board (Young and Sanzone, 2002). The essential attributes used
20 to describe ecological condition include landscape condition, biotic condition, chemical and
21 physical characteristics, ecological processes, hydrology and geomorphology and natural
22 disturbance regimes. The majority of the published literature pertaining to Pb and aquatic
23 ecosystems focuses on the biotic condition, one of several essential attributes of an ecosystem as
24 described in Young and Sanzone (2002). For the biotic condition, the SAB framework identifies
25 community extent, community composition, trophic structure, community dynamics, and
26 physical structure as factors for assessing ecosystem health. Other factors for assessing the
27 biotic condition such as effects of Pb on organs, species, populations, and organism conditions
28 (e.g., physiological status) were discussed in Sections AX8.2.3 and AX8.2.4.
29 For natural aquatic ecosystems, the focus of study in the general literature has been on
30 evaluating ecological stress where the sources of Pb were from urban and mining effluents rather
31 than atmospheric deposition (Poulton et al., 1995; Deacon et al., 2001; Mucha et al., 2003). The
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1 atmospheric deposition of Pb in remote lakes has been evaluated; however, the direct effects of
2 Pb on aquatic ecosystems was not evaluated in many cases (Larsen, 1983; Kock et al., 1996;
3 Allen-Gil et al., 1997; Outridge, 1999; Letter et al., 2002). In other studies, although Pb
4 deposition was studied, the effects of acid deposition on aquatic life were the focus of the study
5 and perceived to be more relevant (Mannio, 2001; Nyberg et al., 2001). Finally, the effects of
6 Pb, other metals, and acidification on phytoplankton have only been inferred based on the
7 paleolimnolical record (Tolonen and Jaakkola, 1983; Rybak et al., 1989). The statistical
8 methods used when evaluating the effects of Pb on aquatic ecosystems are important, as more
9 than one variable may be related to the observed effect. Studied variables include water
10 hardness, pH, temperature, and physical factors such as embeddedness, dominant substrate, and
11 velocity. In most cases single variable statistical techniques were used to evaluate the data.
12 However, in other cases multivariate techniques were used. Therefore, where appropriate, some
13 detail on the statistical methods used is presented.
14 Although most of the available studies discussed in this section focus on the biotic
15 condition, one case study examining multiple components of the EPA conceptual framework is
16 also included. The remainder of this section describes the effects of Pb on the biotic condition.
17
18 AX8.2.5.1 Case Study: Coeur d'Alene River Watershed
19 The Coeur d'Alene River watershed is an area of Idaho impacted by Pb and other metals
20 from historic mining waste releases. Maret et al. (2003) examined several ecological
21 components to determine any negative associations with metals and the watershed communities.
22 The variables examined and associated ecological conditions are presented in Table AX8-2.5.1.
23 In addition to measurements of non-metal variables (e.g., dissolved oxygen levels, water
24 temperature and pH, embeddedness), Cd, Pb, and Zn levels were also compared in affected sites
25 versus reference sites.
26 Some of the above non-metal variables are important to macroinvertebrate communities.
27 For example, a stream with highly embedded substrate can have a lower number of individuals
28 within a species or a different species composition compared to a stream with less embeddedness
29 (Waters, 1995). Macroinvertebrates from the Ephemeroptera (mayflies), Plecoptera (stoneflies),
30 and Trichoptera (caddisflies) (EPT) group inhabit the surface of cobble and the interstitial spaces
31 between and underneath cobble. When substrate is embedded, these interstitial spaces are filled,
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Table AX8-2.5.1. Ecological Attributed Studies by Maret et al. (2003)
in the Coeur d'Alene Watershed
Ecological Attribute
Subcategory
Measure
Landscape condition
Biotic Condition
Chemical/physical
characteristics
Ecological processes
Hydrology/geomorphology
Natural disturbance regimes
Areal extent
landscape pattern
Organism condition
population
structure/dynamics
Chemical/physical
parameters
Channel morphology
and distribution
Basin area (km2)
Production mine density/km2
Caddisfly tissue concentrations
(mg/kg)
Number of EPT taxa
Density of EPT individuals (no./m2)
Dissolved oxygen (mg/L)
Specific conductance (|iS/cm)
Water temperature (E °C)
pH
Water hardness (mg/L)
Total NO3 (mg/L)
Total P (mg/L)
Dissolved NH3 (mg/L)
Sediment Cd, Pb, Zn (mg/kg)
Dissolved Cd, Pb, Zn in water (mg/L)
None measured
Site elevation (m)
Stream gradient (%)
Stream discharge (m3/s)
Stream width (m)
Stream depth (m)
Open canopy (%)
Stream velocity (m/s)
Embeddedness (%)
Dominant substrate (mm)
None measured
4
5
6
leaving less habitat space for EPT taxa. In another example, water temperature is important;
some macroinvertebrates (e.g., stoneflies) are usually only found in cooler water (Harper and
Stewart, 1984).
Of the variables examined only metal concentrations, mine density, site elevation, and
water temperature were significantly different between reference and mine-affected sites.
A Mann-Whitney t-test was used to evaluate statistical differences between reference and test
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1 sites for physical and water quality parameters, while Spearman's rank correlation matrices were
2 used to compare all possible response and explanatory variables. Lead concentrations were
3 significantly correlated with the number of mines in proximity to the watershed. Lead
4 concentrations in sediment and water were strongly correlated to Pb levels in whole caddisflies,
5 r2 = 0.90 and 0.63, respectively. Furthermore, mine density was significantly correlated to Pb in
6 tissue, r2 = 0.64. Although temperature was significantly different between reference and mine-
7 affected sites, temperature conditions were concluded to be non-limiting to aquatic life. For
8 example, reference and mine-affected sites had at least 15 and 13 obligate cold-water taxa,
9 respectively.
10 A significant negative correlation between Pb in the water column (0.5 to 30 |ig/L
11 dissolved) and total taxa richness, EPT taxa richness, and the number of metal-sensitive mayfly
12 species was observed. Similar, significant negative correlations were found between sediment
13 Pb levels (132 to 6252 ug/g) and the same macroinvertebrate community metrics and caddisfly
14 tissue levels. Negative correlations were also found between Cd and Zn in the water and
15 sediment and the macroinvertebrate community metrics. In an analysis of cumulative toxicity,
16 Pb was judged to be the most significant metal in sediment related to the cumulative toxicity
17 measured. This study provided multiple lines of evidence (i.e., mine density, metal
18 concentrations, bioaccumulation in caddisfly tissue and benthic invertebrate assemblage
19 structure) of the negative impacts of mining in the Coeur d'Alene River, suggesting that Pb (and
20 other metals) were primary contributors to the effects observed in the Coeur d'Alene River
21 watershed (Maret et al., 2003).
22
23 AX8.2.5.2 Biotic Condition
24 In an evaluation of the biotic condition, the SAB framework described by Young and
25 Sanzone (2002) identifies community extent, community composition, trophic structure,
26 community dynamics, and physical structure as essential ecological attributes for assessing
27 ecosystem health. The following two sections describe the effects of Pb on community
28 composition, community dynamics, and trophic structure. To date, no available studies were
29 located on the effects of Pb on physical structure (e.g., change in riparian tree canopy height,
30 ecosystem succession).
31
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1 Ecosystems and Communities, Community Composition
2 To measure community composition, an inventory of the species/taxa found in the
3 ecological system must be conducted. According the SAB framework, useful measures of
4 composition include the total number of species or taxonomic units, their relative abundance,
5 presence and abundance of native and non-native species, and information on the presence and
6 abundance of focal or special interest species (Young and Sanzone, 2002). Focal or special
7 species of interest can be those that play a critical role in ecosystem processes such as flows of
8 materials or energy within complex food-webs (Young and Sanzone, 2002). Community
9 composition as assessed in Pb studies has included the following measures.
10
11 • Changes in energy flow or nutrient cycling:
12 o Increased or decreased respiration or biomass
13 o Increased or decreased turnover/cycling of nutrients
14
15 • Changes in community structure:
16 o Reduced species abundance (i.e., the total number of individuals of a species within
17 a given area or community)
18 o Reduced species richness (i.e., the number of different species present in
19 a community)
20 o Reduced species diversity (i.e., a measure of both species abundance and species
21 richness)
22 Investigators have evaluated the effects of Pb on aquatic communities through microcosm
23 and mesocosm studies in natural aquatic systems. Field studies in the general literature have
24 focused on natural systems that were affected by metal stress from various anthropogenic
25 sources. In most of those natural systems, the sources evaluated were from direct mining waste
26 inputs, rather than atmospheric deposition, of Pb. Studies published since the 1986 Pb AQCD
27 (U.S. Environmental Protection Agency, 1986a) that describe the effects of Pb on natural aquatic
28 ecosystems are presented below and summarized in Table AX8-2.5.2. Studies included here
29 evaluated the effects of Pb on watersheds, landscapes, aquatic ecosystems, aquatic communities,
30 biodiversity, lakes, rivers, streams, estuaries, wetlands, and species interaction.
31
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Table AX8-2.5.2. Essential Ecological Attributes for Natural Aquatic Ecosystems Affected by Lead
to
o
o
ON
oo
1
VO
o
H
6
O
2
0
H
O
o
H
W
O
O
HH
H
W
Condition
Category Species Measures
Biotic Condition
Ecosystems and Protozoan community Reduced species
Communities- abundance and
Community diversity
Composition
Protozoan community Reduced species
abundance
Protist community Reduced species
abundance and
diversity
Meiofauna community Reduced abundance
Algal community Increased respiration
Algal community Decreased primary
productivity
Meiobenthic Reduced species
community (primarily abundance
nematodes) No effect on
abundance
Exposure
Medium
Marine water
Freshwater
water
Marine water
Marine
sediment
Freshwater
Freshwater
Marine
sediment
Location
Laboratory microcosm
Laboratory microcosm
Laboratory microcosm
Laboratory microcosm
Domestic water
stabilization pond
Sharana Basaveshwara
Tank, India
Laboratory microcosm
Exposure
Concentrations
0.02- 0.05 mg/L
0.05-1 mg/L
1 mg/L
177 mg/kg dw
25-80 mg/L
6-32 mg/L
1343 mg/kg dw
1580 mg/kg dw
Other
Metals
Present Reference
N Femandez-
Leborans and
Novillo(1992)
N Femandez-
Leborans and
Antonio-Garcia
(1988)
N Femandez-
Leborans and
Novillo(1994)
Y Millward et al.
(2001)
? Jayaraj etal. (1992)
? Jayaraj etal. (1992)
N Austen and
McEvoy(1997)
-------
Table AX8-2.5.2 (cont'd). Essential Ecological Attributes for Natural Aquatic Ecosystems Affected by Lead
^<:
to
o
X
-------
Table AX8-2.5.2 (cont'd). Essential Ecological Attributes for Natural Aquatic Ecosystems Affected by Lead
to
o
o
ON
Category Species
Macroinvertebrate
Community
Fish Community
Ecosystems and Snails and tadpoles
Communities-
Condition Measures
Lead in tissues and
sediment not correlated
to diversity and
richness
Lead in tissues and
sediment not correlated
to diversity and
richness
Lead affected predator-
prey interactions
Exposure
Medium
Sediment
and whole
organism
residue
Sediment
and whole
organism
residue
Sediment
Location
Aquashicola Creek
tributaries, Palmerton,
PA
Aquashicola Creek
tributaries, Palmerton,
PA
Outdoor mini-
ecosystems
Exposure
Concentrations
7.5-59.5 mg/kg dw
(sediment)
0.25-6.03 mg/kg dw
(macroinvertebrates)
7.5-59.5 mg/kg dw
(sediment)
0.1 -0.86 mg/kg dw
(fish)
Not cited
Other
Metals
Present
Y
Y
Y
Reference
Carline and Jobsis
(1993)
Carline and Jobsis
(1993)
Lefcortetal. (1999)
X
oo
VO
VO
Community
Dynamics and
Trophic Structure
Snails and caddisflies
No avoidance of
predator by snail.
Caddisfly did respond
to predator
Water Field microcosm for
snail; in-stream
disturbance for
caddisfly
27.7-277.6 mg/kg
dw (snail tissue)
223-13,507 mg/kg
dw (caddisfly tissue)
Y
Lefcort et al. (2000)
£
£
H
1
O
o
H
O
o
H
W
O
O
HH
H
W
Fathead minnow Feeding behavior Water Laboratory microcosm 0.5-l.Omg/L
altered
American toad No avoidance of lead Water Laboratory microcosm 0.5-l.Omg/L
Mummichog Feeding behavior Water Laboratory 0.3-l.Omg/L
altered and predator
avoidance affected
N Weber (1996)
N Steeleetal. (1991)
N Weis and Weis
(1998)
-------
1 Aquatic Microcosm Studies
2 The examination of simulated aquatic ecosystems (i.e., microcosms) provides limited
3 information on the effects of pollutants on natural systems. Microcosm studies typically focus
4 on only a few aspects of the natural system and do not incorporate all of the ecological,
5 chemical, or biological interactions. Nevertheless, a few microcosm studies have been
6 conducted that indicate potential effects of Pb on the community structure of aquatic ecosystems.
7 Fernandez-Leborans and Antonio-Garcia (1988) evaluated the effect of Pb on a natural
8 community of freshwater protozoans in simulated aquatic ecosystems and found a reduction in
9 the abundance and composition of protozoan species with increasing Pb concentrations (0.05 to
10 1.0 mg/L) compared to controls. Studies with marine protozoan communities in laboratory
11 microcosms indicated that waterborne Pb exposure reduced protozoan abundance, biomass, and
12 diversity at concentrations of 0.02 to 1.0 mg/L Pb. (Fernandez-Leborans and Novillo, 1992,
13 1994).
14 Austen and McEvoy (1997) studied the effects of Pb on an estuarine meiobenthic
15 community (mainly nematodes) in a microcosm setting using sediment samples collected
16 offshore from England. A multivariate analysis of similarities (ANOSEVI) test with square root-
17 transformed data was used to evaluate differences between treatments and controls. Lead was
18 found to significantly affect species abundance at 1343 mg/kg dw relative to a control at
19 56 mg/kg dw, but no significant adverse effects were observed at the highest dose tested,
20 1580 mg/kg dw. The authors did not attempt to explain why the 1580 mg/kg dw dose was not
21 significant while the 1343 mg/kg dw dose was. None of the Pb exposures were significantly
22 different than the controls based on separate univariate tests of abundance, richness, and
23 diversity. There were no other confounding metals in the Pb tests, as the experiments were with
24 a single metal dose. In one other mesocosm study, the effects of a mixture of metals (Cu, Cd,
25 Pb, Hg, and Zn) on a salt marsh meiofaunal community were evaluated (Millward et al., 2001).
26 After 30-days exposure, significant reductions in copepod, gastropod, and bivalve abundances
27 were observed at the highest Pb exposure concentration, 177 mg/kg dw. Ostracods and
28 nematodes were not affected. The authors believed that the response of the meiofauna taxa to
29 metals was in part due to the various feeding strategies in that deposit feeders were most
30 affected.
31
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1 Natural Aquatic Ecosystem Studies
2 Lead stress in aquatic ecosystems has also been evaluated in natural communities.
3 Studies examining community-scale endpoints, however, are complex, and interpretation can be
4 confounded by the variability found in natural systems and the presence of multiple stressors.
5 Natural systems frequently contain multiple metals, making it difficult to attribute observed
6 adverse effects to single metals. For example, macroinvertebrate communities have been widely
7 studied with respect to metals contamination and community composition and species richness
8 (Winner et al., 1980; Chadwick et al., 1986; Clements, 1994). In these studies, multiple metals
9 are evaluated and correlations between observed community level effects are ascertained. The
10 results often indicate a correlation between the presence of one or more metals (or total metals)
11 and the negative effects observed. While, correlation may imply a relationship between two
12 variables, it does not imply causation of effects. The following studies suggest an association
13 between Pb concentration and an alteration of community structure and function (see summary
14 in Table AX8-2.6.2):
15
16 Reduced Primary Productivity and Respiration
17 Jayaraj et al. (1992) examined the effects of Pb on primary productivity and respiration in
18 an algal community of two water bodies. Concentrations of Pb in water (6 to 80 mg/L) were
19 found to significantly reduce primary productivity and increase respiration. The authors
20 suggested that increased respiration indicated a greater tolerance to or adaptive mechanisms of
21 the resident heterotrophs to cope with lead stress.
22
23 Alterations of Community Structure
24 Deacon et al. (2001) studied a macroinvertebrate community in mine-affected waters of
25 Colorado. Initially, transplanted bryophytes were used to assess whether metals could
26 bioaccumulate at various mine-affected and unaffected sites (Deacon et al., 2001; Mize and
27 Deacon, 2002). Lead was bioaccumulated by the bryophytes, and median tissue concentrations
28 at mine-affected sites (34 to 299 ug/g dw) were higher than at reference sites (2.5 to 14.7 ug/g
29 dw). Lead concentrations in surface water and sediment ranged from <0.001 to 0.02 mg/L and
30 145 to 850 mg/kg dw (<63 um fraction), respectively. The same sites were also evaluated for the
31 effects of various metals on macroinvertebrate communities. Values of total abundance, taxa
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1 richness, mayfly, and stonefly abundance were reduced at mining sites. Lead levels along with
2 Cd, Cu, and Zn were correlated with reduced abundance and diversity indices.
3 Macrobenthic communities studied in a Portuguese estuary were affected by Pb at a range
4 from 0.25 to 192 mg/kg dw (Mucha et al., 2003). Species richness was decreased in areas with
5 increased Pb concentrations in the sediment. Interpretation of Pb effects was complicated by
6 other non-metal stressors, namely sediment particle size and organic matter content.
7 Furthermore, other metals were present (e.g., Al, Cu, Cr, Mn, Zn) and may have affected the
8 community (Mucha et al., 2003).
9 The effects of Pb on oligochaetes in the 111 River and its tributaries in France were
10 evaluated by Rosso et al. (1994). Lead in sediment (5 to 16 ug/g dw at affected sites) was
11 positively correlated to the abundance of the oligochaete, Nais sp., and negatively correlated to
12 Tubificidae abundance. Lead was the only metal that was positively correlated to Nais species,
13 while other metals were negatively correlated to Tubificidae (Rosso et al., 1994).
14 The effects of metals and particle size on structuring epibenthic sea grass fauna (fish,
15 mollusks, crustaceans, and polychaetes) was evaluated near a Pb smelter in South Australia
16 (Ward and Young, 1982; Ward and Hutchings, 1996). Effluent from the smelter was the primary
17 source of Pb and other metal contamination. Species richness and composition were evaluated
18 near the Pb smelter along with metal concentrations in sediment. Lead levels in sediment (up to
19 5270 mg/kg dw) correlated with negative effects on species richness and composition, while the
20 other metals evaluated had similar correlations. Therefore, Pb alone could not be identified as
21 the sole metal causing stress.
22
23 Tissue Bioaccumulation Associated with Alterations of Community Structure
24 Several studies have examined the bioaccumulation of lead in aquatic systems with
25 indices of community structure and function. A focused study on changes in Chironomidae
26 community composition in relation to metal mines (New Brunswick, Canada) identified changes
27 in Chironomidae richness (Swansburg et al., 2002). Lead was not detected (detection limit not
28 given for any matrix) in the water column at any site. However, Pb levels in periphyton were
29 significantly higher at mining sites (40.3 to 1387 mg/kg dw) compared to reference sites (not
30 detected [ND], 33.3 mg/kg dw). Furthermore, Pb in chironomids was significantly higher at
31 mine-affected sites (1.6 to 131 mg/kg dw) compared to reference sites (ND,10.2 mg/kg dw). The
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1 concentrations in biota indicate that Pb is mobile and available to the aquatic community even
2 though water concentrations were undetectable. Chironomidae richness was reduced at the sites
3 receiving mining effluent containing Pb, Cd, Cu, and Zn.
4 In another study, macroinvertebrate lead tissue concentrations (32.2 to 67.1 mg/kg dw at
5 affected sites) collected from the Clark Fork River, Montana correlated negatively with total
6 richness, EPT richness, and density (Poulton et al., 1995). Mean Pb levels were as high as
7 67.1 mg/kg dw at sites most affected by lead. However, other metals, including Cd, Cu, and Zn,
8 also were negatively correlated with total richness and EPT richness. Therefore, attribution of
9 the observed effects to Pb is difficult, as other metals may be contributing factors.
10 In Montana, the potential effects of metals on macroinvertebrate communities in the
11 Boulder River watershed were evaluated (Rhea et al., 2004). Similar to the approach taken by
12 Poulton et al. (1995), the effects on richness and abundance of EPT taxa were compared to metal
13 concentrations in tissue (i.e., biofilm and macroinvertebrates). Lead levels in biofilm (32 to
14 1540 mg/kg dw) were significantly correlated with habitat scores and macroinvertebrate indices
15 (e.g., EPT taxa). However, macroinvertebrate tissue Pb levels were not significantly correlated
16 with macroinvertebrate community level metrics. As with most natural systems with potential
17 mine impacts, other metals also correlated with community level effects. However, the authors
18 indicated that Pb concentrations in biofilm appeared to have the most significant impact on
19 macroinvertebrate metrics.
20 A detailed investigation of sediment, macroinvertebrates, and fish was conducted for
21 tributaries in the Aquashicola Creek watershed near a former zinc smelter in Palmerton, PA
22 (Carline and Jobsis, 1993). The smelter deposited large amounts of Cd, Cu, Pb, and Zn on the
23 surrounding landscape during its operation from 1898 to 1980. The goal of the study was to
24 evaluate if there was a trend in the metal levels in sediment, macroinvertebrate and fish tissue,
25 and community indices going away from the smelter. Sites were chosen, from 7.8 to 24.6 km
26 from the smelter. There were no clear associations between proximity to the smelter and Pb
27 levels in sediment, macroinvertebrate tissue, and fish tissue. Furthermore, there were no
28 associations between proximity to the smelter and macroinvertebrate and fish diversity and
29 richness. The authors suggested that the transport of metals in the watershed has decreased since
30 the smelter ceased operation, and thereby no effects were observed.
31
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1 Ecosystems and Communities, Community Dynamics, and Trophic Structure
2 As described in the SAB framework, community dynamics include interspecies
3 interactions such as competition, predation, and succession (Young and Sanzone, 2002).
4 Measures of biotic interactions (e.g., levels of seed dispersal, prevalence of disease in
5 populations of focal species) provide important information about community condition. If the
6 community dynamics are disrupted, then the trophic structure may also be disrupted. According
7 to the SAB framework, trophic structure refers to the distribution of species/taxa and functional
8 groups across trophic levels. Measures of trophic structure include food web complexity and the
9 presence/absence of top predators or dominant herbivores. Therefore, this section discusses how
10 aquatic species interactions can be affected by Pb. Examples of species interactions can include:
11 • Predator-prey interactions (e.g., reduced avoidance of predators)
12 • Prey consumption rate (e.g., increase or decrease in feeding)
13 • Species competition (e.g., interference with another species, increased aggressive
14 behavior)
15 • Species tolerance/sensitivity (e.g., the emergence of a dominant species due to
16 contaminant tolerance or sensitivity)
17 Species interactions are highly relevant to a discussion about the effects of Pb on natural
18 aquatic ecosystems, because effects on species interactions could potentially affect ecosystem
19 function and diversity. Some examples of Pb induced changes in species interactions are
20 presented below (see summary in Table AX8-2.5.2).
21
22 Predator-Prey Interactions
23 Lefcort et al. (1999) examined the competitive and predator avoidance behaviors of snails
24 and tadpoles in outdoor mini-ecosystems with sediment from a metals-contaminated Superfund
25 site (i.e., Pb, Zn, Cd). Previous investigations of aquatic invertebrates and vertebrates yielded Pb
26 tissue concentrations of 9 to 3800 mg/kg dw and 0.3 to 55 mg/kg dw, respectively. Several
27 species interactions were studied in the presence of metal-contaminated sediment:
28 Snails and tadpoles have similar dietary behaviors. Thus, when placed in the same habitat
29 they will compete for the same food items and negatively affect one another. However, when
30 tadpoles exposed to a predator (i.e., through biweekly additions of 20 mL of water from tanks
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1 housing sunfish—10 mL from sunfish-fed snails, 10 mL from sunfish-fed tadpoles) were placed
2 with snails, the tadpoles reduced sediment ingestion, while snails increased ingestion. Thus,
3 snails were exposed to greater quantities of metals in sediment.
4 In an uncontaminated environment, snail recruitment (i.e., reproduction) was reduced in
5 the presence of tadpoles. The addition of tadpoles increased the competition for food in the form
6 of floating algae and the snails switched to feeding on algae that grew on the sediment. This
7 decrease was due to competition alone. The effects on snail recruitment were even higher when
8 tadpoles, the influence of a predator (i.e., sunfish extract), and metals in the sediment were all
9 present. However, the predator effect was indirect in that the tadpoles hid in the algal mats
10 forcing the snails to feed primarily on the benthic algae that grew on the sediment with high
11 metal levels. Furthermore, although not significant, Pb levels in snails were higher when
12 tadpoles and sunfish extract were present than when only metals in the sediment were present.
13 Finally, snail predator avoidance was assessed. Snails (control and lead-exposed) were
14 stimulated with a predator indicator (i.e., crushed snails and an extract of crushed snail). Control
15 snails changed behaviors in the presence of the predator indicator, while exposed snails did not
16 alter their behavior. The authors suggested that metal exposure caused behavioral changes that
17 alter competitive interactions and the perception of predators by the snails. Thus, Pb may affect
18 the predator avoidance response of snails.
19 In further study, Lefcort et al. (2000) examined the predator avoidance behaviors of snails
20 and caddisflies. In separate experiments, the avoidance behavior of the snail, Physella
21 columbiana, and four caddisfly genera (Agrypnia, Hydropsyche, Arctopsyche, Neothremmd)
22 were evaluated. The snails were collected from reference lakes and lakes downstream of the
23 Bunker Hill Superfund site. The snails from the affected lakes generally had higher cadmium,
24 Pb, and zinc tissue levels implying previous exposure to these metals. Snail predator avoidance
25 behavior was tested by exposure to crushed snail extract. Snails from the affected lakes did not
26 reduce their activity when exposed to the snail extract, implying a reduced predator avoidance.
27 The lack of response may make the snails at the affected lakes more prone to predation.
28 The caddisflies were evaluated at 36 sites from six different streams. As with the snails,
29 the caddisflies from the affected streams had higher cadmium, Pb and zinc tissue levels. The
30 time for caddisfly larvae to respond (i.e., how long immobile) to disturbance (i.e., lifted from
31 water for 3 seconds and moved to a new location) was evaluated. There was no correlation
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1 between tissue metal level and any response variable (Lefcort et al., 2000). Therefore, the
2 authors concluded that preexposure to metals did not reduce predator avoidance for caddisflies.
3 Weber (1996) examined juvenile fathead minnows exposed to 0, 0.5, or 1.0 ppm Pb in
4 water during a 2-week preexposure and 2-week testing period (4 weeks total exposure). Feeding
5 behavior was evaluated by presenting two prey sizes (2-day-old and 7-day-old Daphnia magnd).
6 Control fish began switching from larger, more difficult-to-capture 7-day-old daphnids to
7 smaller, easier-to-catch 2-day-old prey by day 3. Lead-exposed fish displayed significant
8 switching at day 3 (at 0.5 ppm) or day 10 (at 1.0 ppm). Thus, exposure to Pb delayed the altering
9 of prey size choices to less energetically costly prey.
10 Lefcort et al. (1998) exposed spotted frogs (Rana luteiventris) to 0.05 to 50 ppm Pb in
11 water for 3 weeks. High levels of Pb reduced the fright response of tadpoles; suggesting a
12 reduced avoidance of predators.
13 Bullfrog larvae exposed to Pb in water (0.78 mg/L) and high or low dissolved oxygen
14 were monitored for respiratory surfacing behavior (Rice et al., 1999). Larvae had a significantly
15 increased number of trips to the water surface regardless of oxygen content. Thus, the authors
16 suggest that Pb may affect oxygen uptake such that larvae are under greater predation pressure
17 due to increased time spent at the surface.
18 Weis and Weis (1998) evaluated the effect of Pb exposure on mummichog (Fundulus
19 heteroclitus) larvae prey capture rate, swimming behavior, and predator avoidance. Prey capture
20 rates were affected after 4 weeks exposure at 1.0 mg Pb/L. The larvae were also more
21 vulnerable to predation by grass shrimp (Palaemonetespugio) at 1.0 mg Pb/L. Finally, the
22 swimming behavior of mummichog larvae was affected at 0.3 and 1.0 mg Pb/L. Once the larvae
23 were no longer exposed to Pb, they recovered their ability to capture prey and avoid predators.
24 Clearly, exposure to Pb does affect the predator-prey interactions and the ability of prey to
25 avoid predators. The effect of Pb on these ecological functions may alter community dynamics.
26
27 AX8.2.5.3 Summary
28 The effects of Pb have primarily been studied in instances of point source pollution rather
29 than area-wide atmospheric deposition; thus, the effects of atmospheric Pb on ecological
30 condition remains to be defined. The evaluation of point source Pb within the EPA Ecological
31 Condition Framework has been examined primarily in relation to biotic conditions. The
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1 available literature focuses on studies describing the effects of Pb in natural aquatic ecosystems
2 with regard to community composition and species interactions. The effects of Pb on the biotic
3 condition of natural aquatic systems can be summarized as follows: there is a paucity of data in
4 the general literature that explores the effects of Pb in conjunction with all or several of the
5 various components of ecological condition as defined by the EPA. However, numerous studies
6 are available associating the presence of Pb with effects on biotic conditions.
7 In simulated microcosms or natural systems, environmental exposure to Pb in water and
8 sediment has been shown to affect energy flow and nutrient cycling and benthic community
9 structure. In field studies, Pb contamination has been shown to significantly alter the aquatic
10 environment through bioaccumulation and alterations of community structure and function.
11 Exposure to Pb in laboratory studies and simulated ecosystems may alter species competitive
12 behaviors, predator-prey interactions, and contaminant avoidance behaviors. Alteration of these
13 interactions may have negative effects on species abundance and community structure. In
14 natural aquatic ecosystems, Pb is often found coexisting with other metals and other stressors.
15 Thus, understanding the effects of Pb in natural systems is challenging given that observed
16 effects may be due to cumulative toxicity from multiple stressors.
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