United States
Environmental Protection
Agency
Health EHects Research
Laboratory
Research Triangle Park NC 27711
EPA-6 00/1-78-034
Mny 1978
oEPA
Research and Develooment
Zinc
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RESEARCH REPORTING SERIES
Research reports of the Office of Research and Development, U.S. Environmental
Protection Agency, have been grouped into nine series. These nine broad cate-
gories were established to facilitate further development and application of en-
vironmental technology. Elimination of traditional grouping was consciously
planned to foster technology transfer and a maximum interface in related fields.
The nine series are:
1. Environmental Health Effects Research
2. Environmental Protection Technology
3, Ecological Research
4. Environmental Monitoring
5. Socioeconomic Environmental Studies
6. Scientific and Technical Assessment Reports (STAR)
7. Interagency Energy-Environment Research and Development
8 "Special" Reports
9 Miscellaneous Reports
This report has been assigned to the ENVIRONMENTAL HEALTH EFFECTS RE-
SEARCH series This series describes projects and studies relating to the toler-
ances of man for unhealthful substances or conditions. This work is generally
assessed from a medical viewpoint, including physiological or psychological
studies. In addition to toxicology and other medical specialities, study areas in-
clude biomedical instrumentation and health research techniques utilizing ani-
mals — but always with intended application to human health measures.
This document is available to the public through the National Technical Informa-
tion Service, Springfield, Virginia 22161.
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EPA-600/1-78-03A
May 1978
ZINC
by
Subcommittee on Zinc
Committee on the Medical and Biologic Effects of
Environmental Pollutants
National Research Council
National Academy of Sciences
Washington, D.C.
Contract No. 68-02-1226
Project Officer
Orin Stopinski
Criteria and Special Studies Office
Health Effects Research Laboratory
Research Triangle Park, N.C. 27711
U.S. ENVIRONMENTAL PROTECTION AGENCY
OFFICE OF RESEARCH AND DEVELOPMENT
HEALTH EFFECTS RESEARCH LABORATORY
RESEARCH TRIANGLE PARK, N.C. 27711
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DISCLAIMER
This report has been reviewed by the Health Effects Research
Laboratory, U.S. Environmental Protection Agency, and approved
for publication. Approval does not signify that the contents
necessarily reflect the views and policies of the U.S. Environmental
Protection Agency, nor does mention of trade names or commercial
products consitute endorsement or recommendation for use.
NOTICE
The project that is the subject of this report was approved by the
Governing Board of the National Research Council, whose members are
drawn from the Councils of the National Academy of Sciences, the National
Academy of Engineering, and the Institute of Medicine. The members of
the Committee responsible for the report were chosen for their special
competences and with regard for appropriate balance.
This report has been reviewed by a group other than the authors
according to procedures approved by a Report Review Committee consisting
of members of the National Academy of Sciences, the National Academy of
Engineering, and the Institute of Medicine.
ii
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FOREWORD
The many benefits of our modern, developing, industrial society
are accompanied by certain hazards. Careful assessment of the relative
risk of existing and new man-made environmental hazards is necessary
for the establishment of sound regulatory policy. These regulations
serve to enhance the quality of our environment in order to promote the
public health and welfare and the productive capacity of our Nation's
population.
The Health Effects Research Laboratory, Research Triangle Park,
conducts a coordinated environmental health research program in toxicology,
epidemiology, and clinical studies using human volunteer subjects.
These studies address problems in air pollution, non-ionizing
radiation, environmental carcinogenesis and the toxicology of pesticides
as well as other chemical pollutants. The Laboratory participates in
the development and revision of air quality criteria documents on
pollutants for which national ambient air quality standards exist or
are proposed, provides the data for registration of new pesticides or
proposed suspension of those already in use, conducts research on
hazardous and toxic materials, and is primarily responsible for providing
the health basis for non-ionizing radiation standards. Direct support
to the regulatory function of the Agency is provided in the form of
expert testimony and preparation of affidavits as well as expert advice
to the Administrator to assure the adequacy of health care and surveillance
of persons having suffered imminent and substantial endangerment of
their health.
To aid the Health Effects Research Laboratory to fulfill the functions
listed above, the National Academy of Sciences (NAS) under EPA Contract
No. 68-02-1226 prepares evaluative reports of current knowledge of selected
atmospheric pollutants. These documents serve as background material for
the preparation or revision of criteria documents, scientific and technical
assessment reports, partial bases for EPA decisions and recommendations
for research needs. "Zinc" is one of these reports.
F. G. Hueter, Ph. D.
Acting Director,
Health Effects Research Laboratory
iii
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OH zmc
COMMITTEE OH MEDICAL AND BIOLOGIC EFFECTS OP ENVIRONMENTAL POLLUTANTS
ROBERT I. HKHCIH, G«org«town University Hospital, Washington, D.C.,
Chairman
JEAN APGAR. U. S. Pl*nC, Soil, end Nutrition Laboratory, Ithaca, New York
JEROME T, COLE, International Lead Zinc Research Organization, Inc.,
Saw York, New York
JOSEPH E. COLEMAN, Yale University, New Raven, Connecticut
CARL H. COTTERILL, Bureau of Mines, tj. S. Department of Interior,
Waahington, D.C.
MICHAEL FUISCHXR, U. S. Geological Survey, Beaton, Virginia
ROBERT A. GOTRR, University of Western Ontario Faculty of Medicine, London,
Ontario, Canada
BEKHARD GREIFER, National Bureau of Standard*, Washington, D.C.
BERNARD D. KHBZBX, Michigan State University. East Lanaing, Michigan
PAUL MDSRAK, University of North Carolina School of Medicine, Chapel Hill,
North Carolina
MAGNUS PISCATOR, Karolinsk*. Institute, Stockholm, Sweden
BRUCE R. STILLINGS, Nabisco, Inc., Fairlavn, New Jersey
JOHN K. TAYLOR, National Bureau of Standard*, Washington, D.C.
DOUGLAS A. WOLFS, Environmental Research Laboratory, National Oceanic and
Atmospheric Administration, Boulder, Colorado
*JOHN F. DAVIS, Michigan State University, East Lanaing, Michigan, Consultant
T. D. BOAZ, JR.. Division of Medical Sciences, National Research Council,
Washington, D.C., Staff Officer
REUEL A. STALLONES, School of Public Health, University of Texas,
Houston, Chairman
MARTIN ALEXANDER, Cornell University, Ithaca
ANDREW A. BENSON, Scripps Institution of Oceanography, University of
California, La Jolla
RONALD F. COBURN, University of Pennsylvania School of Medicine, Philadelphia
CLEMENT A. FINCH, University of Washington School of Medicine, Seattle
EVTLLE GORHAM, University of Minnesota, Minneapolis
ROBERT I. HENKIN, Georgetown University Medical Center, Waahington, D.C.
IAN T. T. HICOINS, School of Public Health, University of Michigan, Ann Arbor
JOE W. RIGKTOWER, Rice University, Houston
HENRY KAMIN, Duke University Medical-Center, Durham, North Carolina
ORVILLE A. LEVANDER, Agricultural Research Center, Beltsvllle, Maryland
ROGER P. SMITH, Dartmouth Medical School, Hanover, New Hampshire
T. D. BOAZ, JR., Division of Medical Sciences, National Research Council,
Washington, D.C., Executive Director
•Deceased
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Acknowledgments
This document is the result both of individual and of coordinated
efforts by members of the Subcommittee on Zinc. Although each member
was responsible for a specific section, as detailed below, each re-
viewed the work of the others; Chapter 14, the summary, and Chapter 15,
the recommendations, represent a consensus of the members of the
Subcommittee.
The Introduction was written by Dr. Robert I. Henkin, Chairman of
the Subcommittee. Chapter 2, on properties and uses of zinc, was
written by Mr. Carl H. Cotterill. Dr. Michael Fleischer contributed
Chapter 3, on natural sources and distribution of zinc, except for the
section on zinc reactions with organic matter in soils, which was pre-
pared by Dr. Bernard D. Knezek.
Except for the passage on sewage in the section on waste disposal,
which was written by Dr. Knezek, Chapter 4 was the responsibility of
Dr. Jerome F. Cole. Chapter 5, on zinc in plants, was the work of
Dr. Knezek, who was solely responsible for the section on aquatic plants,
and jointly responsible with Dr. John F. Davis (since deceased) for the
section on terrestrial plants.
Drs. Douglas A. Wolfe and Bruce R. Stillings prepared the account
on zinc in aquatic animals, set forth in Chapter 6. Chapter 7, on zinc
in humans, was written by Dr. Henkin, and Chapter 8, on zinc in the diet
and the effects of zinc deficiency in animals, was written by Dr. Jean
Apgar. Dr. Joseph E. Coleman prepared Chapter 9, on zinc in raetallo-
protelns. Chapter 10, on clinical aspects of zinc metabolism, was tne
work of Dr. Henkin. Three authors were involved in the preparation of
v
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Chapter 11, on toxicity of zinc; Dr. Magnus Piscator wrote two sections,
those on zinc and cadmium and interactions between zinc and cadmium, and
Dr. Apgar wrote the section on animals. Dr. Robert A. Goyer had overall
responsibility for the chapter, and contributed the other sections.
Thanks are due Mr. Ralph C. Wands, former Director of the Advisory
Center on Toxicology, for obtaining the material that appears as stand-
ards for zinc levels in Chapter 12. The section on solid and liquid
samples in Chapter 13 (on sampling and measurement techniques for ana-
lyzing zinc), was written by Dr. Paul Mushak; the other section in this
chapter, on air, was co-authored by Drs. Bernard Greifer and John K.
Taylor. The material in Appendix A, on the zinc content of foods, was
chosen by Dr. Apgar, and that in Appendix B, on methods of zinc ana-
lysis, was chosen by Dr. Mushak.
Ms. Avis Berman edited the -nanuscript and worked tirelessly with
the authors in resolving the many aifficulties that arose during the
preparation of the voluminous typescript. Ms. Joan Stokes checked for
accuracy all references cited, and was responsible for preparation of
the extensive bibliography.
Free use was made of the resources available at the National Library
of Medicine, the National Agricultural Library, the Library of Congress,
and the Air Pollution Technical Information Center of the Environmental
Protection Agency. Also acknowledged is the assistance given to the
Subcommittee by the National Research Council's Advisory Center on
Toxicology, the National Academy of Sciences Library, the Environmental
Studies Board, and various units of the National Research Council.
vi
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CONTENTS
1 Introduction 1
2 Properties and Uses of Zinc 5
3 Natural Sources and Distribution of Zinc 26
4 Man-Made Sources of Zinc 45
5 Zinc in Plants 99
6 Zinc in Aquatic Animals 134
7 Zinc in Humans 186
8 Zinc in the Diet and the Effects of Zinc Deficiency
in Animals 261
9 Zinc in Metalloproteins 320
10 Clinical Aspects of Zinc Metabolism 336
11 Toxicity of Zinc 377
12 Standards for Zinc Levels 407
13 Sampling and Measurement Techniques for Analyzing Zinc 413
14 Summary 451
15 Recommendations 468
Appendix A: Zinc Content of Foods 474
Appendix B: Methods of Zinc Analysis 487
References 499
vii
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CHAPTER 1
INTRODUCTION
Zinc has been used by man for industrial, ornamental, or utilitarian
purposes for nearly 2,000 years, but its history is clouded until well into
the middle ages. Thompson was able to ascribe the capability of making
a compound of copper, tin and zinc by reduction with charcoal to the
Babylonians of 3000 B.C.
Semitic bronzes found at Gezer in Palestine and dated 1400 and 1000
1211
B.C. contained up to 23% zinc as well as 10% tin. Although brass was
known early in Palestine and was probably used for cymbals and bells, zinc
was probably quite unknown. The brass was likely to have been made by smelt-
ing a zinc ore with copper and charcoal.
The oldest known piece of zinc is the form of an idol found in the pre-
historic Dacian settlement at Dordosch, Transylvania. Analysis of the idol
showed that it was composed of 88% zinc, 11% lead, and 1% iron. In the
ruins of Camiros, destroyed in 500 B.C., two bracelets filled with zinc were
found? in the ruins of Pompeii, destroyed in 79 A.D., the upper part
of a fountain front was discovered, and the finders claimed that it had been
covered with zinc.
The Greeks may also have known about and used zinc. Aristotle was said
to have spoken about a brilliant white copper produced by adding "some kind
1042
of earth" rather than melting tin with copper. Theophrastus, a contemporary
of Aristotle, also mentioned an ore which became superior in beauty and color
1042
when mixed with copper. Strabo (60 B.C.-20 A.D.) described an ore "which
when burned becomes iron and when heated in furnaces with a certain earth
distils *mock silver1 (pseudargyros), and this with the addition of copper
makes the 'mixture* named oreichalkos." Mitchell concluded that
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pseudargyros was zinc and the method of smelting through distillation was
one which would produce zinc. Other historians are not Convinced that
pseudargyros was metallic zinc and suggest that it might have been metallic
. , 1025
arsenic or arsenical copper.
It is not clear when zinc was introduced as a useful agent in the Far
East. However, useful brass products were made by the calamine* process
53
certainly since the beginning of the Christian era, and perhaps even before
12
it. Li suggested that zinc was used by the early Chinese by about the
264
beginning of the Christian era. Forbes speculated that zinc metal was pre-
pared by Indian alchemists in the laboratory in the twelfth century, but the
489
process was not applied industrially. The Chinese also were said to know
how to make a metal from tutty scraped from the sides of smelting furnaces
making lead. The substance was thought to be cadmium, but it was mostly zinc
•A 37°
oxide.
The Romans were probably the first Europeans to make brass when they began
to use it for coins (about 20 B.C.). They discovered that when copper was
smelted with calamine ore, a yellow alloy more gold than bronze could be
718
obtained.
In its isolated form, zinc was not recognized until the fifteenth
century when smelting probably occurred accidentally.12 The word "zinc"
may be derived from the German noun Zinker, meaning "jagged part or tooth."
This word was applicable to the metal because in early smelting endeavors,
*
A zinc ore.
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zinc was deposited in the furnace in the form of pointed parts. Upon slow
cooling, zinc vapor will condense and form clusters of hexagonal crystals
with pointed ends. "Spelter" was also a name given to zinc and it is related
to the word "pewter." The earliest occurrence of the word "spelter" was in 1661.
Commercial smelting began in the eighteenth century when it was clearly
realized that zinc could be obtained from the calamine used to make brass and
that it was the same metal imported from India and China.
No reports of zinc toxicosis in any form were forthcoming from these early
accounts. However, it has been alleged that zinc was applied as an ointment for
skin lesions by several cultures of the ancient world, including the Egyptians and
other Mediterranean peoples. The first documented usage of zinc administered
554
orally occurred in 1826 when zinc sulfate was used to treat gleet and leucorrhea.
The purpose of this document is to attempt to place into perspective the
role of zinc in the environment — its importance as an essential nutrient
for all forms of life and its effects as a toxic agent to some species. The
effects of zinc are many sided. Because zinc is abundantly distributed throughout
the earth's crust and found in many manufactured products, humans come into direct
daily contact with several forms of zinc. Zinc is used to manufacture
motor oils, lubricants, and rubber tires, and it is found in the fuel oil
and coal used for heating and manufacturing purposes; thus, particulate
zinc is present in the atmosphere in rural as well as urban areas.
This particulate zinc has not been specifically identified as a public
health problem. Zinc is essential for the normal activity of DNA polymerase; and
hence, for protein synthesis. It plays an important role in the growth, development,
and metabolism of living species* In plants and animals the balance between excess
and insufficient zinc is important. Plants do not grow well in zinc-depleted or
zinc-absent soils, and insufficient dietary zinc in animals and humans leads to poor
-------
development and other pathophysiologic changes. Dietary zinc
replacement usually will reverse the pathologic events of zinc
depletion in man and animals, but important exceptions do occur.
In rat offspring with congenital malfunctions or
behavioral abnormalities associated with zinc depletion, repletion
with zinc seems to be of little value. In humans, a major problem
is the diagnosis and evaluation of zinc deficiency, particularly
if the case is marginal.
Excessive zinc in the aquatic environment is of particular importance,
because the respiratory systems of fish are very sensitive to the
toxic effects of zinc. Water represents a convenient vehicle for
disposal of human and animal waste products, which can be quite high
in zinc. The toxic effects of zinc in humans are not common
medical problems, although they may appear in some metal workers
and in some children under special conditions. There is increasing
evidence that zinc may be helpful in the treatment of several disorders
of man and animals, including skin lesions and the rare disease
acrodermatitis enteropathica. However, knowledge of zinc metabolism
in humans is still quite limited.
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CHAPTER 2
PROPERTIES AND USES OF ZINC*
Among the major common metals (iron, aluminum, copper, lead and zinc),
all but zinc are easily recognized as such by the consumer. This distinc-
tion is a function of its loss of identity to an end product. Zinc has
been found to be beneficial and necessary for proper nutrition of humans,
animals, and plants, and a deficient intake has been proved deleterious.
Zinc is a bluish-white, relatively soft metal with a density slightly
less than iron (7.133 and 7.86 g/cc, respectively). Its atomic number is
30, atomic weight, 65.37, and it is placed in Group II-B of the periodic
table. The atomic radius of zinc is 1.31 A and its electron configuration
is 2-8-18-2. Zinc is divalent in all its compounds. It is a composite of
five stable isotopes: zinc-64, -66, -67, -68, and -70; measured by
relative abundance, these constitute 48.86%, 27.62%, 4.12%, 18.71%, and
0.69%, respectively, of the whole. Six radioactive isotopes have been
identified: zinc-62, -63, -65, -69, -72, and -73.
The most commonly used artificial isotopes are zinc-65 and zinc-69,
which have half-lives of about 244 days for the zinc-65, 14 h for the zinc-69
isomer, and 58 min for zinc-69 itself. Decay products of zinc-65 and -69
are stable copper-65, manufactured by positron emission and electron
capture from zinc-65, and stable gallium-69 isotopes, produced by negative
*Much of this chapter is derived from the U.S. Department of Interior,
Bureau of Mines publication, The U.S. Zinc Industry: A Historical Perspective
(.1974). 101*> No other references will be provided in the body of the chapter
unless the information comes directly from another source.
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B-emission from the zinc-69 Isotopes. Other characteristics of zinc
isotopes may be found in Weast et a.1.
The structure of the zinc molecule exhibits a hexagonal close-packed
lattice: in A, when a = 2.664 and o = 4.9469, a/o - 1.856.
PHYSICAL PROPERTIES OF ZINC
Cast zinc, which crystallizes in a hexagonal system, is brittle; but
when heated to about 120 C, it becomes ductile and is easily rolled
or drawn. After mechanical shaping at about this temperature,
the metal does not become brittle again upon cooling. Thus, wrought
zinc is used in construction to form roofing and as drawn battery cans for
dry power cells. The high electrochemical activity of zinc is surpassed
among the common metals only by magnesium and aluminum in the electro-
motive series. This activity accounts for one of its major uses: the
galvanizing of iron and steel. In such applications, zinc undergoes
sacrificial corrosion from the surface of the steel, and protects the
substrate from structural degradation. Zinc readily combines with other
metals, imparting characteristics of workability at low temperature, cor-
rosion resistance, and pleasing finishes for use in die-casting alloys,
brass, and other common alloys. Table 2-1 sets forth some physical
properties of zinc.
CHEMICAL PROPERTIES OF ZINC
Pure zinc at ambient temperatures is highly resistant to attack by
dry air, but in temperatures above 225 C the rate of corrosion increases
rapidly. In moist air, attack proceeds at room temperature, and in the
presence of carbon dioxide it accelerates to form the hydrated basic
carbonate, 2ZnC03'3Zn(OH)2' This carbonate
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TABLE 2-1
Some Physical Properties of Zinc
Density:
solid at 25 C, 7.133 g/cm3
solid at 419.5 C, 6.83 g/cm3
liquid at 419.5 C, 6.62 g/cm3
liquid at 800 C, 6.25 g/cm3
Melting point: 419.5 C (692.7 K)
Boiling point (1 atm) : 907 C (1,180 K)
Heat capacity:
solid - Cp = 5.35 f 2.40 x 10~3 T (298 - 692.7 K) cal/mol
liquid - Cp = 7.50 cal/mol
gas (monatomic) - Cp = 4.969 cal/mol
Heat of fusion: 1,765 cal/mol at 419.5 C
Heat of vaporization: 27,430 cal/mol at 907 C
Linear coefficients of thermal expansion!
polycrystalline (20 - 250 C) , 39.7 x 10~6 per C
Volume coefficient of thermal expansion (20 - 400 C) : 8.9 x 10~5 per C
Thermal conductivity:
solid (18 C) 0.27 cal/sec cm C
solid (419.5 C) 0.23 cal/sec cm C
liquid (419.5 C) 0.145 cal/sec cm C
liquid (750 C) 0.135 cal/sec cm C
Modulus of elasticity:
10 to 20 x 106 psi
Surface tension (liquid): Y = 758 - 0.09(t - 419.5 C) dynes/cm
Electrical resistivity:
polycrystalline (t = 0-100 C) R = 5.46(1 + 0.0042t) microhms /cm3
liquid C423 C) 36.955 microhms/cm3
Magnetic susceptibility (diaraagnetic) :
polycrystalline (20 C) - 0.139 x 10~6 cgs electromagnetic units
a Modified from Schuhmann and
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forms a tightly adhering light gray film which tends to protect the zinc
from further corrosion. Halogens react with zinc in the presence of
moisture, but not under dry conditions. Reaction with steam at 350 C or
higher occurs rapidly. Mineral acids also easily attack zinc: the
strongest reaction occurs with sulfuric acid, followed by hydro-
chloric., and nitric acids. Zinc displays a vigorous reducing
power, liberating hydrogen from sulfuric and hydrochloric acids. This
property is the basis for the use of zinc dust or mossy zinc in many commercial
organic chemical processes* . Zinc liberates nitrogen oxides instead of
hydrogen from nitric acid. The metal is amphoteric, reacting with hot
caustic to liberate hydrogen and form zincates. All zinc is inert to
petroleum derivatives and anhydrous alcohol but is- oxidized by
mixtures of alcohol and water. The chemical properties of zinc compounds
make them useful as oxides, carbonates, sulfates, sulfides, chlorides,
phosphates, and organic complexes. Commercial grades of zinc contain
enough trace impurities to make it more reactive than pure zinc.
ZINC METAL AND ALLOYS
Commercial grades of zinc metal have been established by the American
Society of Testing Materials (ASTM)y as listed in Table 2-2.
TABLE 2-2
Grades of Slab Zinc and Chemical Requirements^ "
Grade
Cnor^ al tM o\\ — C-raeto^— .
TH ah CfaAo— — — — —
Prime Western
Lead,
maximum
Onni
On?
0?n
Ofin
1.60
Composition,
Iron,
maximum
0.003
0.02
0.03
0.03
0.05
7.
Cadmium,
maximum
0.003
0.03
0.40
0.50
0.50
Zinc, minimum ••
by difference
99.990
99.90
99.5
99.0
98.0
aData from ASTM standards, specification B6-70.30a
^When specified for use in the manufacture of rolled zinc or brass,
aluminum shall not exceed 0.005%.
°T±n in Special High-Grade shall not exceed 0.001%.
8
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Prime Western was the specification earliest established for use in
hot-dip galvanizing. Brass Special and Intermediate were largely used in
alloying with copper to form brass. High-Grade, and eventually Special-
High-Grade specifications were established for zinc to be used in alloys
containing small-amounts of aluminum. These alloys are us.ed in the die-
. cast ing method
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of the coating shows first a layer of pure iron overlaid by a very thin
layer of FeZn3, over which is a thicker layer of FeZn7, above which is a
layer of essentially pure zinc with a few solid crystals of the FeZny.
These various alloys are quite resistant to corrosion and form an adherent
bond between the steel base and the zinc. If the galvanized sheet is to
be subsequently bent and formed, it is necessary to minimize the alloy
formation to avoid cracking and peeling.
Tin. Tin is deleterious in very small quantities in rolled zinc, because
it causes ruptures during hot rolling. In castings it also promotes
subsurface corrosion. However, small amounts of tin will cause an
esthetically desirable "spangle" on galvanized sheets.
Copper . Copper increases strength, hardness, creep resistance, and
recrystallization temperature. It may bring on corrosion of zinc in dry cells.
ii. At levels of 3.5-4.5%, aluminum reduces grain size, and improves
impact strength and castability of zinc. In galvanizing, die casting, and
protective galvanic anodes, aluminum beneficially inhibits formation of the
zinc-iron alloy.
Magnesium. Magnesium counteracts subsurface corrosion effects of tin and
lead in zinc alloys.
Titanium. Titanium forms a compound rich in zinc. At elevated temperatures,
it decreases the grain size of cast zinc and restrains grain growth in rolled
zinc. Used in newer rolled zinc applications, it greatly increases creep
resistance.
One of the chief uses for high-purity zinc is in zinc-base alloys for
die-casting. The die-casting process enables a dimensionally accurate equipment
part to be produced in a die-casting machine in fractions of a minute.
Therefore, few finishing operations are required before use, and the economic
advantages are obvious. Table 2-3 reports the chemical and physical
properties of die-cast alloys.
10
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TABLE 2-3
Chemical and Physical Properties for
Zinc Die-Casting Alloysa
Chemical
'Requirements
as Ingot*7
Copper, %.
Aluminum, %
Magnesium, %
Iron, % (max)
Lead , % (max)
Cadmium , % (max)
Tin, % (max)
Nickel
Zinc
Chemical
Requirements
as Alloy
Die-castings"
Copper, %
Aluminum , %
Magnesium, %
Iron, % (max)
Lead , % (max)
Cadmium, % (max)
Tin, % (max)
Nickel
Zinc
Alloy #3
(ASTM AG40A)
0.10 (max)
3.9-4.3
0.03-0.06
0.075 '
0.005
0.004
0.002
remainder
Alloy #3
(ASTM AG40A)
0.25 (max)
3.5-4.3
0.03-0.08
0.100
0.007
0.005
0.005
r eraainder
Alloy #5
(ASTM AC41A)
0.75-1.25
3.9-4.3
0.03-0.06
0.075
0.005
0.004
0.002
remainder
Alloy #5
(ASTM AC41A)
0.75-1.25
3.5-4.3
0.03-0.08
0.100
0.007
0.005
0.005
remainder
Alloy #7&
0.10 (max)
3.9-4.3
0.005-0.020
0.050
0.0020
0.0020
0.0010
0.005-0.020
remainder
Alloy #7
0.25 (max)
3.5-4.3
0.005-0.020
0.75
0.0030
0.0020
0.0010
0.005-0.020
remainder
a Alloys are numbered 3, 5, and 7'in ordinary trade practice.
b-Alloy 7 has not been assigned an ASTM number.
0ASTM standards, specification B240-64.
"ASTM standards, specification B86-71.
-------
TABLE 2-3 continued
Typical Mechanical Properties for Die-Casting Alloys
Charpy test impact strength, ft-lb, 1/4 x 1/4-in. (.625 x .625 cm)
bar, as cast
Charpy test impact strength, ft-lb, 1/4 x. 1/4-in.
bar, after 10 yrs indoor aging
Tensile strength, psi, as cast
Tensile strength, psi, after 10 yrs aging .
Elongation, % 2 in. (5 cm), as cast
Elongation, % 2 in. after 10 yrs aging
Expansion (growth) in. /in. (2.5 cm/2.5 cm) after 10 yrs aging at
room temperature
Brinell hardness
Alloy #3
(ASTM AG40A)
43
41
41,000
35,000
10
16
0.00008
82
Alloy #5
(ASTM AC41A)
48
40
47,600
39 ,'300
7
13
0.00007
91
Alloy #7
40
41,000
14
76
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Die-cast alloys are used widely in producing automobile parts such as
carburetors, grills, door handles, and ornaments; appliance control panels,
home washer parts television bezels, transistor radio and camera frames;
and small control gears and many other devices. A zinc-containing alloy
in growing use is an aluminum-based die-casting alloy incorporating 2.7-8%
zinc. Production with this alloy requires higher die temperatures and slower
operating speeds than necessary for the zinc-base alloy, but parts made of
it exhibit tensile strengths up to 80,000 psi. Many solders, especially
those used on aluminum, contain zinc as well as other alloying metals.
Silver and gold solders also are alloyed with zinc.
Zinc is a minor component of the copper-zinc alloys known as brass
(although brass is the oldest known use of zinc). Commercial brasses con-
tain between 5-40% zinc. Brass is superior to copper alone in its greater
strength and ductility, and its resistance to corrosion. It is widely
used in hardware, plumbing accessories, instruments, communication equip-
ment, as well as for aesthetic purposes because of its pleasing yellowish-
gold color. Table 2-4 lists composition and properties of some of the more
popular commercial brasses.
Wrought zinc is composed of commercial grades of zinc to which very
small amounts of other metals have been added to alter the properties of
the zinc. Important properties of these wrought zinc alloys designed for
commercial use include: resistance to corrosion; white and nonstaining
corrosion products; chemical characteristics desirable for dry cells and
photoengraving plates; mechanical properties for easy forming, machining,
and spinning; and good solderability. Table 2-5 lists several of these zinc
alloys, and their composition, characteristics and uses.
13
-------
TABLE 2-4
Properties of Common Brasses0
Nominal Yield
Composition, % Strength,
Material copper zinc other Condition 10 psi
Gilding metal
Commercial bronze
Red brass
Aluminum brass
Admiralty brass
Cartridge brass
Yellow brass
Nickel silver
Muntz metal
Manganese bronze
Architectural bronze
a From the
95
90
85
76
71
70
65
65
60
58.5
57
5
10
15
22
28
30
35
17
40
39.
40
Chemical
Cold-rolled
Cold-rolled
Cold-drawn
2 Annealed
1° Annealed
- Cold-rolled
Cold-drawn
18d Cold-rolled,
hard temper
- Annealed
2 l,l,0.3e Cold-rolled
3' Annealed
Engineers' Handbook.1234
50
54
55
27
20
63
55
70
20
50
20
Tensile
Strength,
103 psi
56
61
70
60
53
76
70
85
54
80
60
Brinell
Hardness
114
125
120
82
60
155
115
170
80
180
95
Density,
g/cc
8.90
8.80
8.75
8.33
8.53
8.47
8.47
8.75
8.39
8.36
8.47
Modulus of
Elasticity,
106 psi
17
17
17
16
16
16
15
18
15
15
14
b
0 Tin
d Nickel
e Iron, tin, and manganese, respectively.
/ Lead
-------
TABLE 2-5
Classification of Wrought Zinc Alloys'2
Composition %
Lead
0.05-
0.10
0.05-
0.10
0.15-
0.35
0.05-
0.10
0.05-
0.10
0.005-
0.10
0.15-
0.35
0.007
max
a From
o Manean
Iron
0.012
max
0.012
max
0.017
max
0.012
max
0.015
max
0.012
max
0.014-
0.025
0.10
max
Cadmiun
0.005
max
0.06
0.15-
0.30
0.005
max
0.005
may
0.05
max
0.15-
0.30
0.007
max
the Metals
ese.
Copper
0.001
max
0.005
max
0.005
max
0.85-
1.25
0.85-
1.25
0.50-
1.50
0.005
max
0 -
3.5
3.
Other
-
~*
~™
""
0.007-
0.02*
0.12-
1.50°
0.005-
0.025*
0.02-
0.10*1
5-4. 5d
Handbook. 959
•
Characteristics
High ductility with low hardness
and stiffness. Very little work
hardening possible.
High ductility with low hardness.
Can be work hardened slightly.
High hardness and stiffness. Uni-
form etching quality. Can be
work hardened.
High hardness and stiffness. Good
ductility. Good creep resistance.
Work hardens easily.
High stiffness and creep resistance.
Can be severely work hardened.
Outstanding creep resistance. Can
be severely work hardened. Lowest
thermal expansivity with the grain.
Very high resistance to grain
growth during annealing.
High hardness. Can be baked without
serious softening. Good etching
characteristics.
High strength and hardness.
0 Titanium.
" Aluminum.
Typical use
Drawn battery cans, eyelets, fuse links,
and many other industrial articles drawn,
formed, and spun.
Drawn battery cans, eyelets and grommets.
Extruded battery cans. Address plates,
laundry tags.
Photo engraver's plates, lithographer
sheets, boiler and ship plates, weather-
strips.
Weatherstrips and drawn and formed
industrial articles requiring stiffness.
Flat or formed commercial articles
requiring high stiffness and strength.
Corrugated roofing, leaders and gutters,
and other uses requiring maximum creep
resistance.
Photoengraver's sheets.
Shearing and forming dies. Extruded rods,
tubings and moldings.
-------
of zinc
Although not a strict alloying function, the enlistment /as galvanizing
to coat iron and steel objects is a major use of the metal. As mentioned,
dipping steel in a bath of molten zinc results in a coating of 1-2 oz zinc/
*\
ft2 (311.1-622.2 g/m ) of steel surface. This coating galvanically protects
the underlying steel from corrosion because of the higher electromotive
potential of zinc over iron. Thus the zinc coating must be entirely corroded
away before the iron substrate begins to rust, and the structural strength
of the article is preserved by the sacrificial corrosion of zinc. Steel
of construction
dipped in zinc is the material /for galvanized garbage cans, barbed and woven
wire fencing, steel woven cable, highway guard rail, radio and electricity
transmission towers, and building structures and bridges. Recent advances
control of the
in continuous-line galvanizing have made possible better/composition of
minor alloying metals in the zinc bath possible. The newer processes are
able to provide superior, more economical coatings on continuous steel
sheet made for protective building siding, downspout and guttering, air
ducts, and automobile and appliance bodies.
INORGANIC ZINC COMPOUNDS
Because of its high reactivity with other elements and amphoteric
character, zinc forms a wide variety of compounds. Zinc sulfates and chlorides
are water soluble, whereas the oxides, carbonates, phosphates, silicates,
and organic complexes generally are insoluble. Properties of common zinc
compounds are tabulated in Table 2-6. In the following discussions of zinc
compounds, substances will be listed in generally descending order of
commercial volume.
16
-------
TABLE 2-6
Properties of Common Inorganic Zinc Compounds'2
Zinc Compound
Oxide
Sulfide
(sphalerite)
Sulfate
Chloride
Ammonium
chloride
Fluoride
Bromide
Iodide
^Acetate
Borate
Carbonate
Chromate
Dichromate
Cyanide
Nitrate
Orthophosphate
Orthosilicate
Fluosilicate
Formula
ZnO
ZnS
ZnSOif
ZnCl2
ZnCl2.2NHttCl
ZnF2
ZnBra
Znl2
Zn(C2H302>2
3Zn0.2B203
ZnC03
ZnCrOif
ZnCr207.3H,0
Zn(CN)2
Zn(N03),
ZnaCPOj,,
Z^SiO^
ZnSiF6.6H20
Molecular Crystal Refractive
Weight Form Indexes
81.37
97.43
161.43
136.28
243.26
103.37
225.19
319.18
183.46
383.35
125.39
181.36
335.40
117.41
189.40
386.05
222.82
315.54
Hexagonal 2.008, 2.029
Cubic 2.368
Orthorhombic 1.658, 1.669
Trigonal 1.681, 1.713
-
Tetragonal
Trigonal
Trigonal
Monoclinic -
Triclinic
Trigonal 1.818, 1.618
Orthorhombic
- -
Cubic 1.470
-
Monoclinic
Trigonal 1.694, 1.723
Trigonal 1.382, 1.396
Specific
Gravity
5.606
4.102
3.54
2.91
1.8
4.95
4.01
4.7364
1.84
4.22
4.398
3.40
-
1.852
-
3.998
4.103
2.104
Melting
Point »
C
1,975
—
600
283
~
872
394
446
200
980
-
-
-
800
-
900
1,509
Boiling
Point.
C Remarks
-
Transition point @ 1,020 C
"•
732 Forms hydrates
~
1,500
650
624
-
-
Evolves carbon dioxide @ 300-
500 C
-
- Hygroscopic
-
- Unstable; forms hydrates
- Forms hydrate and complex
-
Decomposes in water
aData from the Handbook of Chemistry and Physics.17*0
-------
Zinc Oxide
Zinc oxide, one of the most valued zinc compounds, crystallizes in a
white hexagonal form. Combustion control during formation can modify the
particle size and shape to achieve desired properties. A variety of particle
shapes and wide size ranges combine with the large interstitial spaces of the
compound to provide a kaleidoscope of properties to be manipulated. Zinc
oxide is used most in the compounding and vulcanizing of rubber, wherein
the properties of high heat capacity and conductivity serve to cool flexing
rubber in belts and tires. This compound scavenges any free sulfur remain-
ing in the article after the rubber is vulcanized.
The high index of refraction of zinc oxide accounts for its use as a pig-
ment in white paints to bestow high hiding power. For exterior paints, the
ability of a thin film of zinc oxide to completely absorb ultraviolet rays
from the sun is useful; it also acts as an effective mildewcide and prevents
fungal staining.
Zinc oxide commonly is classified by method of production. American
Process zinc oxide is made by carbon reduction of roasted zinc ore, and
the resultant zinc vapor is burned to form zinc oxide; French Process zinc
oxide is produced by burning zinc metal in air; some chemical process
oxide is made by precipitating zinc hydroxide (Zn[OH]2) from solution and
calcining it to form oxide.1607c ASTM specifications and chemical and
physical test values for typical grades of zinc oxide are set forth in
Table 2-7, although it should be noted that many producers manufacture
many more grades than are listed to accommodate certain specific needs and
uses.
18
-------
TABLE 2-7
ASTM Specifications and Physical and Chemical Test Values for Zinc Oxide
A. ASTM Specification for Zinc Oxidea
Zinc oxide, range or minimum %
Total sulfur, maximum %
Moisture and other volatile matter, maximum %
Total impurities, including moisture and other volatile
matter, maximum %
Coarse particles (total residue retained on a
No. 325 [44 urn] sieve , maximum %)
Zinc
American
Process
98
0.2
0.5
2.0
1.0
Oxide
French
Process
99
0.1
0.5
1.0
1.0
ASTM standards, specification D79-44 (reapproved 1974).
31
B. Chemical and Physical Property Test Values for Representative Grades of Zinc Oxide"
Chemical tests, in %
Zinc Oxide
Lead
Cadmium
Manganese
Copper
Chlorine
Acidity as sulfur trioxide
Total sulfur as sulfur trioxide
Water-soluble salts
Insoluble in hydrochloric acid
Loss at 110 C
Physical tests
Average numerical diameter, 1 um
Average surface diameter, 3 ym
Specific surface, m2/g
% fines under .50 ym
% through 325-mesh/in2(6.5 cm2) screen
Specific gravity ,
Apparent density, Ib/ft (kg/m3)
Oil absorption of zinc oxides, g oil/ 100 g
zinc oxide
Rubber
Grade
99.20
0.03
0.01
0.003
0.002
-
0.03
0.05
0.15
0.15
0.30
0.28
0.65
1.69
16.0
99.97
5.65
30.0 (220$
Paint
Grade
99.20
0.03
0.01
-
-
0.02
0.15
0.20
0.20
0.15
0.30
0.26
0.60
1.83
18.0
99.92
5.65
30.0 (220$
14.5
A "typical" chemical and physical analysis of one company's particular grades
of rubber and paint zinc oxides.
19
-------
Zinc oxide is insoluble in water, organic solvents, and neutral oils.
With organic and inorganic acids, it forms simple and complex salts and
soaps. Reaction with alkalies form zincates, because zinc is amphoteric.
The oxide may be used to catalyze some chemical reactions. One of the
oldest uses is pharmaceutical — in treatment of burns, infections, and skin
diseases. A newer use for zinc oxide, which relies on physicochemical
properties, was taken up by the photoconductivity field: carefully pro-
duced zinc oxides are coated on paper and used in office photocopying appli-
cations. The ceramic industry produces frits and glazes in which zinc oxide
provides color for pottery or improves the brilliance of glass.
Other Inorganic Zinc Compounds
Zinc sulfate. Zinc sulfate* ZnSO^ is water-soluble and useful as a hard-
ener in viscose rayon spinning baths, and as a flotation reagent in mineral
concentrations. It has become increasingly popular as a trace element
applied to overcome zinc deficiencies in plants grown in certain areas.
Zinc chloride. Zinc chloride* ZnCl2 has a low melting point and is used
in galvanizing fluxes, and in preserving and controlling inflammability in
wood. It is also an essential ingredient in dry cells, a disinfectant, a
printing mordant, and an aid for mercerizing cotton.
Zinc sulfide. Zinc sulfide^ ZnS. is usually a component of barium litho-
pone, a white paint pigment. Very pure zinc sulfide enjoys widespread use
as a phosphor in cathode-ray television tubes and fluorescent lamps.
Zinc chromate. Zinc chromate^ ZnCrO^ is a wood preservative, an algicide,
and a primer on metal surfaces for protection against corrosion.
Zinc carbonate. Zinc carbonate^ ZnC03j like zinc oxide, is now being
used more and more as a nutritive supplement for swine, sheep and
poultry.
20
-------
Zinc borates. Zinc berates, 3Zn0.2B203j are used as fire retardants and as
fluoborates for insecticides.
Zinc acetate. Zinc acetate. Zn(C2H302).2 chiefly provides mordant for dye-
ing and glazes for porcelain, but it is also an astringent and antiseptic.
Zinc silicate. Anhydrous zinc silicate, Zn^iO^, is used as a phosphor in
television screens.
Zinc fluosilicate. Zinc fluosilicate, ZnSiFg.6H20, has continued to be an
effective laundry sour, concrete hardener, and wood preservative.
Zinc cyanide. Zinc cyanide, Zn(CN)2» serves as zinc carrier in electroplating, and
it is used for medicinal purposes to treat epilepsy, neuralgia, etc. It is
poisonous, and will evolve hydrogen cyanide, HCN> gas if contacted by
mineral acids.
Zinc nitrate. Zinc nitrate, Zn(N03)2, finds use as a mordant in dyeing.
Zinc phosphate. Zinc phosphate, .ZngCPO^.AH^O, mostly is used in dental
cements.
Zinc phosphide. Zinc phosphide, Zn.P-, is a common ingredient in rat and
mouse poisons. It is dangerous to humans if highly toxic phosphltie gas
liberated.
Zinc permanganate. Zinc permanganate, ZnpfoOif)2.6H20, finds some applica-
tion as an antiseptic and astringent.
Zinc peroxide. The powdered form of zinc peroxide, Zn02, functions as a
deodorant, astringent, and antiseptic for wounds and skin diseases.
ORGANIC ZINC COMPOUNDS
Most organic derivatives of zinc are manufactured in very small com-
mercial quantities, whereas many of the inorganic compounds are sold in
lots of thousands of tons. As in the previous section, the list of
compounds is in approximate descending order of commercial volume.
21
-------
Zinc soaps. Zinc soaps—stearates, Zn(c18H3502^2' Palmitates»
and oleates, Zn(C1gH3302)2--are fatty acid salts. They are fine,
bulky, soft, white powders that feel greasy, repel water, and generally are
soluble in benzene and petroleum derivatives. They are used as
lubricants and mold release compounds in rubber and plastic forming, metal
die-casting, and medicine tablet making. Zinc soaps also waterproof con-
crete, paper, and textiles, and serve as flatting agents in lacquers.
Zineb and ziram. Zineb (ethylenebisJ Cj^HgN^Zn) and ziram (.dimethyldithio-
carbamate; CeH^NaS^Zn) are popular agricultural fungicides, because they
are more tolerable to humans than are the mercury, lead, and copper fungi-
cides which the zinc salts replace. Some plants even reap nutritional
value from the zinc.
Zinc bacitracin. Zinc bacitracin is an antibiotic in ointments and prepara-
tions for human use and a growth stimulator in swine and poultry nutrition.
Zinc propionate and caprylate. Zinc propionate, Zn(C~He02)2, and caprylate,
Zn(C0IL C00) , serve as fungicides on adhesive tape coatings, and prepara-
8 15 2 2
tions which combat athlete's foot and other molds, fungi, and bacteria.
Zinc phenolsulfonate. Zinc phenolsulfonate, Z9(ei2H,0OgS2»8H20), is both an
insecticide and an antiseptic for internal treatment of ulcers and wounds.
Zinc salicylate. Zinc salicylate, (C H-0 ) Zn.3H 0, is an effective
astringent and antiseptic.
Many other organic salts are used in medicinal applications because
small quantities of zinc are toxic to many microorganisms harmful to human,
animal, and plant life.
Zinc undecylenate. Zinc undecylenate, c22^38°4Zn' is used as a cutaneous
fungicide to control dermatophytoses.
22
-------
PATTERNS OF USE
In the United States, the use of zinc in all forms has increased at a
rate approximating that of the real growth of the gross national product.
The pattern, of use has shifted from one in which brass and galvanizing were
predominant to one in which a much larger share is constituted by zinc-base
alloys for die-casting applications. Consequently, the proportion of zinc
used for brass, wrought zinc, and other purposes has decreased. Table 2-8
and Figure 2-1 chart some of these trends from 1935-1976. During these
years, the total zinc usage in the United States has more than tripled from
566,000 to 1,753,000 metric tons in 1973. This total for
1973 includes the most frequently used statistic of "slab zinc" consump-
tion (1,364,000 metric tons), plus the zinc content of ores used directly
without going through the cast metal or slab stage (118,000 metric tons), as
well as the recoverable zinc content of alloys and chemical compounds pro-
duced from old and new scrap, residues, etc. (271,000 metric tons).
During the period charted, the four major uses of zinc appear
to have been at or near their peak in 1973 (except for brass, used
heavily during World War II for cartridge cases). These large applications
of zinc are very sensitive to the economic cycle (note 1975 data), because
zinc is important in automobile production (die-casting, galvanizing, and
oxide for rubber tires) and industrial and residential construction
activity (galvanizing, brass, die-casting for appliances, and oxide for
paint pigments).
23
-------
TABLE 2-8
Total Zinc Usage in the United States
Classified by Industry, at Intervals between 1935 and
1976a
Metric Tons
Year
1935°
1940
1945
1950
1955
1960
1965
1970
1973
1975
1976d
Zinc Base
Alloy
58,468
137,639
125,847
277,973
413,367
322,576
602,386
444,535
576,493
312,516
460,900
Galvanizing
176,901
300,464
305,886
400,691
409,268
337,100
437,645
430,232
511,504
341,906
390,600
Brass
139,706
249,403
441,013
272,850
270,790
187,284
258,467
262,904
343,210
236,172
256,900
Oxide
90,598
105,677
125,768
140,478
126,633
101,542
145,388
158,572
189,950
120,048
123,000
Rolled
Zinc5
51,256
54,608
88,531
62,091
46,801
35,104
41,623
37,254
36,980
24,773
27,200
Other
49,318
58,818
77,087
71,071
65,868
67,765
94,868
92,231
94,476
82,069
35,400
TOTAL
566,247
906,609
1,164,132
1,225,154
1,332,727
1,051,371
1,580,377
1,425,728
1,752,613
1,117,484
1,294,000
a Data from U.S. Department of Interior, Bureau of Mines.
b Included in "Other" on Figure 2-1.
a Partially estimated.
d Estimated.
24
-------
2,000 i—
1935 1940 1945 1950 1955 1960 1965
YEAR
1970 | 1976
1973
TTTrinHT 2-1 Total zinc usage in the United States by industry at selected
internals, 1935-1976? Compiled from data of U.S. Department of Interior,
Bureau of Mines.
-------
CHAPTER 3
NATURAL SOURCES AND DISTRIBUTION OF ZINC
Zinc is a moderately abundant element; its concentration
in the continental crust of the earth is generally given as 70
ppm, which would place it as twenty-fourth in abundance of the
chemical elements.
Zinc has ionic radii of 0.68A in fourfold coordination
and 0.83A in sixfold coordination; they are close to those of
magnesium, ferrous iron, and cupric copper, and these elements
commonly form solid solutions with zinc in oxygen salts such
as the sulfates and phosphates.
Although more than 80 zinc minerals are known, only a few
serve as commercial ores of the metal. The principal ores are
the sulfides sphalerite and wurtzite (cubic and hexagonal ZnS)
and their weathering products, especially smithsonite, ZnCO~,
and hemimorphite,'Zn, si9o7(OH) -ELO. Other minor ores are
zincite, zoO, and willemite, Za.SiO,.
Zinc is present in part in igneous and metamorphic rocks
as the sulfide sphalerite, but most of it present is disseminated
as a minor constituent of rock-forming minerals, especially those
rich in iron, such as magnetite, Fe-0^, the pyroxenes
(Mg,Fe)'2Si206 and Ca(M g,Fe) Si20g , the amphiboles, such as
Ca2 01g,Fe) 5Sig022 (OH)2, biotite, K( Mg,Fe>3 AlSi^g (OH,F)y ,
spinel, (Mg,Fe)Al204, garnet, (Fe ,Mg) 3A12 (S104) 3 , and staurolite
26
-------
(Fe,Mg)2Al9Si4023(OH). When igneous and raetamorphic rocks
are weathered, most of the zinc is concentrated in the clay
minerals of the sedimentary rocks and soils formed, especially
in the minerals of the montmorillonite group.
ZINC IN IGNEOUS AND METAMORPHIC ROCKS
Ranges of zinc content and averages for various types of
igneous rocks are assembled in Table 3-1. Zinc generally is
concentrated in basaltic rocks and is somewhat
depleted in granitic rocks. In basaltic rocks, the content of
zinc increases directly with the total iron content.
Data on metamorphic rocks indicate that their contents of
zinc are very similar to those of the unmetamorphosed rocks
in which they were found, that is, no indication of appreciable
mobilization of zinc during metamorphism has been found.
ZINC IN SEDIMENTARY ROCKS
Ranges of zinc content and averages for various types of
sedimentary rocks are listed in Table 3-2. Zinc is concentrated
notably in shales and clays; part of it is thought to be an
isomorphous replacement for magnesium and iron and part may be
adsorbed on the surface. The concentration is clearly higher
1627,1694
in black shales rich in organic matter, but the nature
of the zinc-organic complex and the exact mechanism of its
formation are not known. Zinc exhibits a two- to threefold
791
concentration in iron-rich laterites and bauxites. Jenne
has reviewed evidence indicating that hydrous oxides of manganese
27
-------
TABLE 3-1
Zinc in Igneous Rocks—
Type of Rock
Ultramafic
Basaltic
Intermediate
Granitic
Rhyolitic and dacitic
Alkalic
No. of
Analyses
85
If681
114
1,087
300
252
Range ,
ppm
25-103
42-420
5-127
5-235
15-400
18-1,070
Average ,
ppm
55
100
70
50
48
70
1743a 587a
Si
-Derived from Wedepohl, Gurney and Ahrens, and
1366
Rosman.
28
-------
TABLE 3-2
a
Zinc Contents of Sedimentary Rocks~
No. of Range, Average,
Type of Rock Analyses ppm ppm
Limestones and 490 < 0.1- 180 20
dolomites
Sandstones 150 5 - 170 30
b
Shales low in bituminous 365 46-200 95—
matter
Shales high in 980 15 -1,500 200
bituminous matter
Phosphorites 240 20-750 10 0~
Coal 1,600 7 -1,000 40-80
1743a
a
—Derived from Wedepohl.
—But the average for deep sea clays was about 165 ppm.
^Appreciably higher (average, 300 ppm zinc) in phosphorites of
the Phosphoria formation of Montana, Wyoming, Idaho, and Utah.575
29
-------
and iron are the principal controls of the fixation of zinc
in soils and fresh water sediments.
Zinc is also quite concentrated in marine phosphorites,
and especially high levels of zinc are reported in those from
Idaho, Montana, Wyoming, and Utah.575 These high levels of
zinc also are found in phosphatic fertilizers manufactured from
phosphorites of this area.
ZINC IN SOILS
The zinc content of soils has been the subject of some
review.153'938'1581'1696 Normal soils contain 10-300 ppm zinc,
average about 50 ppm, and in uncontaminated areas, the contents
generally are not very different from those of the parent rock.
A recent study of 863 U.S. soils1 77a noted an average of 54
ppm zinc (see bar graph for Figure 3-1). Zinc contents of
soils near sulfide deposits are generally higher than background;
this indicator is the basis of geochemical prospecting methods
widely used in exploration for ore deposits.
Soils near highways may be contaminated appreciably; zinc
contents of such soils891 are set forth in Table 3-3. The source
of the zinc may be from wearing of tires (containing zinc oxide) and
emissions from motor oil to which zinc dithiophosphite
has been added; these sources of zinc are in addition to
industrial emanations. Klein851 found that soil from industrial
areas near Grand Rapids, Michigan, had a mean content of 56.6 ppm
zinc, whereas nearby agricultural and residential areas had re-
spective means of 22.1 and 21.1 ppm zinc. Davies found similar
conditions in England.359
30
-------
r^'~«.
J...Q? T;-;-T-rA
la" ~*-~-?J. ^ * 0><*
V»
-------
TABLE 3-3
Zinc Content in Soils near Highways"
ppm Zinc Extractable by 1 N Hydrochloric Acid
Meters
Location from Road
Near U.S. 1,
Beltsville, MD
ii
it
Baltimore-Washington
Parkway, Bladensburg,
MD
H
ii
West of Interstate
29, Platte City,
MO
n
H
North of Seymour
Road, Cincinnati, OH
it
ii
8
16
32
8
16
32
8
16
32
8
16
32
Depth of soils, cm
0-5
172
66
54
162
110
44
54
60
15
72
60
34
rt f\ i
5-10
94
48
46
86
28
20
24
21
11
24
16
11
10-15
72
42
42
36
20
18
16
16
14
11
10
8.3
a
—Data from Lagerwerff and Specht.
32
-------
ZINC REACTIONS WITH ORGANIC MATTER IN SOILS
Many organic substances, including bundle and fulvic acids and a wide
711,1*35,1553
range of biochemical compounds, form stable combinations with zinc.
The occurrence of insoluble zinc-organic matter complexes has
been established, and the formation of soluble organic complexes with zinc has a pro-
found effect on the mobility and availability of the metal. The soluble
zinc-organic complexes can leach through the soil to influence weathering and
geochemical distribution in the soil profile. Organic substances are also
important in the transport and ultimate concentration of zinc and other metals
in such important deposits as peat and coal. Some soluble zinc-organic
complexes are so stable that the zinc is essentially unavailable to living
systems, but these extremely stable complexes are rare. Therefore, the chem-
istry of the zinc-organic complex in soil is closely related to the nature and
magnitude of microbial and other biologic activities as well as to the level of
zinc and other competing compounds present.
Natural Chelating Substances in Soil
Two main groups of organic compounds that form stable compounds with zinc
in soil are biochemicals which exist in living organisms, and complex polymers
formed by secondary synthesis reactions to form more complex products. The
first group contains organic acids, peptides, proteins, and polysaccharides, and the
second group contains humic and fulvic acids. The two groups cannot always be
clearly separated because some biochemical compounds are bound tightly to humic
materials. Zinc can function as a linkage in binding these organic compounds,
and it can be chelated by them. Most of the insoluble zinc-organic complexes
are associated with the humic group, especially humic acid.
The most soluble and least stable zinc complexes are with individual bio-
chemical compounds; but the fulvic acid complexes also have high water
1553
solubilities, yet greater stability than the biochemical complexes.
33
-------
Biochemical Compounds
The relative susceptibility of simple biochemical compounds to microbial
decomposition has often been assumed to be of little or no importance in metal
reactions in soils. But recent studies show that the combined total of poten-
tial chelating agents at any one time may tremendously influence the availa-
526,713
bility of zinc and other metals to plants. For example, up to 75% of
the zinc in displaced soil solutions was shown to be associated with low
526,713
molecular weight, dialyzable organic constituents. Natural complexing
biochemicals have also been demonstrated to be of considerable importance in
425
the transport of zinc to plant roots and its movement through the soil profile.
Many organic acids (malic, citric, oxa&cetic, fumaric, a-ketoglutaric,
^
pyruvic, etc.) and free amino acids (alanine, aspartic, glutamic, glycine,
cystine, cysteine, etc.) are the biochemical compounds of primary importance
in the formation of soluble zinc complexes in soils. Other compounds that
chelate zinc in soils include organic phosphates, phytic acid (inositolhexa-
phosphoric acid], chlorophyll and chlorophyll-degradation
products, simple sugars, porphyrins, phenolic compounds, and auxins. The con-
tribution of all these other compounds to zinc-organic complexes from biochem-
ical sources is minor compared to the prevalence of organic acids and free
711,1081
amino acids. Specific reactions have been reviewed in detail.
34
-------
Humic and Fulvic Acids
The binding of zinc and other metals by humic and fulvic acids is influ-
enced greatly by the nature and properties of the acids. The following
differences should be noted between fulvic acid and humic acid:
• fulvic acid is light yellow to yellow-brown, whereas the
color of humic acid ranges from dark brown to gray-black;
• the degree of polymerization is increased for humic acid;
• the molecular weight of fulvic acid is about 2,000, and the
molecular weight of humic acid is about 300,000;
• carbon content increases from 45% in fulvic acid to 62% in
humic acid;
• oxygen content decreases from 48% in fulvic acid to 30% in
humic acid; and
• exchange acidity decreases from 1,400 in fulvic acid to 500
1553
in humic acid.
Generally, humic acid is the material extracted from soil by alkaline solutions
and precipitated upon acidification, whereas fulvic acid is an alkaline-soluble
1553
material that remains soluble after acidification.
Formation of stable humic and fulvic acid complexes with zinc and other
metals is possible because of the high content of oxygen-containing functional
groups such as carboxyl, phenol, alcohol, enol-
hydroxyl, and carbon-oxygen structures. Amino and imino
1553
groups may also be important in zinc binding. Several functional
1314
groups have been identified as being involved in the binding of zinc by
humic acids. Phenolic hydroxyl and carboxyl groups having pKa values between
2.8 and 4.4 accounted for the least stable but greatest adsorbed fraction of
zinc, while more stable fractions were attributed to strongly acidic carboxyl
groups with pKa values below 2.0* Zinc would be more likely to be adsorbed by
the more stable forms.
35
-------
1314 1436 711
Randhawa and Broadbent, Schnitzer and Skinner, and Hodgson
have suggested that humic and fulvic acid fractions of organic matter are
1314
very important in zinc adsorption. Randhawa and Broadbent found that
the species of zinc complexed by the humic acid varied with pH. At pH 7.0,
they concluded that 70% of the metal retained by humic acid was present as
the divalent species, whereas at pH 3.6, 75% of the acid was monovalent.
However, inherent errors in determining the ratio of the complexes by the ion
exchange resin method may cast doubt on their conclusions. Stability constants
for the zinc-humic acid complex were calculated to be 4.42 at pH 3.5, 6.18 at
1315
pH 5.6 and 6.80 at pH 7.0.
1436
Schnitzer and Skinner reported zinc-fulvic acid stability constants
(Io8l0) of 1.73 at pH 3.5 and 2.34 at pH 5.0. Because bonding of metals to
fulvic acid involves carboxyl groups, the' dependence of pH on the stability
constants reflects the competition with hydrogen ions. Whenever possible, it
would be preferable to eliminate the pH dependence on the metal-fulvic acid
complexes in determining stability constants.
The log K value of 5.6 at pH 7.0 for a zinc-soil-organic matter complex
705
reported indicates that most of the zinc may have been adsorbed in the form
of a zinc-humic acid complex. The pH drop of the humic acids on addition of
metallic cations was found to be Mn+2, Ni+2, Zn+2, Cu+ , Al*3, and Fe+3
in descending order. Therefore, copper, aluminum, and iron may be more
tightly bound than zinc to humic acid.
Zinc can be fixed through solid phase adsorption (including complexation)
or the formation of insoluble precipitates. The hydrolysis (log K, » 9.7) and
solubility (log <8O = -16 £) constants for zinc hydroxides indicate that these
insoluble forms of zinc cannot be controlling the levels of soluble zinc in
36
-------
soils. Most of the zinc immobilization is attributed to surface adsorption
711 793
and complexing by organic matter. Jensen and Lamm found a high degree
of correlation between zinc content and organic matter distribution in differ-
ent soils. Destroying the organic matter of a surface soil allowed almost
all of an extra source of zinc to be extracted with dithizone (C6H5'N:N'CS'NH*NH-
CgHs) whereas only 50-75% could be recovered when organic matter was allowed to
87
remain. Thus, removing organic matter appears to decrease the immediate
reactivity of zinc in soils.
Although the immobilization of zinc by stabilized organic matter is well
established, newly formed organic substances (especially biochemical inter-
mediates and fulvic acids) are mobile and can solubilize zinc and increase
the metals availability to plants and other biologic systems. Leachates from
organic soils and humic layers from forest soils often contain considerable
amounts of soluble organic substances during periods of high biologic activity.
Hence, zinc movement and availability can be increased greatly whenever zinc-
enriched sewage sludges, animal manures, and industrial wastes are added to
soils. The possibility of influencing the environmental impact of added or
native zinc through proper management of soil organic matter is of considerable
importance because of the variety of chemical reactions that occur between zinc
and organic materials.
37
-------
ZINC IN WATERS
Most modern determinations of the zinc content of sea
water are in the range of 1-27 y.g/1 zinc, with a median at about
8 Mg. 543, 1291, 1349, 1498, 1530, 1625 Jt has been estimated that
about 700,000 metric tons of zinc are transported to the sea
annually. More than 99.9% of the zinc reaching the sea in
the dissolved form is eventually precipitated with oceanic
sediments, chiefly with clay minerals, but partly with manganese
oxide nodules and phosphorites . Appreciable amounts of zinc
can be precipitated as sulf ide in anoxic waters , such as those
that exist in parts of the Black Sea.^531
The zinc content of fresh waters is more variable, but
uncontaminated fresh waters generally contain 10 yg or less
zinc/1 water. The chemistry of zinc in such waters has been
reviewed recently by Hem, who has compared observed concentra-
tions of zinc with those calculated from thermodynamic data.
Zinc carbonate or zinc orthosilicate were
assumed to be solid phases. Most surface waters appear to be
unsaturated by the carbonate; the equilibrium of zinc silicate
may be more important.
Recent analyses of filtered surface waters of the United
States are summarized in Table 3-4. An earlier study869 pre-
sented similar results. Zinc was detected (sensitivity 20 yg/1)
in 76.5% of 1,577 samples; the maximum content reported was
1,183 yg/1, and the mean content was 64 yg/1.
38
-------
TABLE 3-4
a
Zinc Content of Filtered Surface Waters of the U.S.-
Zinc Content, ug/1
<
10 -
20 -
30 -
40 -
50 -
60 -
70 -
80 -
90 -
100 -
110 -
120 -
130 -
140 -
150 -
200 -
250 -
300 -
500 -
1,000 - 4
19,000
42,000
TOTAL
10
19
29
39
49
59
69
79
89
99
109
119
129
139
149
199
249
299
499
999
,500
40Q
No. of Samples
205
112
104
57
37
35
23
12
11
11
11
13
11
9
7
20
17
2
8
2
5
1
1
714
—Data from Durum et al.
39
-------
High amounts of zinc in surface waters represent indus-
trial and urban pollution, from such sources as zinc dissolved
from galvanized pipes and the dumpings of plating baths.
Streams that drain from areas of mining activity also may have
424,1068
zinc contents up to 21,000 yg/1. High amounts of zinc
were found in waters of the South Fork of the Coeur d'Alene
River (in northern Idaho) in an area where thousands of tons
1068
of ground tailings of zinc ores were exposed.
Waters of such streams tend to purify themselves by pre-
cipitating zinc with clay sediments or with hydrous iron and
manganese oxides. Quantitative data are insufficient to permit
an overall estimate of the proportion precipitated, but the
424
effect of such precipitation was shown by Elderfield et al.,
who found 1,000-10,000 ppm zinc (average was 3,700 ppm) in
69
sediments of the Conway River in Wales, and by Banat e£ al., who
examined seven German rivers for zinc content, and reported
highest values for the sediments of the Weser River, which
contained 400-3,100 ppm zinc, with an average of 1,572 ppm.
1225
Perhac found that although the zinc content of suspended
matter in two Tennessee streams was much higher than the con-
centration of dissolved zinc, most of the zinc was transported
as dissolved material.
Zinc is picked up by water in the distribution system and
household plumbing, and almost all drinking water has a de-
tectable concentration of zinc. All but three of 2,500 samples
40
-------
of water collected at consumers' taps had more than I p% zinc/1. The
survey of 969 water systems located in nine geographic areas in the U.S.
reported the average zinc content of drinking water as 194 yg/1. The
highest concentration detected was 13,000 vg/lf and 0.3% of the 2,595
1646
samples exceeded the drinking water standard limit of 5 mg/1.
When the intakes of food and water are compared, it is found that
drinking water would provide 4.3% of the average food-zinc intake in the
United States.
ZINC OOMTEMT OF CQKLS
Although hundreds of samples of coal have been analyzed for zinc,
a meaningful average content is not easy to obtain. Nearly all the
available analyses were made by spectrographic analysis of the residual.
ash of the coal; there is considerable danger of volatilizing part of
the zinc during ashing unless special precautions are taken. Furthermore,
the spectrographic method for zinc is comparatively insensitive; various
5,925,1823
workers have given their limits of detection of zinc as 50 to
200 ppm zinc in the ash. As indicated in Table 3-5, a substantial per-
centage of the analyses report zinc as "not found." Because all such
determinations were calculated as zero, all averages given are minimal
figures.
No attempt has been made to average all the data reported. Tables
3-5 and 3-6 give data based on reports in which analyses are given of a
substantial number of samples, and in which the percentage of ash was
given, so that results could be calculated back to the zinc content of the
289
coal proper. Earlier data have been reviewed by Clarke and Swain and
4
by Abernethy and Gibson.
41
-------
10
TABLE 3-5
Zinc Content of Coals of the tftiited States, by Regions
Area
b
Eastern province-
Appalachian region-
. b
Interior province—
. . . d
Eastern interior region—
b
Western states-
Western region^
Northern Great Plains-
No, of
Samples
600
378
123
475
104
44
221
Average Zinc in
a
Coal, ppiu
21.4
8.2
78.0
44.0
25.3
28.0
59.0
No. of Samples
Reporting "Not Pound"
-
296
-
260
-
-
199
TOTAL 1,945
AVERAGE 32.6
=In these calculations, "not found" was calculated as zero; the averages are therefore all
minimal figures. 5
^Derived from Abernethy et al.
^Derived from Zubovic et al.
^Derived from Zubovic et al.
^Derived from Zubovic et al.
Derived from Zubovic et al.
-------
TABIE 3-6
Zinc Content of Coals from Major Regions of Dcxxmtentation
APRS
b.c
Alabama— —
Arkansas-
fa
Colorado-
Illinois^'-
Illinois Basin (111., Ind., Ky.)-
b,e
Indiana
lowa^'-
b,c
Kentucky--
Ohio-'—
Oklahoma—
b
Pennsylvania—
b
Ttennessee-
UtahS
Virginiar-
. . . b,f
W. Virginia
East Germany^-
Nova Scotia-
No, of
Samples
~ii^^_jab^ij__M_^^
137
67
40
319
82
123
25
198
208
93
117
40
23
51
938
494
182
Average Zinc in
a
Coal, pptr-
9.7
7.2
33.0
141.0
313.0
33.0
41.0
10.7
20.5
13.7
22.2
17.3
76.0
22.7
32.9
149.0
25.0
a
~When zinc was not found, it was calculated as zero; the averages, are
therefore all minimal figures.
^Derived from Abemethy et al.
0«_ . . * * • ^ ,"1824
^Derived from Zubovic et al.
^Derived from Zubovic et al.
^Derived from Zubovic et al.
^Derived fron Headlee and Hunter.
Derived from Leutwein and Rosier.925
3382a
from Ruch et al.
43
-------
The average content of zinc in U.S. coals given in Table 3-5 (32.6
107
ppm) is of the same order of magnitude as other recent estimates: 50 ppm,
1816 1016
20 ppm for hard coal, and 60 ppm for brown coal. The last two
estimates were based on a survey of world literature, weighed according
to the total estimated reserves in each area.
The nature of the bonding of zinc in coal has been discussed by
1821a 1382a
Zubovic and Ruch et al. There are notable areal variations in
the proportion of zinc bound to the organic and to the inorganic con-
stituents of coal, but a large proportion of the zinc is bound to inorganic
constituents, and especially to the occurrence of sphalerite.
Data on zinc content of petroleum are scarce. A recent estimate for
107
oil was 0.25 ppm, and an average of 4.2 ppm zinc was given for 3 resid-
369
ual heating oils.
44
-------
CHAPTER 4
MAN-MADE SOURCES OF ZINC
MINING AND CONCENTRATING
There are over 30 mines and smelters where zinc is mined and produced
in the United States. Primary areas of zinc production are mapped in
Figure 4-1 and the specific locations are listed in Table 4-1.
Zinc ore, primarily sphalerite, ZnS, is mined by conventional under-
ground mining methods, then crushed in mills, and concentrated by dif-
ferential flotation. The choice of concentration techniques depends upon
the chemical composition of the zinc compounds and also the other con-
stituents (e.g., lead, copper, iron) in the ore. Losses of zinc to the
atmosphere from mining, milling, and concentrating are comparatively small,
but some do occur during blasting, ore handling, crushing, and wind loss
from tailings. During grinding and flotation, the ore is wet and atmospheric
emissions are small. It has been estimated that zinc emissions to the
369
atmosphere from mining and milling are 0.1 kg/metric ton of zinc mined.
Based on the 1973 total of 435,318 metric tons for U.S. mine production of zinc,
the total zinc emissions to the atmosphere from mining and milling would have
1644 a
been 43.5 metric tons. However, mining and milling, including flotation,
can be an important source of water-borne zinc, whenever water pumped from
the mine is utilized in the concentrating process. Figure 4-2 is a flow
diagram of mining and milling waste water from a lead-zinc mine in Missouri,
and Table 4-2 shows the concentrations of zinc found at various stages along
1796
that path.
45
-------
PRIMARY ZINC PRODUCTION AREAS IN THE UNITED STATES
FIGURE M—1 Primary zinc production areas in the United States. Prepared by the Zinc
Institute, Inc., June 1975.
-------
Primary
TABLE 4-1
STATE
Arizona
Colorado
Idaho
Maine
Missouri
New Jersey
New Mexico
New York
Pennsylvania
Tennessee
Utah
Virginia
Wisconsin
tine Prodi
NO.
©
©
©
©
©
©
©
©
©
®
©
©
©
®
®
®
©
©
@
©
®
@
®
©
©
($'
(*j)
©
©
iction Areas in the United
MINES
DISTRICT
Bruce
Eagle
Idarado
Leadville
Bunker Hill
Star Unit
Blue Hill
Brushy Creek
Buick
Fletcher
Magmont
Ozark
Viburnam
Sterling
Ground Hog
Balmat
Edwards
Friedensville
Coy
Elnrwood
Idol
Immel
Jefferson City
' New Market
Young
Zinc Mine Work
Burgin
Austinville
Shullsburg
States
COUNTY
Yavapai
Eagle
Our ay -
Lake
Shoshone .
Shoshone
Hancock
Reynolds
Iron
Reynolds
Iron
Reynolds
Iron
Sussex
Grant
St. Lawrence
St. Lawrence
Lehigh
Jefferson "
Smith
Hancock
Knox
Jefferson
Jefferson
Jefferson
Jefferson
Utah
Wythe
Lafayette
-------
TABLE 4-1
(Continued)
SMELTERS
STATE NO. DISTRICT COUNTY
Idaho 00 Kellogg Shoshone
Illinois (3 Sauget St. Clair
Oklahoma 03 Bartlesville Washington
Pennsylvania @J Monaca Beaver
gj Palmerton Lehigh
Texas @ Corpus Christi Nueces • •
a
Prepared by the Zinc Institute, Inc., June 1975.
48
-------
FLOTATION
REAGENTS
ADDED
LEAD-ZINC
ORE
FROM
MINE
vo
WATER
PUMPED
FROM
MINE
CONCENTRATE
THICKENERS
7///77//7///777A
FLOTATION
CELLS
EFFLUENT
DISCHARGE
OR
RECYCLED
I
SETTLING AND
TREATMENT LAGOONS
CYCLONED FOR FILL OR DAMS
DISCHARGE
TO
RECEIVING
STREAM
FIGURE 4-2 Flow diagram of mining and milling wastes. Reproduced from Wixson and Bolter.
1796
-------
TABLE 4-2
a
Mean Zinc Concentrations in Mined and Milled Waste Water
Mine A
(Lead, zinc, and
copper ore)
Mine B
(Lead and zinc
ore)
Locations
Around
Mine
Mill
and
Lagoon
Discharge
Stream
Mill
LC
Lagoon
Discharge
Stream
Early Mining
Activity
zinc, ppb
11
43
46
24
139
137
123
98
Present 1971
Mining Activity
zinc, ppb-
180
1,320 b
1,045 b
231
449
262
411
328
a 1796
Data from Wixson and Bolter.
Mill discharged through tailings direct to stream during construction of
new lagoons.
50
-------
Wixson and Bolter hold that the differences between the final
stream concentrations of zinc in the early and present mining situations
were caused by reduced retention time in recent years as the lagoons filled
up with tailings. However, the data in Table 4-2 indicate that the concen-
tration of zinc in the mill water, the point furthest up the waste stream
sampled, may be the determining factor of the eventual discharge concen-
tration. It also was noted that the zinc content of the streams returned
to baseline levels within a few miles of the source.
A study of the Coeur d'Alene River system in northern Idaho confirmed
that nonferrous metals mining and related activities can result in high
concentrations of zinc in waters that receive effluents. ^^ Zinc concen-
trations during low volume flow ranged from less than 0.1 mg/1 for the North
Fork of the Coeur d'Alene River, which has little mining activity, to 21.0
mg/1 at a station in the South Fork, which has considerable mining activity.
Concentrations of zinc in the South Fork were generally between 1-2 mg/1; in
the main stream the concentrations of zinc were between 2-5 mg/1.
51
-------
Although mining and concentrating activities are only minor sources of
zinc to the atmosphere, such activities can cause significant quantities of
zinc to be discharged into waterways. It should be noted that sedimentation
may cause concentrations of zinc in steams receiving effluent from mining
and concentrating activities to be reduced with distance downstream from
the point of discharge.1676
SMELTING AND METALLURGIC OPERATIONS
Primary Zinc Production
There are three basic types of primary zinc smelters in the United
States: horizontal-retort distillation units, vertical-retort distillation
units, and electrolytic plants. Regardless of the type of smelting process
zinc may be released into the air during concentrate handling, open storage,
and conveying. In each of these smelting processes, the concentrated zinc
ore must go through a roasting procedure to drive off sulfur dioxide, SOo,
and convert the zinc sulfide, ZnS, to zinc oxide, ZnO, Roasting may create
large amounts of dust, but because that operation is enclosed, the dust may
be readily collected.
The horizontal-retort distillation units are essentially batch processes
in which a roasted concentrate and coke mixture are heated in a retort to
approximately 1,100 C; natural gas is the usual fuel. Zinc is then reduced
from the roast (zinc oxide) to zinc metal in vapor form. The zinc vapor
passes into a condenser where it condenses into a liquid; it is then drained
off at intervals into molds.
52
-------
The vertical-retort distillation process produces highly pure zinc con-
tinuously. The basic process is essentially the same as for the horizontal-
retort process, except that the charge, consisting of briquettes of zinc
ore concentrate and coke, is fed into a charge column, which is a vertical
extension of the retort. The charge moves down through a combustion zone
where zinc vapor, carbon monoxide., and particulates are evolved. The
gases pass out through the top of the column into a condenser, where the
zinc vapor is condensed into zinc metal.
The electrolytic process is quite different from the other two
systems. After the ore concentrate is roasted, the concentrate is leached
with sulfuric acid to form a zinc sulfate solution. The liquid is
then pumped into electrolytic cells where the zinc deposits on aluminum
cathode. The cathodes are removed periodically and the zinc is stripped off.
The use of particulate collecting devices, primarily baghouses and
electrostatic precipitators, is highly efficient (> 95%) in recovering zinc
particulates. However, some emission sources are uncontrolled and little
control is exercised over emissions associated with concentrate unloading,
handling, and storage.3" The particle size of zinc emissions (composed of
zinc oxide and sulfur complexes) from the retort of a horizontal-retort
operation were as follows: 34% of the particles were 2.5 ym in diameter,
35% were 2.5-5.0 ym, and 31% were greater than 5.0 ym.-'69 Other estimates,
however, have indicated smaller particle sizes. The particle size for zinc
fume has been estimated at 0.01-0.3 ym in diameter and metallurgic dust
and fumes at 0.001-100 ym in diameter.317
Few quantitative data exist on the concentration of airborne zinc near
primary zinc smelters in the United States. Schrenk ejt al, estimated
that the total pollution load of zinc oxide from a zinc smelter involved in
the air pollution episode in Donora, Pennsylvania in 1948 was 11,068 kg
53
-------
zinc oxide/day. Another study of two vertical-retort zinc plants estimated
198
that the daily emissions of zinc since 1960 ranged from 6,000-9,000 kg.
However, particulate control systems can remove 99.5% of the particulate matter.1238
Based on conditions existing in 1969, the following estimates for emission
factors for the three types of zinc processes were made: horizontal-retort
smelting created 77 kg zinc emissions/metric tons zinc produced; vertical-retort
smelting released 36 kg zinc emissions/metric tons zinc produced; and the electrolytic
369
process produced 27 kg zinc emissions for every metric tons zinc. The study also
estimated that primary zinc smelting activity accounted for the discharge of
45,454 metric tons of zinc into the atmosphere in 1969. Since 1969, however,
five zinc smelters in the United States have ceased operations, thereby reducing
the zinc-producing capacity of the United States, as well as the total emissions
of zinc to the atmosphere. ^"^
Although data are not available on the concentration of zinc in the air
near zinc smelters, soil analyses are available. Table 4-3, which summarizes
the zinc content of soils near three smelters, shows that concentrations are
much higher than the average 50-54 ppm found in normal soils. Although most of
the zinc is in the top few centimeters of soil, there is appreciable downward
movement, as shown by the data in Table 4-4.
Great contamination has been shown in the upper 15 cm of the soil
1 QQ
profile near a zinc smelter. ° The range of zinc content of soil
within 1 km of the plant was 50,000-80,000 ppm zinc in the soil. Organic
matter in the same area contained as much as 135,000 ppm of zinc. It
has been estimated that the zinc deposition rate in this area was 1.75-5.25
^
g/mj/month, or approximately 207-621 kg/ha/yr. The concentration of zinc in soil
fell off sharply with distance from the plant. Background levels* of zinc were found
The level of zinc normally found in the soil if there has been no contamination
from any metallurgic operation.
54
-------
Location
Swansea, W. England
Avonmouth, England
E. Helena, Montana
traverse C
traverse A
TABLE 4-3
Zinc Contents of Soils near Smelters
Depth of
samples, cm
0-5
0-5
0-5
0-5
1-5
1-5
1-5
1-5
1-5
1-5
2.5-10
2.5-10
2.5-10
2.5-10
2.5-10
2.5-10
2.5-10
2.5-10
Distance from
smelters, km
1.5
3
6
16
0.32
1.13
4.5
6.9
9.5
12.7
1.61
3.22
6.44
12.87
1.61
3.22
6.44
12.87
Zinc Content
ppm
543a
310a
150a
45*
5,000
1,400
450
250
150
90
450^
25fl£
140?
82*
210&
14o£
91~
6f^P
^J^
a
Zinc extractable by 0.5 N acetic acid.
"Calculated from equation fit to data.
546
^Derived from Goodman and Roberts.
d 203
Derived from Burkett et al.
e
1043
Derived from Miesch and Huffman.
55
-------
TABLE 4-4
Variation with Depth of Zinc Content of Soils near Smelters
Location
Annaka, Japan
900 m from smelter
Poland
200 m from smelter
Avonmouth, England
250 m from smelter
Depth, cm
0-2
5
10
20
30
40
60
0-10
15 - 30
40 - 50
60 - 80
0-3
3-6
6-9
9-12
12 - 15
15 - 18
Zinc Content,
ppm
1}680
1,590
1,310
540
140
80
62
12,200
1,230
467
57
1,000
720'
280'
175'
250'
250a
aZinc extractable by 2.5% acetic acid.
862
^Derived from Kobayashi.
a 565
Derived from Greszta and Godzik.
, 946
jJerived from Little and Martin.
56
-------
25 km to the east (downwind) and 16 km to the west (upwind).198 Therefore,
airborne zinc emanating from smelters can cause significant local contamination
near smelters. However, the concentrations of zinc in soil noted above are the
consequences of metallurgic operations dating back to 1898. The installation
of control equipment and improved control technology should mean that the area
near the smelter now receives contamination at a much lower rate than in the past.
The potential waste waters from electrolytic refining are spent electrolyte
solutions and slurries formed by removing impurities in the electrolyte solution
by precipitating them in thickeners. According to Tallmadge, little or no zinc
appears in waste water, as spent electrolyte solutions, and supernatant liquids
1594
from slurries are usually recycled for re-use in the leaching step.
A survey of lead-zinc smelting and refining operations in the United States
606a
has shown that waste waters can have zinc concentrations ranging from 0.01-25 mg/1.
Tables 4-5 and 4-6 list the composition of waste waters from these processes. By
comparison, a 1970 survey of surface waters in the United States indicated that
most waters contained less than 0.05 mg/1; some exceeded 5.0 mg/1, and the highest
value was 42 mg/l.1116a
Another important primary source of zinc—accounting for 68,471 metric tons of
zinc in 1972^°a—is zinc oxide from fuming furnaces. Zinc is recovered from
lead blast furnace slag by heating the slag to high temperatures and blowing
coal and air through it. Zinc is reduced, volatilized, reoxidized, and then
collected as zinc oxide in bag filter units. Information is not available on
the quantity of zinc emitted to the atmosphere during zinc fuming operations.
However, the efficiency of the collecting units probably determines the magnitude
of zinc arising from fuming furnaces. One survey of a fuming operation found the
particulate emissions to be negligible because of the efficient collection by a
baghouse unit.1646a
Because zinc is a constituent in other ores and raw materials, zinc dusts
and fumes may be produced when other metals are refined or produced, primarily
lead, copper, and steel. Athanassiadis has reported that considerable quantities of
57
-------
TABLE 4-5
Composition of Waste Waters and Receiving Streams
in Lead-Zinc Metallurgic Processes for Lead Smelters and Refineries, mg/1
Upstream
pH 7.6-8.1
Arsenic 0 . 11
Cadmium 0 . 002
Copper 0 . 13
Iron
Manganese —
tn Nickel —
oo
Lead 0.03
Zinc 0.08-0.13
Sulfate ion (S0^=)
1*
Outfall Downstream Intake
7.4-8.2 7.6-8.2 7.8
0.15-0.46
0.02-1.09 0.03-0.113
0.13 0.13 0.34
0.08
0.2
—
0.07-0.157 0.03-0.05 0.14
0.11-2.00 0.08-0.43 0.019
— — 6.5
llc
Receiving Stream
7.8-11.1
—
0.0-0.007
0.0-0.015
0.0-0.03
0.004-0.05
—
0.0-0.01
0.01-0.048
9.2-23.0
IIId
Neutralized
Intake Acid Plant Water
7.2 5.0
—
— 7.7
0.06
5.0 7.4
0.9 0.7
0.2
0.5
8.0
126 960
Other
Waste Water
8.6
—
0.5
4.0
1.3
0.5
0.06
11.0
2.0
200
606a
from Hallowell
al.
Effect of waste discharge on stream: outfall was combination of processed and cooling waters. Major contributor
to processed water impurities was effluent from gas conditioning operation in which fumes and dusts were mois-
turized by spraying them with water before entering electrostatic precipitator. Spray water was collected in
sump below conditioner and discharged.
eCompany studied contribution of waste to stream over and above background impurities by analyzing intake
water and the receiving stream below the outfall. Major source of contamination in effluent was water dis-
charged from slag granulation operations.
^Company operates lead smelter in conjunction with sulfuric acid (H? S04) plant. It provided data on intake water
neutralized acid plant water before discharge, and other waste water, which includes slag granulation and cooling
water.
-------
Composition of Waste Waters and Receiving Streams
Ln
vo
in Lead^Zinc Metallurgic Process.es fpr. Zinc Smelters and Refineries, mg/1
a
pH
Arsenic
Cadmium
Copper
Iron
Manganese
Lead
Zinc
Sulfate
Cyanide
Dissolved Solids
Suspended Solids
Effluent
7.7
0.39
1.0
8.5
583
10
Intake
5.0 - 7.0
1.0 - 1.2
0.4 - 1.0
5.7
136 - 179
372 - 391
14 - 18
Discharge
5.2 - 6.8
0.9
0.02 - 1.35
1.3 - 19.7
0.4 - 1.75
18 - 25
182 - 291
1.3 - 1.5
304 - 534
14 - 164
606a
from Hallowell et al.
^Combined waste stream from zinc smelter includes coke plant, cooling water, gas scrubbers,
spills, clean-up, etc. No sulfuric acid plant.
eZinc smelter in conjunction with sulfuric acid plant. Analyses show composition of water
supply and combined waste discharge.
-------
zinc can be emitted into the air from the production of steel. Average
airborne zinc concentrations in four communities with steel-producing plants
have been reported in Table 4-7.1590 Concentrations during and after strikes
demonstrate the impact of the industrial activity on the zinc content of the
air.
Therefore, primary production of zinc and other metals may release zinc
into the environment. These contributions are appreciably reduced by
dust and fume control equipment and by water pollution control systems. How-
ever, even in situations under little control, the emission of zinc into the
environment is likely to be confined to a limited area near the smelter.
Secondary Zinc Production
Secondary or recycled zinc is an important source of zinc in the United
States. In 1972, 438,150 metric tons of recoverable zinc were produced by
American mines. In addition, 83,736 metric tons of secondary zinc were
produced, nearly 19% of the total U.S. zinc production.
Zinc scrap materials are composed of metallic scrap and residual scrap
materials (skimmings, residues, and drosses from metallic baths). Because
these materials differ considerably in zinc content and form, several differ-
ent processes are employed to reclaim the metal. Scrap brass and bronze
are also recovered by remelting. Bronze contains little zinc and is poured
at temperatures substantially below the boiling point of bronze. Brass,
however, contains 15-40% zinc and is poured at a temperature near the boiling
point of brass. Therefore, major zinc emissions may occur from brass
remelting operations.-^9 Analysis of various brass smelter flue dust samples
indicated zinc concentrations of 47.0-70.4%, when approximately 0.9-1.8 metric
tons of flue dust were collected daily.512a>1269a
60
-------
TABLE 4-7
Zinc Concentrations in Air During and After Steel Industry Strikes
a
in Four Communities in the U.S. (1956) and Corresponding Probability Levels
Average Concentrations,
yg/m-* No. of ,
Community After Strike Strike Difference Samples Probability^
Birmingham, Ala. 700 200 500 9 0.028
Donora, Pa.C 11,800 100 11,700 15 < 0.001
East Chicago, Ind. 1,600 300 1,300 9 0.002
Allegheny County, Pa. 1,100 100 1,000 8 0.009
1590
from Tabor and Meeker.
^Probability of obtaining the observed differences by chance alone; the
limit of statistical significance is p=0.05.
cThe only community with a zinc plant (closed during the strike) .
61
-------
Zinc scrap materials are sorted and sometimes pretreated to drive off
moisture, oil, and other organic impurities. After pretreatment, the scrap
zinc-bearing material is sweated and/or distilled. In sweat processing, heat
is applied to scrap materials to melt the metal. A flux may or may not be
used. A molten-metal bath is formed from the metallic zinc and dissolved alloy
metals. The molten metal may then be cast directly into blocks for subsequent
further processing, fed directly to a distillation furnace, or cast into ingots
to specifications. The distillation process is similar to the distillation
systems used in primary zinc production. Either a retort or a muffle furnace
is used, each equipped with condensers."°°
The principal source of airborne zinc emissions in secondary zinc process-
ing occurs from vaporization of the metal in melting and pouring, and through
escape of zinc fume from the sweating and distillation processes. It has been
0£Q
estimated00 that zinc emissions to the atmosphere vary from 5 g-62 kg/metric
tons of products, averaging 9 kg/t. The total zinc emissions to the atmosphere
369
came to 3,455 metric tons in 1969. Although the amount of secondary zinc
seems small when compared to emissions from primary zinc production (45,455
metric tons in 1969), as secondary zinc evolves as a more important source of the
metal, its contribution to total zinc emissions will increase.
Zinc Oxide Production
Zinc oxide is used extensively in industry. In 1974, 228,356 metric tons
of zinc oxide were produced in the U.S. and used for many purposes, the most
important being in the production of rubber.135^a Zinc oxide may be produced
chemically, or by direct or indirect pyrometallurgic means. As mentioned, in
the direct (American) process, zinc vapor is produced from ore or scrap and
62
-------
then oxidized to form zinc oxide. The indirect (French) process uses zinc
metal, which first is vaporized and then oxidized to form zinc oxide. One
survey has indicated that zinc emissions to the atmosphere during the produc-
tion of zinc oxide in 1969 ranged from 9-85 kg zinc/metric ton of product. It
was estimated that in 1969 emissions from zinc oxide production totalled 7,330
metric tons.369
Manufacturing and Fabricating
The major end uses of slab or metallic zinc are galvanizing, manufacturing
brass products, and die-casting alloys. During galvanizing, zinc is primarily
released into the atmosphere when ammonium chloride flux is added or
when the flux layer over the hot zinc bath is disturbed.
Brass is a copper alloy containing up to 40% zinc. Bronze contains a
much smaller percentage of zinc. During processing of both products, the
metals are melted together and poured at elevated temperatures (649-1,316 C).
Zinc fume is released to the atmosphere during heating and pouring. Similarly,
when zinc die-castings are produced, the zinc alloys are melted and poured into
369
dies and then cast into the desired forms. Table 4-8 lists the amounts and
sources of zinc emissions into the atmosphere from manufacturing and fabricating.
Industrial operations using zinc contribute to the zinc content of streams
into which waste water flows, of course, but few quantitative data exist
from individual sources.
ZINC IN THE COMMUNITY ENVIRONMENT
Available data on air levels of zinc in urban communities without mines
54
or smelters indicate a general decline in airborne zinc from 1954 through 1964.
914a
Lee and Lehmden report concentrations of zinc particulates in urban areas
63
-------
TABLE 4-8
Airborne Emissions from End Uses of Zinc'
a
Use
Zinc-based alloys
Zinc-coating
Brass and bronze
Rubber tires
Photocopying
Paint
Zinc sulfate
Miscellaneous
Major Source
of Emission
Melting
Molten zinc baths
Melting
Abrasion
Waste disposal (burning)
Production
Production
Emissions,,
metric ton/yr (1969^
2,727
864
164
7,836
1,364
9
27
1,000
369
Adapted from Davis.
64
-------
throughout the U.S. ranging from 0.1-1.7 pg/m3, and data from the National
Air Sampling Network have recorded annual average airborne zinc concentrations
that i645a
throughout the U.S. /are generally less than 1 ug/m3, as shown by Table 4-9. *y
Motor vehicles , fuel oil and coal combustion, incineration, soil erosion,
and industrial, commercial, and construction activities contribute to zinc in
urban atmospheres. Estimates have been made of the amounts of zinc released
into the urban atmosphere by these activities and products. For example,
tire wear and leakage and combustion of fuels and lubricants containing zinc
additives account for most of the zinc released by motor vehicles. Rubber
tires contain 1.5% zinc by weight jH72a according to Davis, 3<^ ztnc wears off
rubber tires at a rate of 45 g zinc/tire/yr, or 1.2 kg zinc/million km.
Fuel and lubricating oils contain 30-1,500 ppm zinc;^72a the zinc arising
from motor vehicle emissions is preferentially attached to small (submicron)
particles in amounts of 0.1-10 ppm.^l^a
The amount of zinc contributed from motor vehicles appears to vary from
city to city. Gordon, in College Park, Md.t and Fried lander, in Los Angeles,
reported that the fraction of zinc in the atmosphere accounted for by motor
vehicles wae 23% and 22%, respectively. 72a However, Creuson et_ al.335 found
that neither site differences, distances, nor depth gradient effects were
significant for dust fall zinc when comparing samples representative of various
Industrial and urban patterns collected from roadside sites in Cincinnati.
Ondov £lt aJL, H72a compared zinc: lead ratios for urban area roads and a
tunnel, which provided a system for accurate measurement of background
air. Sample! from the tunnel gave a zinc: lead ratio of 1:100, whereas the urban
areas had a zinc: lead ratio of 1.10(0.25; hence, motor vehicles account for
only a small percentage of the zinc observed in city air. Moreover, they are
not the major source of zinc on suspended particles in urban atmospheres.
65
-------
TABLE 4-9
Urban Air Samples and Yearly Zinc Averages, yg/m3 —
Birmingham, Alabama
Paradise Valley, Arizona
Texarkana, Arkansas
Bakersfield, California
Bridgeport, Connecticut
Atlanta, Georgia
Boise, Idaho
Moline, Illinois
Beverly Shores, Indiana
Dubuque, Iowa
Ashland, Kentucky
1.09 Baltimore, Maryland .34
.05 Brockton, Massachusetts .11
<.01 Kalamazoo, Michigan .04
<.01 Moorhead, Minnesota <.01
1.60 Lincoln, Nebraska .50
.52 Bridgeton, New Jersey .08
<.01 Albuquerque, New Mexico <.01
<.01 Akron, Ohio .48
.20 Altoona, Pennsylvania .27
<.01 Baymon, Puerto Rico <.01
.54 Portsmouth, Virginia .08
^Adapted from the National Air Sampling Network.
1645a
66
-------
Fuel oil is used in homes for heating and by industries, electric
utility companies, railroads, oil companies, and the military. Based on an
average zinc content of 4.17 ppm for fuel oil, Davis369 has calculated a zinc
emission factor for fuel oil of 0.64 kg zinc/1,000 barrels of oil burned. In
the U.S. in 1969, combustion of fuel oil accounted for 410 metric tons of zinc
emitted to the atmosphere, or approximately 0.28% of total zinc emissions.
The probable forms of the zinc particles in the flue gas are zinc oxide and
zinc sulfate.860a
Davis369 also provides estimates for zinc emissions contributed by burn-
ing coal. The average zinc content of various U.S. coals is 54.6 ppm (see
Chapter 3). Based on U.S. figures for 1969, the calculated zinc emission
factor for burning coal is 8.5 kg zinc/1,000 metric tons of coal burned. This
means that an estimated 3,922 metric tons of zinc were emitted, that is, 2.7%
of the total zinc emissions from milling and nonmining sources. Most of the
zinc is discharged with the airborne fly ash.^^Oa
Small but significant amounts of zinc are released during incineration.
otq
DavisJ y calculated that 25,435 metric tons of zinc, or 17.5% of total zinc emissions,
were emitted into the atmosphere in the U.S. in 1969 from incinerations. This
estimate is based on an average zinc content of about 2,400 ppm a in sewage,
sludge, and 23,842 metric tons of refuse and garbage that were ultimately incinerated.
Much of the zinc particulate material released into urban atmospheres
from all of these sources settles on pavements as "fallout" and is washed
into natural and man-made drainage systems as rainfall runoff. A study of
runoff from street surfaces following a moderate-to-heavy storm in seven
cities revealed zinc values ranging from a low of 0.03 kg/curb mi to a high
67
-------
of 0.95 kg/curb mi, with an average of 0.34 kg/curb mi (see Table 4-10).1414a
The zinc was of no particular size or particle range, and it was one of the
metals present in the greatest amount in the runoff, regardless of land use
category (see Table 4-11). Zinc loading intensities were heaviest in indus-
trial areas and lightest in commercial areas (see Figure 4-3).' However, such
distinctions disappear when the contribution of zinc is considered in terms of
percent by weight of total solids, that is, pounds (grams) of metal per 100
pounds (kilograms) of dry solids, as shown in Figure 4-4.
Drinking water samples from throughout the United States indicate that
despite the contribution of airborne zinc to water systems as fallout or in
precipitation, the drinking water standard of 5 mg/1 was exceeded on the
average in only one of 969 water systems tested. Of a total of 2,595 distri-
bution samples, the 5 mg/1 standard was exceeded in 8 cases, or 0.3%. The
1646
maximum zinc concentration found was 13 ppm.
Available data indicate that significant zinc contamination of the
environment in specific limited areas near point sources exists. However,
there do not seem to be major mobile sources of zinc nor does zinc appear to be
an increasing environmental contaminant.
ZINC USES AND MARKETS
Zinc consumption in the United States during the 1960fs and on into
the 1970's has averaged over 1,183,000 metric tons/yr, making zinc the fourth
most used metal after steel, aluminum, and copper. The consumption rate is
high because of the wide availability and advantageous properties of zinc.
Consumption of zinc is sensitive to economic cycles, particularly
because the metal is mostly used in automobile production (for die-casting,
68
-------
TABLE 4-10
Concentrations of Zinc in Runoffs from Street Surfaces
City
San Jose - I
San Jose - Ii
Phoenix
Milwaukee
Baltimore
Atlanta
Tulsa
Seattle
Zinc Loading Intensity,
kg/curb mi
0.63
0,13'
0.16
0.95
0.59
0.05
0.03
0.17
Average
0.34
1414a
Derived from Sartor and Boyd.
"San Jose was tested twice, once in December 1970 and again
in June 1971; other cities were tested in the summer of 1971.
69
-------
TABLE 4-H
Distribution of Heavy Metals by Land Use Category from
Metal
Chromium
Copper
Zinc
Nickel
Mercury
Lead
Cadmium
Street Surface
Residential
5
10
38
1
10
36
_LMB
Contaminants ,
Industrial
8
14
44
5
4
25
__
% by Weighta
Commercial
5
20
24
3
20
28
— _
Total
7
11
40
3
4
35
^•v
100%
100%
100%
100%
a 1414a
Derived from Sartor and Boyd.
The figure reported here as "residential" was computed by combining
all of the observed data for the four residential land use categories
sampled in each city. "Industrial" and "commercial" figures were
computed similarly.
70
-------
0)
I-l
1
o
25
20
H 60
H AS
H >-"
CO
8
15
10
RESIDENTIAL
INDUSTRIAL
COMMERCIAL
FIGURE 4-3 Zinc loading intensities on street surfaces: variations with
land use. All seven cities are considered together. The metric equivalent
for residential areas is about 12.24 kg/km; for industrial areas, it is
about 17.28 kg/km; and for commercial areas, the figure is about 2.16
kg/km. Reproduced from Sartor and
71
-------
.o
c/j
o
O
in
c
2
LU
O
O
o
NJ
LAND USE CATEGORIES
FIGURE 4-4 Varying zinc concentrations according to land
use. All seven cities are considered together. Reproduced
from Sartor and
72
-------
galvanizing, and oxide for rubber tires) and in industrial and residential
construction activity (for galvanizing, brass, die-casting for appliances,
and oxide for paint pigments). This sensitivity can be observed by noting the
recent change in the consumption pattern for zinc in the United States. Since
the 1960's, die-casting alloys accounted for the largest consumption of zinc,
galvanizing has been second, and brass a distant third. These patterns
of consumption are charted in Figure 4-5.
In 1974, however, the U.S. consumption of zinc for die-casting alloys
dropped to a level below that for galvanizing. Because of anticipated growth
in the galvanizing markets and a probable lower growth rate in zinc die-
casting, consumption of zinc for die-casting is likely to remain at a lower
level (personal communication, T. F. Shaffer, Zinc Institute, Inc.). Consump-
tion patterns vary throughout the world, but in general, galvanizing is the
leading market, especially in Japan and Australia, where zinc consumed for
galvanizing surpasses all other zinc markets combined. In Europe, galvanizing
is also the leading zinc market in all countries except the United Kingdom,
where brass is the largest single consumer of zinc.
73
-------
1400
1925
FIGURE 4-5 Consumption of slab zinc in the United States. Reproduced from
the annual review of the U.S. zinc industry.13543
74
-------
Zinc Die-Casting
For many years, zinc alloys have been the most widely used metal in die-
casting, because of their combination of favorable properties, adaptability
to the process, and low cost. The low melting point of zinc also preserves
the life of the dies longer.
It has been estimated that over 455,000 metric tons of zinc were used in 1974
by die-casters in the United States.1354a The metal is used in major appli-
ances and automobiles for functional parts like pumps, impellers, carburetors,
and housings. It is also employed for decorative door handles, trim, frames,
and control panels. Toys, hardware, plumbing fixtures, and industrial com-
ponents are other major markets for zinc die castings.
The development of ILZRO 12, a general foundry zinc alloy, in the late
1960's has opened new casting markets for zinc. Components previously cast
in iron, copper-base, or some aluminum alloys are now being produced with this
zinc alloy.
New die-casting techniques developed in the early 1970's have made it
possible for conventional zinc castings to be made with cross sections only
about half as thick as those made in previous years. Thus, material costs
and weight of the product are reduced, yet its tensile, fatigue and impact
strengths are retained or even improved.
rlention of this product does not imply any endorsement or recommendation by the
National Academy of Sciences or the National Research Council.
75
-------
Wrought Zinc
Depending on the ultimate product requirements, numerous "compositions
and alloys are used for wrought zinc applications. Various alloying metals
are used to improve various properties (such as stiffness and creep resistance*)
for specific applications.
The conventional grades of slab or ingot zinc are rolled into sheet,
strip, ribbon, foil, plates, or rod according to their uses. For instance,
dry cell battery cases, are made from wrought zinc rolled into a strip. The
zinc also may be continuous cast into rod or bar. Because of its properties,
rolled zinc is easily workable into various shapes and forms by common fabri-
cating methods, such as stamping, forming, and spinning. It may be polished,
plated, painted, or left to weather naturally.
Major markets for wrought zinc include the electrical and electronics
industries, which make products such as brackets, shielding, capacitor cans,
and heat sinks; general manufacturing, for products like drawer knobs, bezels,
and golf club sole plates; the jewelry industry for medallions, jewelry, and
buckles; and construction, for flashing, trim, and decorative panels.
Galvanizing
One of the oldest functions of zinc has been as a protective coating on
steel to prevent rusting and its ultimate destruction. It is estimated that
over 450,000 metric tons of zinc were consumed in the U.S. in 1974 for coating of
*Creep resistance is the ability of metal to resist elongation over long periods
of time under sustained loading stress. See also Chapter 2.
+
Continuous casting is a process for converting molten zinc directly into rods
without interruption by casting the metal between two counter-rotating steel
wheels with V-slots on their faces.
76
-------
Continued improvements in the galvanizing process and related areas
have increased consumption totals markedly for conventional applications.
In the late 1930's, for example, the introduction of the continuous hot-dip
galvanizing line led to wide installation of such lines around the world,
turning out millions of tons of galvanized sheet and strip. But the traditional
method of dipping a steel product into a bath of molten zinc is still very much
in use, particularly for prefabricated and very large parts. Over the years,
galvanizing has proved effective in protecting such steel products as bridges,
railroads, automotive vehicles, ships, wire, cable, pipe, hardware, containers,
trash cans, and roofing.
Although hot-dip galvanizing is the most popular method for protecting
steel against corrosion, metallic zinc can be applied to steel in several
other ways: by electrodeposition, metallizing or spraying of molten metal,
sherardizing, and painting.
The all-galvanized bridge became a reality in the late 1960fs when it
was demonstrated that galvanized bolted connections were technically and
economically feasible.31a The use of galvanized steel reinforcement for
concrete also became widely accepted then. Galvanized steel reinforcements
are integral parts of major governmental and institutional buildings, office
buildings, bridges and roads, chemical plants, storage tanks, and other build-
ings where prevention of concrete spalling or rust staining from black steel
reinforcement is necessary.
Zinc-rich paint has become one of the fastest growing new markets for
zinc in the past few years. Such paints are most often used to supplement
galvanized steel for automotive underbody protection against corrosion, for
77
-------
bridges and other structures, for ship hulls, and for various buildings. Zinc-
rich paint provides sacrificial galvanic protection of steel similar to that of
galvanized coatings and is recommended whenever economics or size and shape of
products make conventional galvanizing impractical.
»
Large-scale applied research in the early 1960's on whether zinc anode
systems should be used for marine tankers showed zinc to be superior to mag-
nesium anodes in performance and economy. That zinc does not spark was a
•
significant advantage, and now zinc anodes are the preferred sacrifical anode
for protection against corrosion in this application. Related research led
to a later development of zinc anode systems for galvanized hot water storage
and glass-lined tanks, which provided superior service life at an economic
price. Protective zinc anodes also were developed for crab pots. a
ZincChemicals
Zinc and its compounds are used more and more in the chemical-metallurgic,
ceramic, fertilizer, paint, paper, plastics, rubber, textile, and electronics
industries. As mentioned, zinc oxide is the most important zinc chemical;
it is employed primarily as a reinforcing pigment in rubber, where it provides
good heat conductivity and resistance to aging by sunlight. Zinc oxide is
a necessary substance for the copying industry, where it is used as a thin
electrostatic coating material. Although only a micron-thick layer of zinc
is applied when used, the zinc oxide consumed by the copying industry has
been estimated to be around 33,000 metric tons in 1972.
Recently, zinc has become a supplement in plant and animal feed. Trace
amounts of zinc are essential for the growth and development of plants and
animals, and this essentiality will be discussed in Chapters 5, 6, and 8.
78
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WASTE DISPOSAL
Tailings
Tailings are the gangue and other refuse material left over from the washing,
concentration, or treatment of ground ore. Tailings from the processing of zinc
and lead ores may include considerable amounts of zinc, as well as sulfates,
chlorides, copper, iron, lead, cadmium, arsenic, and selenium. These tailings
are dissolved or suspended in process waste waters which have come into direct
contact with raw materials and intermediate or final products or by-products.
Table 4-12 shows some of the concentrations of metals contained in waste water
discharge streams.1643
Most major waste water streams from processing plants contain sulfuric
acid as well as trace metals. Addition of a lime slurry to the water raises
the pH from around 2 to between 10 and 11.5, causing precipitation of several
of the metals as hydroxides. The water is channeled through an enclosed system
to a settling pond where the suspended solids settle; absorption of carbon
dioxide from the atmosphere leads to formation of carbonates, gradually re-
ducing the pH of the pond. The effluent which evolves contains less than
10 mg/1 of total suspended solids.1°^3 Table 4-13 shows typical concentrations
of various constituents from a lime and settle treatment plant.
Waste water is often cycloned before liming and settling. As the water
passes through cyclone pumps, coarse sand is separated from finer material.
The coarse sand is used to build the retaining walls of the disposal dams;
the fine material is dumped into the lagoon with the mine water or other waste
stream water and left to settle. Waste water from flotation concentrators,
thickeners, and zinc scavenging circuits is treated by cycloning and then
allowed to settle. Cooling water from metal casting operations is usually
just settled, whereas acid-plant blowdown can be treated in several ways:
79
-------
TABLE 4-12
Gross Concentrations in Discharge Streams0
Waste Material Concentration"
Dissolved solids 455-4,485 mg/1
Suspended solids 25-249 mg/1
Sulfates 175-2,221 mg/1
Chlorides 60-620 mg/1
Arsenic 0.1-0.68 mg/1
Cadmium 0.02-2.4 mg/1
Copper 0.01-0.34 mg/1
Iron 0.02-1.93 mg/1
Lead 0.02-1.35 mg/1
Selenium 0.007-1.8 mg/1
Zinc 5-243 mg/1
a 1643
Data from the U.S. Environmental Protection Agency.
Data summarized from six smelting/refining plants.
80
-------
TABLE 4-13
Effluent Concentrations from Lime and Settle
Treatment of Mixed Wastes'2
Concentrations, mg/1
Constituent
PH
Total solids
Sulfur
Chloride
Cadmium
Lead
Selenium
Zinc
JflT tl TTTlt ITO
9.3
1,430
250
140
0.03
0.5
0.8
1.0
Maximum
10.8
4,050
730
490
0.7
1.8
5.0
8.8
Average
9.6
-
650
480
0.3
0.7
-
2.0
Data from the U.S. Environmental Protection Agency.
Except for pH.
81
-------
by a combination of liming and settling, by mixing with other waste streams,
settling, and liming, or by mixing and settling. The Akita zinc smelter in
Japan, designed with zero pollution emission as a primary goal, treats its
waste waters by using a two-stage neutralization with ground limestone* CaCO,,
and calcium hydroxide, Ca(OH)2, followed by settling. The final discharge
water from such treatment has a zinc content of 0.5 ppm or less. Table 4-14
QOQ
compares the efficiencies of various methods which remove zinc from effluents. JJ
The clarified water that remains in tailings ponds after settling occurs
is left to evaporate, to percolate through the bed of the pond and underlying
soil, or to be diluted by rain and snow before being decanted and released
to the natural watershed. Or, it will be discharged directly to the watershed
if the concentrations of trace contaminants are low enough.
Few specific data are available on the concentrations of zinc contributed
to ground water from water percolating through the beds of tailings ponds.
Concentrations of zinc in the natural waters of zinc mining areas have been
found as high as 50 mg/1, although in most ground and surface fresh water,
zinc is present only in trace amounts. 1434 A median value of 20yig/l has been
reported in the surface waters of the United States. ^
Studies of the translocation of zinc in soils demonstrated the relative
immobility of zinc added to sandy soils, even at pH 4.^90 Similarly, Wheatland
and Borne, applying river water to soils of the Bunter Sandstone area of England,
found the concentration of zinc in the percolate to be considerably less than
the concentration in the water applied (see Table 4-15).
82
-------
TABLE 4-14
Treatment Processes for Removal of Zinc
Possible Effluent Concentration after
Treatment Processes Treatment, mg/1
CPa (with lime) + Sd 0.5-2.5
CP° (with lime) + Sd + F6 0.1-0.3
CP° (with lime) + Sd + Rf/ +
Fe + AC^ 0.4
o, 933
Data from Lin and Lawson.
b
Treatment efficiencies vary according to initial concentrations of raw
wastes.
^Chemical precipitation.
Sedimentation.
filtration
•'Recarbonation.
^Activated carbon adsorption.
83 ,
-------
TABLE 4-15
a
Mineral Analyses of River Water and Percolate Applied to the Soakage Area
Mineral, ppm
Total hardness (as
CaC03)
Total dissolved solids
Calcium
Magnesium
Sulfate
Chloride
Iron
Nickel
Total chromium
Manganese
Zinc
Lead
Copper
Water Applied
9/30/57
262.0
456.0
76.0
18.8
88.0
49.0
0.5
0.16
0.04
0.25
1.00
0.025
0.035
Percolate
1/31/58 9/30/57
268.0
— 457 .0
80.0
18.0
94.0 89.0
49.0 49.0
0.3 0.2
0.11 0.03
0.015 0.015
0.01 nil
0.25 0.06
0.01 0.012
0.02 0.01
1/31/58
—
—
—
—
90.0
49.0
0.24
0.04
0.017
nil
0.02
0.008
0.02
a 1762
From Wheatland and Borne.
84
-------
Zinc was mobilized in soil by continuous flooding in the presence of
918
fermenting organic material, but immobilized when the soil was reoxidized.
Lehman and Wilson observed similar results, using suburban residential sewage
effluent filtered through sandy soil material. When lysimeter columns were
intermittently treated with effluent, less translocation of trace metals
occurred than when the columns were treated by continuous flooding. The tests
performed on both field plots and lysimeter columns proved that:
• zinc was present in approximately the same amounts in both
studies; concentrations of zinc at the 15- and 30- cm depths
of the field site were nearly the same as the concentrations
at comparable depths of the continuously flooded lysimeter
columns;
• in the lysimeter study, zinc was removed from the applied
effluent at or near the soil surface, and only minor amounts
of zinc were detected in the filtrates; and
• some zinc was translocated when filtration rates were high,
as during the first few weeks of irrigation and when the
Q1 O
soil was saturated. °
85
-------
Sewage
Zinc is one of the trace elements found in highest concentration in
municipal waste waters. Depending upon the treatment process utilized, the
zinc in waste water will be separated into effluent and sludge. On a concen-
tration basis, the sewage sludge is usually enriched, whereas the effluent is
lower in zinc than the incoming waste water. Even though effluents may be
relatively low in zinc content, the great amounts of effluent which are dis-
charged at larger sewage treatment plants can be responsible for discharging
significant portions of zinc to receiving streams, irrigation sites or other
effluent-receiving systems. The zinc concentration permitted in industrial
effluents is regulated by discharge limits, so the metal is removed from
contaminated industrial waste waters and often forms industrial sludges which
are highly contaminated by zinc as well as other metals. These sludges
are usually disposed of by landfill, but some are incinerated.
Pound and Crites^^SSa have listed effluent concentrations of zinc
measured from differing stages and types of sewage treatment in California.
The values given for primary effluent were 0.83 ppm zinc, trickling filter
effluent, 0.16 ppm zinc, activated sludge effluent, 0.32 ppm zinc, and pond
effluent, 0.39 ppm zinc. These data are based on limited samples, but
Blakeslee*23a nas presented data on effluents ranging from 0.01 to 4.7 ppm
zinc in a survey of 58 sewage treatment plants in Michigan. However, most
effluent values for zinc are well below 1.0 ppm, and a value of less than
0.5 ppm zinc has been cited as typical, '"a when sanitary and storm sewers
are combined, the addition of zinc to sewage by fallout of zinc particulate
material onto urban pavements is significant (see Figure 4-3 and Table 4-9).
Therefore, many loading factors of zinc must be considered, and the zinc
concentration in the effluent from a single treatment plant may vary greatly,
depending on water use and amount of zinc received.
86
-------
If all sewage were treated to meet secondary treatment requirements
set by the EPA, about 8.1 million dry metric tons of digested sludge would be
produced in the U.S. each year; the current amount is about 4.5 million dry
metric tons (personal communication, R. Bastion). Because of the variation in
wastes arriving at the sewage treatment plants, the sludges range widely in
zinc concentrations. Many municipalities have industries that produce waste
water high in zinc and more than 98% of it is segregated into sewage sludge.
Sludge zinc concentration increases during anaerobic sludge digestion, because
unstable organic components are lost and zinc forms an increasing percentage
of the remaining material. Therefore, the zinc is effectively concentrated
on a dry-weight basis.
2S6
Chaney has found zinc concentrations in digested sewage sludge
ranging from 500 to 50,000 ppm. Dean and Smith3713 have shown that the
distribution of zinc values in sludges tend to be logarithmically
normal: some extremely high values are traceable to industrial effluents.
A geometric mean gleaned from over 100 studies was
2,420 ppm zinc, and 80 additional samples from sewage treatment plants in
the U.S. yielded a geometric mean of 6,380 ppm zinc.371a Berrow and Webber
have described the composition of British sludges and the range of values for
the 42 sludges sampled was 700 to 49,000 ppm zinc, with a mean value of 4,100
and a median value of 3,000 ppm. By comparison, normal values for zinc in
283
soils range from 10 to 300 ppm with a typical level of 80 ppm. Swedish
workers have reported a mean value of 2,800 ppm zinc in 1968 and 1969 and
11 'JR
2,500 ppm in 1970 from routine samplings of sludges at 57 treatment plants.
The five sludges highest in zinc in Sweden ranged from 5,700 to 17,200 ppm
zinc.11^ Blakeslee^3a reported zinc values for sludges from 58 municipal
sewage treatment plants in Michigan. They ranged from 72 to 16,400 ppm, with
87
-------
a median value of 3,200 ppm zinc. Recently, Page1191a and Peterson et al. a
also have summarized published data on the metal content of sludges.
Depending upon the disposal or utilization of the sludge material,
the presence of zinc in sewage sludge may or may not be a potential problem.
Zinc contamination should cause no problems in well-designed landfill sites
where there is no opportunity for leaching into surface waters or groundwater
supplies. As mentioned, significant amounts of zinc may be released to the
atmosphere when sludge is incinerated. Sludge which will be applied to agri-
cultural or other lands that support plant growth may be the most dangerous
problem. Indeed, zinc is such a common contaminant of sewage sludges that
283
Chumbley expressed the permissible levels of toxic metals in sewage used
on agricultural land in terms of a zinc equivalent.
Solid Wastes
In addition to the effluents discharged to tailing ponds, zinc-bearing
converter dust and retort waste are produced as solid wastes during smelting
of zinc and lead ores. These wastes used to be dumped, but now the zinc is
recovered."6 Converter dust is a mixture of zinc, lead, iron, and copper
oxides. Leaching with a sulfuric acid solution removes the copper;
addition of a bleaching powder precipitates the iron. The remaining zinc
sulfate reverts to zinc through electrolysis. Retort residues,
consisting of unconsumed coal, coke, and unreduced ore from incomplete distilla-
tions, may contain 5-15% zinc. Because the composition varies so greatly,
each residue must be separately analyzed to determine the best method of
treatment.
88
-------
Many foundry and metallurgic operations produce wastes that can be
treated to recover zinc. Skimmings from galvanizing operations may contain
20% metallic zinc and 35% zinc chloride; the zinc drosses that form
at the bottom of galvanizing vats may yield intermediate grades of zinc;
crude zinc oxide can be recovered from the leaching vats in which skimmings
from galvanizing are treated. Zinc ashes and flue dust from foundries and
smelters can be mixed with zinc ore and be distilled into slab zinc, or they
can be used to make oxides and pigments; die-cast drosses and turnings may
contain 40-70% recoverable zinc.^^ The reclaimed zinc products are used
for galvanizing vat charges, producing rubber, making paint pigments and
ceramic glazes, and for rolled zinc.
Trace amounts of unrecoverable zinc will still remain in waste material
even after reprocessing. This material frequently is deposited in municipal
sewer systems and treated with the rest of the sewage. If the sludge from
the sewage treatment is dried and incinerated, trace quantities of zinc
may be released into the atmosphere. Zinc oxide is also a potential effluent
from the incineration of vinyl products in which the zinc was used
as a heat stabilizer. Data on the concentration of zinc emitted from one
stack of a twin-stack municipal incinerator are set forth in Tables 4-16 and
4_17.1813
89
-------
TABLE 4-16
Metals Analysis of Samples Extracted from Various Points
of One Stack of a Two-Stack Municipal Incinerator^
Cadmium
Lead
Metal Concentrations
Zinc
U8
yg/g yg
yg/g
yg
yg/t
Copper
yg
yg/g
\o
o
Probe backwash,
suspended
solids, 0.0192 g
dissolved
solids, filtrate
Glass fiber
filter, 0.2097 g
Impinger
contents
345.5 ml
TOTAL
0.845 44 124.9 6,507 78.2
00 0 0 —
401
4,077
0.04
6.91 0.02 0 0
408.8 16,413
0.47
..IxlO5
8.18 426
0 0
1,912 16,288 77,677 l.lxlO5 556,340 319 1,519
0.04
327.2
•%>ata from Yost et al.1813
-------
TABLE 4-17
Metals Emission Rate from One Stack of a Two-Stack Municipal Incinerator-
Metal Mass Emission Rate, g/sec—
Cadmium 0.017
Lead 0.683
Zinc ' 4.580
Copper 0.014
1813
•Data from Yost et al.
' 3 3
%ig/sec = yg/m x m /sec
91
-------
Nuclear Reactors and Zinc-65
A major component of any nuclear reactor Is the coolant material, which is
essential in removing the heat released by the fission process. Water is a
popular coolant because it can also serve as a moderator to slow down neutrons.
Although the water is purified before circulating through the reactor, small
amounts of trace elements dissolved in the water are not removed. Also,
corrosion products from reactor construction materials and feedwater systems
are present. These impurities concentrate on the fuel cladding surfaces where
they are bombarded by the intense neutron flux in the reactor and become radio-
active. Zinc-65 is one of the radioactive contaminants formed in reactor coolant
water in this way, particularly in those reactors in which admiralty brass is
used in the feedwater system. Zinc from aluminum alloys also provides
n
the target material. The nuclear reactions involved are copper-63 ( EtfT))
zinc-65 and zinc-64 (n,^)^ zinc-65.
The radioactive material is subsequently removed from the coolant water
by ion exchange. Since it is generally classed as low-level, that is, it
contains 10 uCi/ml or less of activity, "^ this material may be contained
for a brief period of time to allow shorter-lived radionuclides to decay;
then it is discharged into a system that allows dilution of the remaining
radioactivity. Discharge into rivers, lakes, and oceans provides ample dilution
of the radioactive waste to below the maximum permissible limits. It is in this
manner that waste containing zinc-65 is handled.
+2
When it is discharged from a reactor, zinc-65 is a soluble cation, Zn
If discharged into a river, it becomes associated with particulate matter in
the water. Studies of the transport of zinc-65 by the Columbia River from
Hanford reactors to the Pacific Ocean reveal that changes in physical form
and concentration occurring in transit are caused primarily by adsorption on
92
-------
suspended sediments. Most of these suspended particles settle in the pro-
1977
tected and slack water regions of the river. Perkins et_ al., i7 sampling
Hanford reactor effluent water from the Hanford reactors and river water at
three downstream river locations, demonstrated an increased percentage of
zinc-65 in particulate phase from 1.8% at the point of discharge to 14% at
Pasco, Washington (56 km downstream) and 64% at Hood River, Oregon (336 km
downstream), to 76% at Vancouver, the third point (448 km downstream). They
also observed that total zinc-65 concentration at downstream locations de-
creased, except during spring and early summer when flooding caused scouring
of the river bed and resuspension of the deposited radioactivity.
Annual samples of average metal concentrations from 1966 to 1970 taken
at Richland, Washington and Bonneville Dam (48 and 384 km below the Hanford
reactors, respectively) also showed that zinc-65 in the water decreased with
distance because of settling (See Table 4-18). ^Oa Because all but one of
the production reactors were closed during the time tests were conducted, the
zinc concentration diminished from year to year.
As river water mixes with ocean water, the amount of zinc-65 activity
associated with particles decreases as salinity of the water increases (see
/ o *?
Figure 4-6). This effect and the distribution of dissociated zinc in ocean
water have been investigated by the Laboratory for the Study of Radioactive
Contamination of the Sea at Fiascherino, Italy. The laboratory has reported
that 10-20% of the naturally occurring zinc in sea water is in an ionic state,
30-50% is in particulate form, and 40-50% is in a complexed form. If the
ionic form of zinc-65 (65Zn ) is added, its distribution among these forms is
not equal. Exchange with the ionic and particulate forms is rapid, but even
after 2 years, exchange is incomplete with the complexed forms; complexed
zinc-65 is not in chemical equilibrium with the ionic and particulate zinc-65.
93
-------
0)
4J
«
1
so
zinc-65
16
Salinity, 7.
24
o
a
•4
10
8
6
4
3
J 1
zinc-65
1 1
1 1 1
*"• "Pafticulatc
~ —'—Soluble —
-X
' V
^^
f» >*.
^* * ^^
1 1 *""""'*
"fc*^»»«—
nn ~*~:'.-g>-
r.'fS.-74^Sj.--£K;x|.^
-
-
-
_
Is.
Salinity, %
FIGURE 4-6 Soluble, particulate, and percentage of particulate levels
in the Columbia River estuary in April 1968. Reproduced from Forster.4"92
94
-------
Observations of the Columbia River where it merges with the Pacific Ocean
have provided additional information on the fate of zinc-65 in estuarine and
marine environments. Jennings and Osterberg7^ reported in situ radioactivity
measurements of estuarine sediments and overlying water; more zinc-65 activity
was found in the sediments, and the highest radioactivity was observed in sedi-
ments in protected areas of upstream portions of the estuary where fine-grained
silts and clays would tend to settle from suspension. Gross570 and Osterberg,1181
comparing estuarine sediments with offshore sediments, found that radioactivity
decreased rapidly with distance from the mouth of the river and the surface of
the water.
This distribution of zinc-65 was further evidenced by radioactivity levels in
birds, fish, crustaceans, mollusks, algae, and plankton from along the Washington
and Oregon coasts.1738 Concentrations were highest in samples at the mouth of
the river, and they decreased very rapidly with distance from the river mouth,
QQO
as seen in Figure 4-7. Kujala analyzed 132 samples from 5 species of Pacific
salmon and noted distinct differences in radioisotope concentration by species
and area of capture. Chinook, chum, and pink salmon that spend most of their
ocean life in the Bering Sea and the central north Pacific had zinc-65 activity
less than 2.0 pCi/g, while Chinook and coho salmon that live within the
Columbia River plume areas had levels as high as 49.2 pCi/g and 59.3 pCi/g, re-
spectively. The highest level, 81.9 pCi/g, was found in Chinook salmon near
Eureka, California, which has a nuclear power plant that disposes low-level wastes
into the Pacific. Samples of sockeye salmon did not show any increase in zinc-65
activity close to the Columbia River; since these fish are thought to spend most
of their ocean li£e beyond the influence of the Columbia River, they would not
be expected to be appreciably influenced by zinc-65 from the river.
Radionuclides in the Columbia River water have been used as convenient
tracers to study the dispersion of the river water itself in the Pacific Ocean.
95
-------
COOS BAY.
ORE.
(320)*
1957
1959
1960
1957
BOILER BAY,
ORE. 1959
060)*
1960
SEASIDE,
ORE.
(19.2)*
ILLWACO,
WASH.
(0)*
1957
1959
1960
1957
1959
1960
-
S//S/////S//y7/7//YS////////r/////S//S///s'7f/r/ft
WSMft^aWxiWaiSK
iS&S&SSi^^
LONG BEACH. 195?
WASH. 1959
(12.8)*
1960
1957
WILLAPABAY,
WASH. 1959
(57.6)*
1960
1957
KALALOCH,
WASH. 1959
(152)*
1960
DUNG EN ESS. 1957
S5T-
1960
E.
?777\ Algae (Fucus Sp.)
t^a Mussels (Mytilus Spp.)
i i Oyster (Crassostrea gigas)
•• Razor Clam (Siliqua patula)
I
I
I
I
I
I
I
•Distance from river fj 20 40 60 80 100 120 140 160 180
mouth in kilometers
At/uc/9 WET WEIGHT
FIGURE 4-7 Concentration of zinc-65 in marine organisms near the mouth of the Columbia River.
Reproduced from Watson et al. 1738.
96
-------
Since zinc-65 is taken up by most aquatic organisms and is usually the predominant
Y-emitting radioelement found in biota samples, mapping the distribution of
zinc-65 by analysis of samples of marine organisms has been particularly useful
as an indication of the dispersion pattern of the river water. Such studies^92*1738
have shown that the dispersion follows two seasonal patterns: in summer, because
of winds from the north, the flow is offshore toward the southwest as far as 320
km; and in winter, the flow is inshore, north of the river mouth along the
coast of Washington, because of the southerly winds.
Research on the distribution and levels of zinc-65 has been prompted by a
concern for zinc-65 because it is radioactive, not because it is zinc. Although
levels of zinc-65 discharged in effluent are below permissible levels, the
potential exists that the radioactivity may be brought back to man in dangerously
high levels if it becomes concentrated through food chains. Many studies have
been undertaken to evaluate this possibility, including studies using controlled
food chains,72 experiments with oysters raised in reactor effluent,^397 an
-------
TABLE 4-19
Permissible Levels'2 for Nonoccupational
Exposure (Unrestricted Areas)**
Type of
Exposure _ Air, uCi/ml Water, yd/ml
Soluble zinc-65 4 x 1(T9 1 x 1(T4
Insoluble zinc-65 2 x 10~9 2 x 10~4
Atomic Energy Commission Standards.
Data from U.S. Atomic Energy Commission.16^3
98
-------
CHAPTER 5
ZINC IN PLANTS
AQUATIC PLANTS
Aquatic plants, especially algae, have been used extensively to
investigate the role of zinc in plant metabolism However, much of the
research involved laboratory investigations and less is known about zinc
in aquatic plants in natural systems. The research on aspects of zinc
in aquatic plants other than metabolism has been largely devoted to the
fate of zinc-65 as a radioactive contaminant in water systems. Hence,
much of the available literature deals with zinc-65 in aquatic plants.
A requirement for zinc in the alga Stichococcus bacillaris was demonstrated
421
in 1926, and the metal is now considered to be required by all algae for
normal growth and development. Besides the continuing research on metabolic
functions and uptake of zinc-65, work is now being done on the ability of
aquatic plants to concentrate zinc by several orders of magnitude from
ambient waters. Interest has developed in using aquatic plants to remove
zinc from contaminated waters as well as in the toxicity of zinc to the
T695
plants. Vinogradov demonstrated that plants living in the ocean remove
zinc from the water to produce zinc levels in the ocean which are more
acceptable for the normal growth of other organisms.
Zinc Uptake and Concentrations
Uptake. There are two separate mechanisms of zinc uptake by aquatic plants.
These are sorption processes (adsorption, absorption, and ion exchange) and
metabolic assimilation. Both mechanisms may exist in the same organism,
but the pathway utilized often is determined by metabolic requirements or
environmental conditions.
99
-------
Evidence was provided that the uptake of zinc-65 in algae was
63
directly related to the rate of photosynthesis and metabolically controlled.
The involvement of adsorption processes in zinc uptake by algae was confirmed
when the green alga Golenkinia paricispina was used to show that zinc uptake
involved ion exchange sites created by the photosynthetic removal of carbon
62
dioxide. Living and freshly killed algae were placed in the same
concentration of zinc-65 and zinc uptake was determined by zinc accumulation
in cells over time. Dead cells accumulated more zinc-65 than the living
algae. These results were confirmed in studies of freshwater phytoplankton
345,346
and periphyton. Such results indicate that sorption mechanisms
accumulate more zinc than is necessary for normal metabolic functions.
590
Gutknecht suggested that metabolic zinc uptake by algae may be a
secondary process that occurs only after sorption has taken place. Much of
1452
the zinc sorption takes place in the cell walls of aquatic plants.
The sorption processes are probably responsible for the large concentrations
of zinc which are found in algae and other aquatic plants when compared to
the zinc concentration of the ambient water.
Uptake of zinc by aquatic plants is influenced by the structures in
the plants. Algae are often covered by organic excretions that can function
146
as ion exchangers. Because of their suspended state, planktonic algae
and free-floating vascular plants can take up zinc from the water but not
from sediments. Benthie algae or rooted vascular hydrophytes which are
submerged in water may take up zinc from either the sediments or the
ambient water. Sediments are usually several orders of magnitude higher
in zinc than the ambient water, but the relative importance of sediment
and solution in supplying zinc to rooted aquatic plants is unknown.
100
-------
Concentration. The presence of zinc in aquatic plants has been established
123,646,1695 0
for some time. Some of the earliest analyses of aquatic plants
OT Q
which showed traces of zinc were conducted in 1919. It was variously
demonstrated that the concentration of zinc in brown seaweed (Fucus vesi-
_. t 123,196,591,1815
culosus) varies with the concentration in the seawater.
187,188
This observation was confirmed for another brown seaweed, Laminaria digitata.
275
Similar results had been obtained earlier.
187
Bryan, in studies with Laminaria digitata, found 2.2 mg zinc/1
of seawater and a growing tissue concentration of 5.4 mg zinc/g, resulting
a concentration ratio f mg zinc/g tissue ]of zinc between plant and
Vitag zinc/g ambient solution"'' ^23
seawater to be 2,455. Earlier studies by Black and Mitchell produced
lower concentration factors, listed in Table 5-1. In a review of mineral
1163
nutrition of algae, O'Kelley reported zinc-65 concentration factors
in marine algae ranging up to 1,200. However, concentration ratios for
344a
zinc-65 in Columbia River plankton were reported to range from 300-19,000.
649
Similar high concentration factors were reported for freshwater algae.
In later research with brown seaweed, a tissue concentration factor of
146
3,240 for zinc was reported. Bowen cited ratios found by Russian
workers who studied the uptake of zinc-65 by 32 species of algae, bryophytes,
and vascular hydrophytes. The concentration factors usually ranged between
2,000-4,000 in tissue as compared to ambient water, where concentrations
259
were low. Chapman et al. found similar conditions determining zinc
149
concentration factors. Boyd found that certain aquatic plants concentrated
zinc from the environment in early stages of growth, but uptake decreased
as the growing season progressed. Boyd's work does not necessarily disagree
101
-------
TABLE 5-1
Concentration Factor, or Ratio of Zinc Content in Algae
6L>
to Zinc Content in Seawater
Species
Concentration Factor
mg zinc/g tissue
mg zinc/g solution
Pelvetia canaliculata
Ascophylltun nodosum
Fucus vesiculosus
Fucus serratus
Laminaria digitata frond, Atlantic Bridge
Laminaria digitata stipe, Atlantic Bridge
Laminaria digitata frond, Ardencaple Bay
Laminaria digitata stipe, Ardencaple Bay
1,000
1,400
1,100
600
400
600
1,000
900
from Black and Mitchell.
123
102
-------
with the research denoting a continued uptake throughout the growing
season, because variations in uptake rate within a specific growing season
were seldom determined. That aquatic plants can concentrate zinc from
ambient water is well documented.
The content and concentration factor for zinc in algae grown under
591
similar conditions was shown to be species-dependent by Gutknecht.
How concentrations of zinc may vary by species is illustrated in Table 5-2.
Much of the difference in zinc concentration factors by aquatic plants
146
is caused by differences in the ion exchange properties of their surfaces.
One of the primary sites of zinc binding in aquatic plants is in the cell
1452 991
wall. In studies with Chlorella fusca, Matsku and Broda concluded
that some zinc-65 taken up by the organism can be removed by exchange with
unlabeled zinc, although another fraction is more tightly bound. Exchange-
able zinc is probably in the free space, whereas the bound zinc is in the
protoplast. There are probably marked differences in the metabolic
assimilation of aquatic plants as well as in their ion exchange properties.
Zinc concentrations assimilated by freshwater and marine organisms reported
the - __ researchers
in/ literature before 1964 have been summarized, and other/ have studied
zinc concentration in aquatic plants and reported results similar to those
f „ , , „ 105,512,908,1339,1815
of Matsku and Broda.
The seasonal changes in zinc concentrations of aquatic plants must
be considered when data are compared and interpreted. Table 5-3 contains
data on zinc variations by season in several species of seaweed.
Considerable variation in zinc content was evident in some species
with different seasonal samplings, but other species contained relatively
constant amounts of zinc through all seasons. Sampling techniques probably
103
-------
TABLE 5-2
Uptake and Concentration of Zinc-65'
and Total Zinc in Seaweeds
Species
Ulva lactuca
Codium decorticatum
Fucus vesiculosus
Dictyota dichotoma
Gracilaria foliifera
Agardriella tenera
Hypnea musciformis
Zinc-65
Concentration Factor
290
30
3,300
280
210
395
150
Total Zinc,
Fresh Weight
23.80
0.96
124.00
5.70
5.83
9.78
3.54
rag/kg
Dry Weight
158.0
17.0
472.0
35.0
37.7
91.4
23.2
a- 591
Data from Gutknecht.
1Q4
-------
TABLE 5-3
Seasonal Variation in Zinc Content
0,
of Oven-Dried Seaweeds
Sample
Felvetia canaliculata
Fucus spiralis
Ascophyllum nodosum
Fucus serratus
Fucus vesiculosus
Laminaria digitata frond
Laminaria digitata stipe
Laminaria coloustoni frond
Dates Sampled
1/12/49 5/26/49 6/27/50
mg zinc/kg dry weight
40
103
79
99
no data
117
47
62
60
70
60
64
62
76
90
no data
116
63
105
59
92
a 123
D?ta from Black and Mitchell.
105
-------
account for some of the seasonal variability. However, data have been
presented which confirm, a seasonal fluctuation in zinc content of Fucua
196 859a
veslculosua. Knauer and Martin found that the seasonal variations
of zinc in both water and phytoplankton in Monterey Bay, California, were
related to the upswelling of the ocean and possibily to the availability
of zinc from organic matter on the ocean floor. Other biologic, chemical,
and environmental factors that can Influence the seasonal variation of
zinc in aquatic plants are growth rate, supply of other nutrients, and
temperature.
Zinc toxicoses in aquatic plants are distinct possibilities in areas
where vine-contaminated surface waters exist. Complete destruction of
aquatic plant communities because of zinc toxicosis in areas associated
110
with mining or processing of ore was reported by Besch and Roberts-Pichette
in their study on the effects of zinc mining pollution in the Miramlchi
River system of northern New Brunswick, Canada.
527,1769
A laboratory bioassay approach was developed to determine
critical levels of zinc that would be potentially toxic to many aquatic
plants. The sampling, analysis and correlation studies that have been
completed show promise, but variables such as pH, light, temperature,
and the presence of other species can influence the reliability of
384
a critical metal level. Dietz conducted bloassays on mosses to assess
the trend of water pollution by metals and found that the mosses were
considerably enriched by zinc because the water had been contaminated
by it. The amount of enrichment also was dependent upon plant species
527
and specific plant selectivity. Gerloff used plant analysis to
106
-------
evaluate nutrient supplies and growth-limiting nutrients for the aquatic
weeds Elodea occidentalia and Ceratophvllum dejmuraum in lakes and streams.
The critical concentration of zinc was 8 ppm in the second 2.5-cm segment of
main branches and laterals of Elodea occidentalis.
1540
Sprague used a bioassay that established a critical level of
0.75 mg zinc/1 of ambient water as a value which caused lethal effects in
aquatic plants. The 0.75 mg zinc/1 value applied only when zinc was in
an ionic form; the presence of nitrilotriacetic acid (NTA) also
reduced the toxiclty of the zinc to the plants. Therefore, the form in
which zinc exists in solution is an Important factor in evaluating potential
zinc toxicity to plants.
1615
Tlerney did not find a clear relationship between concentration
of zinc in Elodea anadensls and Ceratophyllum demursum and the zinc content
of the ambient waters, and he observed that more detailed field and laboratory
studies were needed to identify the specific factors which control availability,
uptake, and accumulation of zinc by aquatic plants.
Factors Affecting Zinc Uptake and Accumulation
Temperature. Variations in temperature significantly influence the metabolic
366
rate of aquatic organisms. Organisms which obtain zinc through food
conuumption were shown to increase their zlnc-65 accumulation as their rate
338,637,1332
of food consumption increased at elevated temperatures. Similar
Increases In zinc uptake by aquatic plants can be expected as rates of
metabolism increase with increasing temperature of ambient water.
107
-------
590,592
pH. Zinc uptake and accumulation by algae is dependent upon pH. Gutknecht
found accumulation of zinc-65 by Ulva lactuca to be twice as high at pH 8
as at pH 7 and three times as high at pH 9 as at pH 8. These data and later
research, in which zinc uptake was found to follow the Freundlich adsorption
equation, demonstrated that pH-dependent adsorption exchange
591 183
was an important mechanism of zinc uptake by aquatic plants. Brungs
observed that the solubility of zinc-65 decreased with increasing pH in
freshwaters. These observations were confirmed by the calculation of
solubility products, but the precipitation of zinc would be low enough to
provide adequate zinc for aquatic plant growth and development at near
neutral pH values.
Ambient concentration of zinc. Accumulation of zinc in aquatic plants
123,187,
was found to be directly related to the amount in the ambient water.
188 196 591 1815
The log of the zinc concentration in Chlorella cells was
discovered to be proportional to the log of the zinc concentration in the
860
medium over a hundredfold range in concentration of zinc. Halving the
ambient concentration of zinc-65 available to Chlorella resulted in a 50%
345
decrease in uptake rate and total amount accumulated. Previously
62 592
Bachmann and Gutknecht had reported similar results with certain
other algae species, and similar relationships have been demonstrated for
392
mercury in vascular aquatic plants.
Competing.-ion. The presence of other cations in waters containing zinc
have been shown to reduce the amount of zinc taken up by aquatic plants.
108
-------
62
Bachmann found that zinc-65 uptake In the green algae, Golenkinia, was
reduced when concentrations of calcium, magnesium, potassium, and sodium
345
were increased. Gushing and Rose reported that zinc-65 uptake by periphyton
was reduced 50% when the amount of magnesium in solution was doubled. Also,
doubling the stable zinc concentration decreased the zinc-65 uptake by 25%
(probably caused by isotopic dilution). The competition between zinc-65
and other cations for available binding sites on the plants is responsible
345
for such occurrences.
Light. If minimum requirements for photosynthesis are met, increased light
590,592
intensity will cause little increase in zinc uptake or accumulation.
Functions of Zinc
1296
Zinc functions primarily in plants as a metalloenzyme. Several
zinc-requiring dehydrogenases, proteinases, and peptidases have been identified
1295
in various organisms, including aquatic plants. Price reported that
several dehydrogenases were sensitive to zinc deficiency in Eugjlena gracilis.
Therefore, the metabolism of aquatic plants can be influenced markedly
by changes in the availability of zinc. Data indicate that zinc deficiency
rapidly reduces the amount of RNA as well as the ribosome contents of
ly be as a
1289,1296
1296
cells. Another function for zinc in plant metabolism may be as a
stabilizer of the cytoplasmic ribosomes of Euglena gracilis.
It also has been suggested that zinc aids in auxin B production,
but a definite connection between zinc and indolacetate has not been
1296
determined. For additional information on the metabolic role of
zinc in connection with metalloenzyme activity, see Chapter 9.
109
-------
TERRESTRIAL PLANTS*
Reports of the use of zinc as a nutrient in plant fertilizer
790 157
appeared in the literature as early as 1912, and Brechley described
zinc deficiency in higher plants in 1914. That zinc was essential to
1523
plant life was established by Sommer and Lipman when they reported
that zinc was required for three plant species. Zinc was discovered to
be necessary for trees when zinc deficiency was identified on fruit trees
255
in California, and when pecan rosette (a malformation of the new leaves
14
of growing tips) in the South was found to be caused by zinc deficiency.
Knowledge of the nature of zinc made the identification of many nutritional
problems in higher plants possible. Zinc deficiency is now the most
938
common micronutrient deficiency in the United States. Using zinc
fertilizer to improve crop growth also is well established. Although it
is possible for zinc toxicity to be a problem to plants, zinc deficiency
in plants is far more likely than toxicosis. Crops that are sensitive to
zinc deficiency are citrus and deciduous tree fruits, pecans, pine (in
Australia), grapes, kidney beans, hops.soybeans, corn, lima beans, flax,
castor beans, and onions. Cotton, potatoes, tomatoes, alfalfa, clovers,
sorghum, sudan grass, and sugar beets are mildly sensitive to zinc
deficiency, and peas.small grains, peppermint, asparagus, mustard and
other crucifers, forage grasses, safflower, and carrots are not
1686
sensitive to it.
*The research discussed in this section appeared in the literature before
April 1975.
110
-------
Availability of Zinc in Soils
Climatic and soil conditions responsible for zinc deficiency in crops
956
in the United States were reviewed in detail by Lucas and Khezefc. Zinc
1387
deficiency in European countries was reviewed earlier. Some important
factors associated with or contributing to zinc availability in soils are
938 956
summarized below. Further details may be found in two reviews. *
Soil low in zinc reserves. Quartz is low in zinc, so sandy soils often
938
contain only 10-30 ppm total zinc. A similar observation has been
reported for peat and muck soils, where zinc in the plant root is separated
from the mineral reserves below the organic surface layer. Under acid
conditions, zinc may be rapidly leached from the soil where rainfall is
956
great.
High soil pH. Most disorders in plants caused by zinc deficiency occur in calcareous
soils with a pH of 7.4 or higher, because the solubility of zinc decreases
as soil pH increases. The total zinc content of calcareous soils is often
equal to or higher than that of noncalcareous soils. In addition to
precipitation at high soil pH values, adsorption of zinc by carbonates
may contribute to low availability of zinc to plants on certain calcareous
939,956
soils.
Limited root zones. Zinc deficiencies are frequently found in soils with
restricted root zones. They may be caused by hardpans, high water tables,
or other factors. Equipment passage may compact soil and cause zinc defi-
956
ciency in certain agricultural soils. The root development of rice
111
-------
grown on flooded and nonf looded soils in Alabama determined the amount of
535
zinc available to plants.
Microbial fixation and low soil organic matter. Zinc deficiency in
beans and corn on calcareous soils is often observed on old corral sites
and barnyards and has been attributed to the rapid growth of microorganisms
956
that may tie up available zinc. Zinc deficiency in corn is sometimes
more severe when it has been planted after a crop of sugar beets than when
938
planted in soil used for corn or many other crops. DeRemer and
378
Smith observed that plowing down sugar beet tops reduced available
zinc to the succeeding bean crop. Therefore, organic matter or soil
microorganisms help determine the zinc fixation by soils. Zinc deficiencies
are often reported in areas where surface soil containing organic matter
572 938
has been removed. Lindsay concluded that deficiencies increased
because the exposed subsoil was lower in organic matter and higher in pH
and carbonate than the surface soil. Indeed, a close correlation
487,
between soil organic matter and extractable zinc has been well documented.
More observations of zinc deficiencies were reported in Michigan
956
where the surface soil was disturbed and the subsoil exposed.
Soil temperature and moisture. Cool, wet spring weather often aggravates
956
or induces zinc deficiency in field crops. Limited plant root growth,
which brings about a limited root feeding zone and reduced microbial
activity so that zinc is not released from organic matter during the cool,
1719
damp weather, may cause such deficiencies. Wallace, et al. have found
112
-------
86
that zinc uptake increases with temperature. Bauer and Lindsay concluded
that decreased solubility of zinc in the soil,rather than a biologic effect^
was the primary reason for more pronounced zinc deficiencies in cool weather.
523
Additional research supported those results. Soil moisture and aeration
432
influence zinc availability through indirect soil reactions and root
metabolism.
High phosphate levels in soil. A phosphorus-induced zinc deficiency has
. 131,435,523,977,1121,1384,1465,1686 A ,. ,
been widely reported. Application of
phosphate fertilizer and high phosphorus levels in soils have been linked
to zinc deficiency problems for many years. ' Since zinc extractable
from soil is not decreased by the addition of phosphorus to soils, the
formation of insoluble zinc phosphate compounds in soils
939,1168
is probably not the cause of a phosphorus-induced zinc deficiency.
The deficiency may result from the formation of an iron phosphate at the
root surface that excludes zinc uptake where soil zinc levels already
. 432
are low.
Influences of nitrogen in soil. The acidifying influences of nitrogen
fertilizers — which make zinc solubility greater and possibly increase
the cation exchange capacity of roots when the nitrogen supply increases —
132,396,939,1687
have been cited as causes of enlarged zinc uptake in plants.
Movement of Zinc to Plant Roots
Zinc is believed to move through the soil to plant roots by mass
938
flow (convection) and diffusion. Mass flow is the movement of nutrients
113
-------
along with the soil solution, whereas diffusion is the movement of nutrients
1775
through the soil solution because of a concentration gradient. (Olsen
1169
and Kemper provide a detailed review on movement of nutrients to plant
73,74,238,290,371,425,426,434
roots.) Various workers have shown that
diffusion gradients are formed between the soil solution and plant roots
and that most zinc movement to plant roots is by diffusion. Warncke and
1733-
Barber have explained how the rate of zinc diffusion in soil is controlled.
1735
Soil moisture, bulk density, and zinc adsorption are all important
1776
in determining final zinc diffusion rates. Wilkinson et al. have
reported that tripling the transpiration rate in wheat seedlings did not
yield an increased uptake of zinc, and concluded that zinc movement to
plant roots was independent of mass flow and dependent upon diffusion.
607
These results were confirmed for several species. The movement of
zinc to plant roots is dependent upon soil solution concentration and the
938
ability of soil to replenish zinc removed from solution. Lindsay and
941
Norvell reported that increasing soil pH decreased solubility of zinc
939
in soils, and Lindsay stated that the diminished zinc concentration
and subsequent reduced concentration gradient would reduce zinc availability
1028
and uptake by plants. Melton et al. have confirmed the influence of
soil pH on zinc diffusion and found a limited influence of phosphorus on zinc
diffusion.
The effects of chelates upon zinc solubility and availability to
_ . J 170,174,175,212,372,425,426,574,712,938,
plants have been well documented.
941 1149 1717
' ' Chelating agents increase the solubility of zinc in soils
938
and increase the movement of zinc to plant roots by both mass flow and diffusion.
114
-------
The ability of the plant root to compete with the chelating agent for the
zinc will determine if the actual amount of zinc uptake by plants will be
increased or decreased from chelation. Adding chelating agents to soils
will usually increase zinc availability to plants because the solubility
is increased; but the addition of a chelating agent to nutrient solutions
938
may reduce the uptake of zinc that is already in solution.
Zinc Uptake, Translocation, and Concentration by Plants
938 1078
Uptake. Zinc absorption by plants has been reviewed, ' and factors
influencing plant absorption of zinc have been identified. ' Ranges
of zinc uptake from 2-4,000 ng/day on a fresh weight basis
239,1428 695
have been reported. Hewitt reported optimal levels of zinc
for plants growing in nutrient solution to range from 0.3-3.0 yM, Carroll
238
and Loneragan observed that a variety of plants grew well at 0.01 yM
with maximum growth at 0.25 uM zinc. Zinc absorption by sugar cane
147
has been investigated with similar findings.
Considerable uncertainty still exists about the roles of active
938,1078
and passive mechanisms in zinc uptake by plants. Conclusions of
workers who failed to separate passive exchange absorption from active
166,441,1078,1317,
cellular accumulation are the source of much conflict.
' For example, some researchers of zinc translocation found that
xylem exudates from tomatoes and soybeans contained higher concentrations
27,1617
of zinc than did the nutrient solution in which the plants were grown.
Such results suggest an active accumulation component in the plant if it
is assumed that the zinc in the xylem exudate is present in a form similar
115
-------
to that in the external nutrient solution. Most of the evidence based on
study of metabolic inhibitors points toward an active accumulation component
536
crucial to zinc uptake.
Translocation. Zinc is translocated from the roots to the tops of
plants through the xylem, and a limited amount of zinc is retranslocated
938,1617,1618,1721
from the leaves through the phloem. (For a review of
1618
the translocation in plants, see Tiffin. ) It is intermediately mobile
938
within plants compared with other micronutrients. Zinc is localized in
238
plant roots when zinc supply is normal, but it moves from roots to plant
984,1340
tops when in limited supply. Also, some plants increase the
efficiency of the zinc they have stored by redistributing it from seeds
200,938,1088,1618
to leaves and from older to newer plant tissue.
1168
Olsen reviewed the interactions of zinc and other elements in
plants. Generally, zinc and phosphorus interactions reduce the uptake
111,113,169,827,1084,1502,1559,1604
and translocation of zinc by plants.
Also, a high level of available nitrogen often causes increased zinc
deficiency. Much of this deficiency was attributed to increased formation
of zinc-protein complexes in plant roots that inhibited normal transloca-
1168
tion to plant tops. Excess copper or iron in a plant also will bring
about intensified zinc deficiency. The influence of zinc in suppressing
copper uptake and translocation by plants usually is greater than the
263
suppressive effect of excessive copper upon zinc translocation.
786
Iron levels were reported to be high in zinc-deficient sweet corn.
The application of phosphorus decreased zinc concentrations in plants and
116
-------
26
plants low in zinc were high, in iron. Ambler and Brown have observed
that lowering zinc concentration in solution has increased the translocation
of iron and phosphorus into bean leaves. Because the iron and phosphorus
consistently increased in bean plants, the phosphorus evidently did not
1737
inhibit iron translocation. Similar findings have been reported for corn,
and decreasing the amount of zinc applied increased the phosphorus uptake
nearly sevenfold in leaf tissue, but only threefold in stems and roots.
Zinc uptake and translocation in tomato, soybean and squash plants
960
were increased when the salt concentration of the growth medium was increased.
Hence, there may not be an antagonistic relationship between soluble salts
and zinc in nutrient culture. Nor has additional research on iron, man-
452
ganese, and zinc interactions (using isotopic techniques) been conclusive.
Macronutrient cations, as well as copper and hydrogen, suppressed zinc uptake
and translocation by wheat seedlings, but iron and manganese had no
9fi1 9fi9
effect on zinc uptake. ' Therefore, the evidence for zinc interactions
with various elements is not solid and varies according to plant species
and other minerals present.
For example, Brown e£ al. reviewed the differential susceptibility
of various plant genotypes to zinc deficiency. A study of zinc in temperate
crops and pasture grasses revealed that species differ in their ability
540
to take up and utilize zinc. Similar findings have been reported for
navy beans,431'803'1258'1317 cats,209 rice,534 soybeans,1716 sugar cane,128
and wheat.1 A definite genetic basis for zinc uptake, distribution,
603,984
and utilization by corn plants was confirmed by others.
117
-------
1417 800
Concentration. Sauchelli and Jones summarized ranges of zinc
concentration in plants. When zinc drops to levels below 20 ppm dry weight
in leaves, deficiencies may occur. The normal concentration range is 25-150 ppm.
Zinc and toxicoses often occur when levels exceed 400 ppm. Zinc
concentration ranges for several specific plant species have been
85,258,540
reported. The concentration of zinc in plant tissue usually is
greatest in young plants; it decreases because of dilution and redistribu-
550,799,1088,1341 _
tion as the tissues grow and mature. Crops were
1689
classified into three groups by Viets et al. according to their zinc
concentration. Table 5-4 sets forth the variety of zinc concentrations
possible which do not produce visible symptoms of deficiency or toxicosis.
Zinc Disorders in Plants
Deficiency. Zinc deficiency symptoms have been described in detail for
1417 1612
several plants, and zinc deficiency and its control have been reviewed.
Because zinc is intermediate in mobility within the plant, a variety of
symptoms, including chlorosis, stunting, and necrotic tissue may indicate
zinc deficiency. In corn and sorghum plants, chlorotic streaks appear in
older leaves. The older leaves of navy beans and soybeans turn yellow, and
the entire plant becomes stunted and fails to produce a normal crop when
the deficiency is severe. In citrus, peach, pecan, and tung frees, irregular
chlorosis and bronzing occurs and the new growth becomes stunted. Severe
deficiency may kill the leaves and twigs and the growing tip may be deformed,
exhibiting a symptom known as "resetting."
118
-------
TABLE 5-4
Zinc Content of Crop Plants^
Crop Plant
Zinc, ppm
Deficient Low Normal High Toxic
Corn leaves, vegetative stage
Soybean leaves, vegetative stage
Wheat, barley oats, vegetative stage
(7.5-30 cm)
Cotton, vegetative stage
Tobacco, vegetative stage
Sugar beets, vegetative stage
Potatoes
Alfalfa tops
Grass, vegetative stage
0-10 11-20 21-70 71-150 150
0-10 11-20 21-70 71-150 150
0-10 11-20 21-40 41-150 150
20-30
0-20 21+
0-10 11-20 21-70
0-16 17-40
0-8
70+
30+
9-14
15-80
Beans (leaves)
Totatoes (leaves)
Citrus (leaves)
Florida
Tung (leaves)
Apples (leaves)
Peaches (leaves)
Pears (leaves)
Grapes (petioles)
0-20
0-10
0-15
0-10
0-15
0-16
0-10
—
—
11-20
16-25
11-26
16-20
17-20
11-16
0-30
—
21-120
26-80
—
21-50
21-50
17-40
31-50
—
121+
81-200
—
51+
51+
41+
51+
—
—
200
—
—
—
—
—
aDerived from Sauchelli14^ and Bishop and MacEachern.^O
119
-------
Symptoms of zinc metabolic disorders also are expressed cytologically
and morphologically. There are more free amino nitrogen and amides in
1267
zinc-deficient plants than in healthy ones. Inorganic phosphate
also is higher in the region outside the stele and in the phloem of
1321
deficient stem tissue. In many plant species, zinc deficiency is
258
shown by interveinal chlorosis because the chlorophyll
1453 1464
formation has been disrupted. According to Seatz"3nd Jurinak,
the palisade cells of leaves are larger and transversely divided,
rather than columnar; reduction in number of chloroplasts; the
absence of starch grains; the presence of oil droplets in the
chloroplasts and the presence of calcium oxalate crystals and
accumulation of phenolic materials in the leaves.
are morphologic changesvthat plants undergo when deficient ia ziae,
Diagnosis and correction of deficiency. Plant and soil analyses can be
useful tools in diagnosing zinc deficiency in many plants. A current
review which includes soil testing and plant analysis techniques for zinc
1724
has been edited by Walsh and Beaton. Zinc deficiency usually is
suspected if plant tissue concentrations are below 20 ppm zinc, but the
critical level varies with species, stage of growth, and plant part
1026
sampled. Levels of zinc in solutions obtained from soil extractions
with acids or chelating agents have been correlated with the occurrence
of zinc deficiency. Usually, about 50 ml of solution is used to extract
zinc from 10 g of soil. Use of 0.1 N hydrochloric acid has been
accepted for several years, but solutions containing ethylenediaminetetraacetic
acid (EDTA) and diethylenetriaminepentaacetic acid
(DTPA) have also been used. Critical zinc levels in
the extracting solution that have been related to zinc deficiency in plants
21,1724 631
range from 0.5-2 ppm zinc. Hag and Miller have compared four
120
-------
zinc extractants on 65 soils and found EDTA and DTPA extractions to be
445
most useful in predicting zinc deficiency. But Evans et al. found few
differences in correlations between zinc uptake and soil zinc extracted
with several extractants. The reliability of soil tests to predict zinc
availability in soils is dependent upon many soil and plant factors and
no single extractant works best in all situations.1623 Table 5-5 summarizes
recommendations of zinc fertilizer applications on specific crops by state.
With most plants, zinc deficiency can be overcome by soi,l or foliar
927,955
application of zinc salts. Zinc sulfate or finely powdered zinc
oxide are the inorganic zinc salts usually applied. However, Wallace
1718
and Romney have used zinc sulfate, alnc NTA and zinc EDTA with great
success; seven different zinc sources were evaluated extensively and found
to be effective during the first few months after application. However,
the zinc apparently reverts to forms of equal availability after 1-3 yr.
Banding the zinc fertilizer 2.5 cm to the side of and 5.0 cm below the seed
801,802
provides the most efficient use per pound of zinc by responsive crops,
and banded and broadcast applications of zinc to soils for crop response have
963,1693
been successfully used. Zinc has been applied to corn seed at
980
planting time without injury, and the plants have responded to the
zinc fertilizer, but many crops may suffer from salt toxicosis during
germination. Applying zinc in the form of synthetic chelates increased
128,1693
zinc efficiency from two*to fivefold compared to zinc sulfate.
But.certain zinc chelates are relatively soluble and can be leached out
892
of the plant root zone when the soil is excessively moist. Most crops
use less than one kilogram of zinc per hectare of soil per year.
121
-------
TABLE 5-5
Recommendations for Applying Zinc Fertilizer
State
Alabama
California
Colorado^
Florida
Georgia
Indiana
Iowa
Kansas
Kentucky
Louisiana
Maryland
Michigan
by_
Crops
pecan
corn
all tree crops and
field crops
corn
tree crops
vegetable and field
crops
ornamentals
pecans
peaches
corn
corn
corn, soybeans
corn, sorghum
corn
citrus, pecans, tung
corn
beans
State0
Recommendations for Zinc
1.8 - 2.25 kg ZnS04/tree/yr
.11.25 kg ZnS04/ha
tree: 1.02 - 2.96 kg ZnS04/100 liter water
field: 11.25 - 22.5 kg ZnS04/ha
5.63 - 11.25 kg ZnS04/ha
360 g ZnS04, plus 120 g lime/100 liter water
90 - 225 g ZnO every 4-5 yr
90 - 270 g ZnO every 4-5 yr
0 . 9 kg ZnS04/cm trunk diameter
0.9 - 1.35 kg ZnS04/tree
11.25 - 22.5 kg ZnS04/ha
0.9 kg ZnS04/ha applied as plow down
11.25 - 22.5 kg ZnS04/ha
11.25 - 16.88 kg ZnS04/ha
4.5 kg ZnS04 in row
360 g ZnS04, plus 120 g lime/100 liter water
11.25 kg ZnSO,/ha
2.25 - 4.5 kg ZnSO./ha
122
-------
TABLE 5-5 (Continued)
State
Minnesota*7
Mississippi
Montana*
Nebraska*
North Carolina
North Dakota*
Oklahoma*
Oregon*
South Carolina
Tennessee
Texas
Utah2*
Crops
corn
pecans, tung,
corn
cherries, apples
castor beans, corn,
fruit, sugar beets,
soybeans
peaches
corn, potatoes
pecans
corn, lima beans
all tree fruit
cherries
pear
barley
corn
pecans
vegetables
corn
pecans
fruit
Recommendations for Zinc
11.25 kg ZnS04/ha
2.25 - 4.5 kg ZnS04/tree
11.25 kg ZnS04/ha
16.88 kg ZnS04/ha
5.63 -22.50 kg ZnS04/ha
44.2 g ZnS04/100 liter water every 3 wk
16.88 kg ZnS04/ha
4.5 kg ZnS04/ha
5.86 - 12.38 kg ZnS04/ha
204 g/100 liter water
1.42 - 1.77 kg ZnS04/liter water
590 g ZnS04/ioo liter water
16.88 kg ZnS04/ha
11.25 kg ZnS04/ha
4.5 - 5.4 kg ZnS04/mature tree
5.63 kg ZnS04/ha
11.25 - 22.5 ZnS04/ha
foliar: 360 g ZnS04/100 liter water
11.25 kg ZnS04/ha
1417
?Derived from Sauchelli.
Deficient levels are less than 16-20 ppm zinc in plant, oven-dry basis.
123
-------
Thus, 1.35-4.5 kg/ha inorganic zinc salts/yr gradually builds up the zinc
reserves in the soil. Adding 28.13 kg zinc/ha as zinc sulfate just once
provides an adequate zinc reserve for 10 years of maximum crop production
1693
on severely zinc-deficient soil. Similar residual effects of zinc
129 6*52 9^8
fertilizer have been reported. * ' ' Foliar application of zinc
to plants, often used to provide supplemental zinc, is usually imposed
927,1062 90
at rates of 1.13 kg zinc/ha/application. It also has been reported
that certain surfactants enhance foliar absorption of zinc. Once absorbed,
233 1795
zinc is not easily leached from vegetative parts of plant tissues .
130
Toxicosis. It is well established that excess zinc can be toxic to plants.
133,256,258,519,913,1095,1320,1546 Staker and Cummins1546 found zinc
toxicosis in onions, spinach and potatoes in some peat soils in New York.
The soils had accumulated several thousand ppm zinc (normal soils contain
97
10-300 ppm). However, few areas of natural zinc toxicity exist. Toxic
amounts of zinc
/usually occur in acid soils or in contaminated areas such as mine spoil
banks, industrial areas, waste disposal sites, or areas of mine seepage.
Zinc was toxic to corn and cowpeas when added to soil at rates
519
of 560-1,120 kg zinc/ha. A concentration of 50 ppm zinc in Sanilac
field beans can be poisonous; toxicosis at this concentration was produced
1027
mainly with acid soils. Zinc toxicosis has been reported in several
256
plants in greenhouse studies when excessive levels of zinc were added.
130 133
Boawn and Boawn and Rasmussen have investigated potential zinc
toxicosis in many plants, and their results are provided in Tables 5-6
124
-------
TABLE 5-6
Zinc Concentration in Leafy Vegetables Grown
at Normal and Excessive Rates
Zinc Fertilization a
Crop
Head Lettuce
(Lactuca sativa, var. capitata)
Leaf Lettuce
(Lactuca sativa. var. crispa)
Romaine Lettuce
(Lactuca sativa, var. longifolia)
Romaine Lettuce (Lactuca sativa)
Endive (Cichorium endivia)
Parsley
f~J (Petroselinum hortense)
Ol
Swiss Chard
(Beta vulgar is, var. cicla)
Spinach
(Spinacia oleracea)
Chinese Cabbage
(Brassica pekinensis)
Chinese Cabbage
(Brassica pekinensis)
Mustard (Brassica juncea)
Collard (Brassica oleracea,
var. acephala)
Cabbage (Brassica oleracea,
var. capitata)
Brussel Sprouts (Brassica
oleracea, var. gemmifera)
Variety
Imperial 847
Grand Rapids
Parr is Island
Parris Island
Green Curled
Extra Curled
Dwarf
Lucullus
Improved Thick
Leaf
Chihli
Chihli
Florida Broad-
leaf
Vates
Ear liana
Long Island
Imperial
Sample
Description
Market size
Market size
Whole plant
heading
Market size
Market size
Market size
Market size
Market size
Whole plant
heading
Market size
Market size
heads
plants
before
heads
plants
plants
plants
plants
before
heads
plants
Rosette of young
leaves
Market size
Market size
heads
heads
of
Zinc Treatment, kg/ha
0
38
38
32
48
32
58
80
139
54
46
32
33
22
50
11
45
46
40 .
50
38
50
72
119
48
42
32
34
20
47
55
64
64
56
62
73
86
153
148
68
60
36
38
23
56
112
ppm
94
125
78
76
142
107
325
175
84
71
43
42
28
50
224
144
157
108
100
247
188
615
240
89
112
58
63
34
62
448
165
239
146
117
308
296
704
344
112
248
131
104
54
73
896
248
269
179
122
343
438
862"
340-'
114
389
364
366
73
79
aData from Boawn.13°
the indicated rate of zinc fertilization, plants
showed normal color but were stunted.
-------
842
and 5.7. King and Morris reported zinc toxicosis in rye at soil applica-
tions of 600 kg zinc/ha in sewage sludge on sandy loam soil, but no toxicosis
841
was observed in Bermuda grass at the same zinc levels. Zinc levels in
the plants were in excess of 300 ppm at the highest rate of zinc application.
Zinc toxicosis was induced in pine trees on three coastal plain soils of
1673
Florida when 200-300 ppm zinc was added to the soil. Tissue concentra-
tions reached 300 ppm zinc at the highest application rates.
267
Conversely, Chesnin observed that field experiments in Nebraska
involving zinc applications up to 700 ppm zinc for corn on acid and alkaline
soils did not bring about toxicosis. However, corn is thought to be
moderately tolerant to excessive zinc levels in soil. In Michigan, a
single application of 140 kg zinc/ha did not induce zinc toxicosis to navy
1693
beans on a calcareous soil. Nor did additions of 387 kg zinc/ha result
in toxicosis in corn, cucumbers, or snap beans grown on a sandy soil in
1725
Wisconsin. Six annual additions of 11.1 kg zinc/ha on a sandy soil
979
in Virginia did not cause toxicosis in soybeans.
Tolerance. Certainly many plant and soil factors influence the
susceptibility of various species to zinc toxicosis. However, certain
species have a relatively high tolerance to zinc in soils. Antonovics
43
et al. have reviewed the zinc tolerance of various plant species growing
upon zinc-contaminated soils. Plant species that can tolerate zinc are:
Rumex acetosa, Festuca ovina, Agrostis stolonifera, Agrostis canina,
Viola lutea, Alsine verna, Silene vulgaris, Plantago lanceolata, Linum
catharticum. Campanula tenuis, Festuca rubra, Holcus lanatus, Anthoxanthum
43
odoratum, Thlaspi alpestre, and Armoria maritima.
126
-------
TABLE 5-7
10
Gram at Excessive Rates
Zinc added,
ppm
Field corn
(Zea mays) ,
Idahybrid 330
Sweet Corn,
Golden
Cross Bantam
cine in yield sine in
tops, ppm decrease Z tops, ppa
10 37
100 205
200 314
300 484
400 576
500 763
Z yield decrease for significant
•c .05 probability level:
Zinc added,
ppm
0
0
13
20
26
42
26
Wheat
(Tritlcum
vuleare) . Gaines
zinc in yield
tons, ppm decrease Z
10 51
100 185
200 345
300 522
400 682
500 909
Z yield decrease for significant
at. .05 probability level:
Zinc added';
ppa
Pea (Piaum
sativum) ,
Perfection
0
1
3
18
30
45
10
41
255
367
475
695
713
yield
decrease Z
0
8
12
32
55
48
29
of Zinc Fertilization"
Sorghum (Sonthum bicolor) ,
NR-125
sine in
tops, ppm
34
380
506
748
917
1,029
yield
decreaseZ
0
10
30
43
62
80
15
(Fhaseolus vulgar is)
Field beans,
Bin Bend
cine in
tops, ppm
24
66
101
151
213
257
yield
decrease Z
0
0
0
0
0
10
ns
Pea, Alaska
cine in yield
tops, ppm decreases
10 37 0
100 132 0
200 197 4
300 285 6
400 367 7
500 489 8
Z yield decrease for significant
at .05 probability level; ns
Zinc added,,
ppa
Potato
White Rose
sine in yield
tops , ppm decre
10 28
100 67
200 95
300 138
400 2"
500 3*6
Z yield decrease Cor significant
at .05 probability level:
sine in
tops , ppm
36
104
166
236
379
522
yield
decreaseZ
0
0
1
9
10
30
22
Sugar beet
(Beta vulgar is) .
HonoKerm Hybrid
aseZ
0
2
1
5
0
' 8
ns
cine in
tops, ppm
39
162
355
509
851
1,067
yield
decreaseZ
0
17
14
13
23
40
22
Snap beans,
Yaklma
sine in
tops, ppm
21
46
69
111
142
213
Lettuce (
sativa) .
zinc in
tops, ppm
34
96
152
250
390
665
yield
decreaseZ
0
11
8
8
14
12
ns
Lactuea
New xork
yield
decreaseZ
0
18
4
21
18
31
27
RS-626
cine in
tops, ppm
32
357
571
646
975
1,140
yield
decreaseZ
0
7
11
50
66
70
19
Alfalfa
(Hedicago
satlva). Vernal
zinc in
tops, ppm
27
71
97
142
232
345
Spinach (Sj
oleracea) ,
leaf
zinc in
tops, ppm
72
338
452
640
775
945
yield
decreaseZ
0
0
3
0
17
22
13
jlnacta
Thick
yield
decreaseZ
0
0
1
12
19
32
10
Barley
(Hordeum
vulgare) , Trail
zinc in
topa, ppm
70
220
530
910
1,237
2,112
Clover
(Trtfolium
pratense) .
zinc in
tops, ppm
28
81
109
161
202
252
yield
decreaseZ
0
10
16
42
59
76
15
Ladino
yield
decreaaeZ
0
0
2"
7
0
9
OS
Potato (Solanun
tuberosum) . Russet
Burbank
zinc in
tops, ppm
33
79
125
163
236
327
yield
decreaaeZ
0
0
0
0
0
0
ns
Tomato
(Lvcoperaicon esculentum) ,
Royal Ace
mine in
tops, ppm
51
150
257
316
381
514
yield
decreaseZ
0
0
5
8
18
26
24
"Data from Boawn and
-------
Plants that have been used as indicators of high levels of soil zinc
are listed below. Although many plants are sensitive to high soil zinc
levels, certain plants can provide vegetative cover on zinc-contaminated
soils because of their tolerance for high zinc levels in soils. In Europe,
Minuartia verna. Armeria vulgaris (and halleri, elongata, martima, etc.),
and Viola calaminaria announce the presence of zinc; in Australia, Tephrosia
sp. nov., Tephrosia affin. polyzyga, Polycarpaea synandra var. gracilis,
and Gomphrena canescens are such reliable markers that they are used as
such in prospecting for heavy metals. Philadelphus sp. is a similar
43
signal in the state of Washington.
The mechanisms of zinc tolerance in colonial bent grass (which is
readily tolerant to zinc) and creeping bent grass (which is not) have been
well examined.1237'1633'1634 Similar amounts of zinc entered the tolerant
and intolerant plants and similar amounts reached the tops, but the tolerant
plants appeared to deactivate zinc. Most of the cytoplasmic zinc in the
1237
tops of colonial bent grass existed as a stable anionic complex.
Considerable differences were noted among the physiologic responses of
the bent grass species. ' The binding of zinc in the cell wall
fraction was suggested as the mechanism of zinc tolerance in the colonial
bent grass. Species other than bent grass have developed zinc-tolerant
563
ecotypes, but they have not yet been studied in detail.
The relationships between cadmium and zinc in grasses are quite
744
important to human and animal nutrition. Huffman and Hodgson determined
the zinc:cadmium ratios in wheat and perennial grass samples from 19 states
128
-------
east of the Rocky Mountains. No zinc: cadmium ratios appeared to be determined
by regions of the country in which the plants grew. These data are summarized
in Table 5-8. High levels of zinc may suppress cadmium uptake by plants,
but at levels of zinc normally found in soils, suppression does not seem to
256,598,1632
occur. There is evidence that each element may enhance the uptake of the
other even when relatively low levels of cadmium and zinc are present in
256
the soil.
Zinc in Plant Metabolism
The specific functions of zinc in plant metabolism are not completely
defined, but much progress has been made in the last 30 years. Established
1296
functions of zinc in plant metabolism have been reviewed.
The best defined and most important role of zinc in plants is as
*
an enzyme cof actor. Carbonic anhydrase was the first zinc metalloenzyme
to be identified in plants and considerable research has been and is still
55,56,371,415,450,554a,1130,
conducted on zinc and carbonic anhydrase activity.
1312,14 8,1624,1803 ^^ metalloenzymes that require zinc have been
described . Zinc binds pyridine nucleotides to the protein
portion of enzymes and zinc atoms stabilize the structure of yeast alcohol
59 8a
dehydrogenase . It has been recognized as an essential component of a
variety of dehydr ogenases , proteinases, and peptidases, as well as the zinc
metalloenzyme carbonic anhydrase. In plants they include alcohol
dehydrogenase, glutamic dehydrogenase, L-lactic dehydrogenase, D-glyceraldehyde-3 -
phosphate-dehydrogenase, glutamic dehydrogenase, D-lactic dehydrogenase, D-lactic
cytochrome c^ reductase and aldolase.
* For more information on zinc and enzymatic activity, see Chapter 9.
129
-------
TABLE 5-8
Average Cadmium Concentrations and. ZincjjCadmium.
-^ — - * J
Ratios
State
Alabama
Colorado
Connecticut
Georgia
Illinois
Iowa
Kansas
Kentucky
Louisiana
Massachusetts
Mississippi
Nebraska
New York
North Carolina
Ohio
Oklahoma
South Carolina
Texas
Virginia
Summary
Found in Wheat
Crop
Wheat
Grass
Wheat
Grass
Wheat
Grass
Wheat
Grass
Wheat
Grass
Wheat
Grass
Wheat
Grass
Wheat
Grass
Wheat
Grass
Wheat
Grass
Wheat
Grass
Wheat
Grass
Wheat
Grass
Wheat
Grass
Wheat
Grass
Wheat
Grass
Wheat
Grass
Wheat
Grass
Wheat
Grass
of Data from S
and Perennial
Number of
Samples
-
8
9
2
-
7
1
23
1
8
-
7
7
6
1
2
-
1
-
7
-
2
5
3
-
6
4
4
-
8
2
1
-
4
2
21
1
-
amples Taken
. Grasses
Cadmium,
—
0.17
0.19
0.27
—
0.13
0.18
0.16
0.34
0.21
—
0.26
0.16
0.15
0.12
0.11
—
0.18
—
0.13
—
0.44
0.32
0.25
—
0.14
0.22
0.20
—
0.17
0.12
0.14
—
0.18
0.14
0.15
0.25
—
East of the
Zinc: Cadmium,
PPm wet weight
-
122
80
80
—
187
94
131
58
107
_
88
94
138
108
147
—
172
_
191
—
70
55
76
—
130
96
133
—
152
97
143
-
115
168
166
60
-
Mississippi
Wheat 8 0.22 83
Grass 79 0.17 139
Overall 87 0.17 136
Summary of Data from Samples Taken West of the Mississippi
Wheat 25 0.20 87
Grass 41 0.18 137
Overall 66 0.19 ' 118
Summary of All Data
Wheat 33 0.20
Grass 120 0.17
Overall 153 0.18
88
137
126
^
a Data from Huffman and Hodgson.
-------
Tomato plants grown in zinc-deficient media have provided correlations
between low zinc, carbonic anhydrase activity and protein level. Zinc
supplied to a deficient citrus plant increased RNA and protein synthesis but
838 1764
decreased ribonuclease activity in the leaves. Subsequently, White et al.
reported that RNA synthesis was a prerequisite for protein synthesis. Zinc
supplied to deficient Neurospora increased the activity of alcohol dehydrogenase
only in the presence of a nitrogen source; apparently protein synthesis must
occur to affect enzyme activity. The presence of a nitrogen source appears
1294
to be a general requirement for the recovery of zinc-sensitive functions.
When zinc is insufficient in growing organisms, metabolic lesion occurs.
1433,1710
First, RNA falls to form, followed by protein, total nitrogen and DNA lesions./
1294
In Euglena severely deficient in zinc, the absolute amount of RNA decreases.
837,838
RNA hydrolysis has been found to increase in citrus leaves lacking zinc.
Zinc in plants is known to be closely associated with proteins, '
798 1488
and it is bound to some enzymes. Sibly and Wood found that zinc
was not removed by dialysis against water from plant carbonic anhydrase.
Zinc is more concentrated in the protein fraction of the Russett Burbank
potato tuber rather than in the tuber as a whole, indicating an association
926
between zinc and protein.
Chelating agents remove or combine with zinc cofactors in enzymes,
so that enzymatic activity is lost. In yeast alcohol dehydrogenase, 1,
10-phenanthroline does not remove zinc from the enzyme but
forms a dissociable zinc-protein-chelate complex that inhibits enzyme
activity. This reaction is common to many other zinc metalloenzymes
1653,1658
dependent upon pyridine.
131
-------
Zinc is so firmly bound in carboxypeptidase that it is not removed by
prolonged dialysis against water; however, 1,10-phenanthroline will
remove the zinc. Carbonic anhydrase also binds zinc so firmly that its
712
zinc will not exchange with zinc-65 for over 32 days.
928
There are two types of interaction between zinc and proteins.
Zinc metalloenzymes are enzymes in which zinc atoms are specifically and
firmly incorporated into the protein, so that they can be considered as a
single physical entity in their native state, and homogeneous metalloenzymes
can be isolated and identified. In contrast, zinc metal-protein complexes
are formed with enzymes which may require zinc as one of several units for
activity. The enzymes are more weakly bonded with zinc and cannot be
928
isolated. Less zinc is bound by enzymes or whole plant extracts as the
798
pH of the medium decreases.
1296
Price et al. stated that if zinc were essential to cytoplasmic
ribosome stability, then it would be necessary to provide another function of zinc in
plant growth and development. Such a function was proposed when it was found
that cytoplasmic ribosomes of Euglena gracilis contained large amounts of
1289
zinc and that the ribosomes became unstable when the Euglena lacked zinc.
Early research showed that zinc and auxin contents are linked in
1491 1630
higher plants and Tsui concluded that zinc was essential for the
synthesis of tryptophan in tomato and indirectly
for its auxin synthesis. Later, Nason found that zinc was needed to
form tryptophan from indole and serine.
132
-------
Because zinc is primarily an enzyme cofactor, it is likely that zinc aids
1 / C O
auxin synthesis by activating enzymes. For example, growth was stimulated
1394
in zinc-deficient corn seedlings by adding tryptophan. Therefore, zinc
may be needed to synthesize tryptophan rather than in the direct formation
of auxins. The role of zinc in auxin formation is certainly not clear at
present. Research also has been reported which postulates that zinc may
804,805
be involved with starch metabolism in bean plants. Again, the
research is not yet sufficiently advanced to evaluate this newly proposed
function for zinc.
133
-------
CHAPTER 6
ZINC IN AQUATIC ANIMALS
Zinc is distributed throughout freshwater and marine aquatic environ-
ments and occurs in all organisms analyzed. Interest in the accumulation
and distribution of zinc in aquatic organisms has arisen from a concern
about heavy metals or radioisotopes in human food products of marine or
freshwater origin. However, in terrestrial organisms, interest always has
been sparked by the need to know nutritional requirements for domestic stock
and fowl as well as humans. Therefore, much of the information on zinc in
aquatic environments is found in the literature on radioecology and pollution
rather than in biochemistry and nutrition reviews. Literature about the eco-
logic cycling of zinc in aquatic environments and distribution of zinc in
62 1339 1695
aquatic organisms has been written by Bachman, Rice, Vinogradov,
105 1223 1797,1800
Bernhard and Zattera, Pentreath, and Wolfe.
CONCENTRATIONS OF ZINC IN AQUATIC ANIMALS
Marine Animals
Zinc is present in significant concentrations in all marine animals.
The range of concentration among species is relatively narrow, except for
.oysters, in which very high amounts may occur. Whereas the zina content of
most seafoods ranges from 3-30 ppm, oysters contain tOOfc2,QOO ppm.
The National Marine Fisheries Service has conducted extensive investiga-
tions into the content of zinc in numerous species of marine fish, mollusks
and crustaceans. These results are summarized in Table 6-1. The average
zinc content of all finfish was about 6.5 ppm, and means for individual
134
-------
TABLE 6-1
Zinc Content of Fish and Shellfish
from Coastal Waters of North America^
Species
FISH
Albacore
(Thunnus alalunga)
Albacore
(Thunnus alalunga)
Anchovy, Northern
(Engraulis mordax)
Bass, Black sea
(Centroprlstes striatus)
Bass, Striped
(Roccus saxatllls)
Bluefish
(Pomatomus saltatrlx)
Bluefish
(Pomatomus saltatrlx)
Bocacclo
(Sebastodes pauclsplnls)
Bocacclo
(Sebastodes pauclsplnls)
Bonito, Pacific
(Sarda chlliensls)
Butterfish
(Poronatus trlacanthus)
Catfish, Channel
(Ictalurus punctatus)
Catfish, Gaff -topsail
(Bagre marinas)
Catfish, Sea
(Arlus fells)
Cod, Pacific
(Gadus macrocephalus)
Area6
C
w
C
s
C
A
S
C
W
C
A
S
G
G
W
Samples
analyzed0
8
10
30
24
17
19
21
21
21
26
29
10
19
30
37
135
Mean
4.05
6.90
23.06
4.12
3.70
9.04
8.14
3.86
4.04
6.67
7.35
6.33
10.13
14.06
4.00
Zinc, ppm
Standard
Deviation
1.15
9.93
5.39
1.70
1.06
4.09
10.78
0.75
0.91
1.43
2.30
2.76
3.75
6.33
1.09
Low
2.00
2.64
15.55
1.43
2.14
2.94
3.06
2.21
2.14
2.06
3.87
4.42
4.44
4.94
2.50
Hifih
5.67
35.00
44.38
9.64
6.79
19.28
54.95
5.00
5.70
8.57
14.28
13.93
17.32
30.37
7.14
-------
TABLE 6-1
(Continued)
Species
Cod, Atlantic
(Gadus morrhua)
Croaker, Atlantic
(Micropogon undulatus)
Croaker, Atlantic
(Micropogon undulatus)
Dogfish, Smooth
(Mustelus canis)
Dogfish, Spiny
(Squalus acanthias)
Dolphin
(Coryphaena hippurus)
Dolphin
(Coryphaena hippurus)
Drum, Black
(Pogonias cromis)
Drum, Red
(Sciaenops ocellata)
Eel. American
(Anguilla rostrata)
Flounder, Gulf
(Paralichthys albigutta)
Flounder, Summer
(Paralichthys dentatus)
Flounder, Winter
(Pseudopleuronectes americanus)
Flounder, Yellowtail
Area*
A
G
S
A
W
H
S
G
G
A
G
S
A
A
Samples
analyzed0
46
10
10
11
44
21
21
23
32
10
20
32
22
29
Mean
3.71
3.94
8.14
4.37
3.67
3.46
5.60
4.81
4.08
17.14
3.34
4.83
4.68
4.73
Zinc, ppm
Standard
Deviation
1.74
0.98
11.00
0.93
1.54
1.04
1.27
1.44
0.94
4.02
0.90
1.12
2.23
2.00
Low
0.87
2.50
3.20
2.94
1.61
1.31
3.23
20.6
2.24
8.75
1.40
2.68
0.69
1.44
High
10.50
5.59
39.30
6.07
8.50
5.53
8.44
7.50
6.43
21.89
5.51
7.51
12.50
13.06
(Limanda ferruginea)
136
-------
TABLE 6-1
(Continued)
Species
Grouper, Red
(Epinephelus morio)
Grouper, Black
(Mycteroperca bonacl)
Grouper, Scamp
(Mycteroperca phenax)
Haddock
(Melanogrammus aeglefinus)
Hake, Pacific
(Merlucclus productus)
Hake, Red
(Urophysia chuss)
Hake, Silver
(Merluccius bilinear is)
Hake, White
(Urophysis tenuis)
Halibut, Pacific
(Hippoglossus stenolepis)
Herring, Pacific
(Clupea harengus pallasi)
Herring, Pacific
(Clupea harengus pallasi)
Jack, Crevalle
(Caranx hippos)
Jacksmelt (whole)
(Atherinopsis calif orniensis)
Lingcod
Area*
G
G
S
A
W
A
A
A
W
K
W
G
W
W
Samples
analyzed
10
18
14
32
41
19
17
26
26
9
14
36
31
49
Mean
3.93
4.85
5.18
3.99
5.08
2.97
3.47
3.01
3.67
14.32
5.38
6.73
12.60
4.74
Zinc, ppm
Standard
Deviation
0.86
1.50
5.94
3.15
6.89
1.15
0.64
0.91
1.47
3.92
1.78
8.66
4.12
1.36
Low
2.50
3.10
1.87
0.13
2.21
0.40
2.44
1.21
1.31
8.13
3.21
0.48
4.63
1.54
High
5.12
8.31
25.51
18.50
47.10
5.62
4.64
5.44
7.50
22.50
8.12
56.30
25.00
9.19
(Ophiodon elongatus)
137
-------
TABLE 6-1
(Continued)
Species
Mackerel, Atlantic
(Scomber scombrus)
Mackerel, King
( Scomberomorus cavalla)
Mackerel, King
( S comberomorus cavalla)
Mackerel , Chub
(Scomber j aponicus)
Mackerel , Spanish
( S comberomorus macula tus)
Marlin, Blue
(Makaira nigricans)
Marlin, White
(Makaira albida)
Menhaden, Atlantic (whole)
(Brevoortia tyrannus)
Menhaden, Atlantic (whole)
(Brevoortia tyrannus)
Menhaden , Gulf (whole)
(Brevoortia pa tr onus)
Menhaden, Gulf
(Brevoortia patronus)
Menhaden, Yellowfin (whole)
(Brevoortia smithi)
Mullet, Striped
(Mugil cephalus)
Mullet, Striped
Area*
A
G
S
C
S
S
S
A
S
G
G
G
G
S
Samples
analyzed
8
22
20
12
29
11
15
23
9
17
11
12
36
20
Mean
6.84
5.15
5.11
7.47
5.78
8.30
5.05
18.38
15.15
23.85
12.67
17.17
5.56
5.95
Zinc, ppm
Standard
Deviation
1.85
1.27
1.13
2.56
1.51
3.15
1.92
5.52
5.50
10.70
10.29
3.51
1.33
2.33
Low
4.50
3.31
2.25
4.25
3.82
4.00
2.56
7.75
9.25
11.88
5.75
13.13
2.80
3.45
HiRh
9.42
8.03
6.78
14.25
9.64
13.57
8.57
30.35
23.43
57.14
33.93
25.00
9.64
10.75
(Mugil cephalus)
138
-------
TABLE 6-1
(Continued)
Species
Perch, Ocean
(Sebastes marinua)
Perch, Silver
(Bairdiella chrysura)
Perch, White
(Morone americana)
Pollock
(Pollachius virens)
Pompano , Florida
(Trachinotus carolinus)
Pompano, Florida
(Trachinotus carolinus)
Rockfish, Canary
(Sebastodes pinniger)
Rockfish, Canary
(Sebastodes pinniger)
Rockfish, Yellowtail
(Sebastodes flavidus)
Sablefish
(Anoplopoma finbria)
Sablefish
(Anoplopoma fimbria)
Salmon , Chinook
(Oncorhynchus tshawytscha)
Salmon, Chinook
(Oncorhynchus tshawytscha)
Salmon, Chum
Area6
A
S
A
A
G
S
C
W
w
C
K
K
W
K
Samples
analyzed"
16
16
9
22
9
15
10
32
19
39
8
22
55
15
Mean
3.80
12.19
12.28
4.16
6.50
8.60
6.10
4.34
4-30
3.19
2.98
4.83
4.46
4.66
Zinc, ppm
Standard
Deviation
2.40
21.74
3.66
2.32
1.03
5.64
2.92
1.21
1.59
1.65
0.80
2.27
1.56
0.91
Low
1.82
4.82
6.62
1.12
5.54
1.43
3.37
2.50
2.29
0.00
1.87
2.63
1.38
2.88
High
12.10
93.60
17.90
11.20
8.81
26.64
10.71
8.13
8.57
9.06
4.50
13.75
8.93
6.34
(Oncorhynchus keta)
139
-------
TABLE 6-1
(Continued)
Species
Salmon , Coho
(Oncorhynchus kisutch)
Salmon , Coho
(Oncorhynchus kisutch)
Salmon, Pink
(Oncorhynchus gorbuscha)
Salmon , Sockeye
(Oncorhynchus nerka)
Salmon, Sockeye
(Oncorhynchus nerka)
Seatrout, Spotted
(Cynoscion nebulosus)
Seatrout , Spotted
(Cynoscion nebulosus)
Shark, Blacktlp
(Carcharhinus limbatus)
Snapper, Red
(Lut janus campechanus)
Snapper, Red
(Lut janus campechanus)
Snapper, Red (Ehu)
(Etelis marchi)
Snapper, Vermilion
(Rhomboplites aurorubens)
Sole , Dover
(Microstomus pacificus)
Sole, Dover
Area3
K
W
K
K
W
G
S
S
6
S
H
S
C
W
Samples
analyzed0
18
25
15
22
21
28
29
10
8
16
9
25
28
38
Mean
5.52
3.03
5.45
6.24
7.56
4.49
4.34
4.01
3.87
4.90
4.46
3.30
3.92
3.99
Zinc, ppm
Standard
Deviation
1.65
0.92
1.44
2.93
10.10
1.43
1.19
1.75
1.38
2.11
0.76
0.93
1.34
1.67
Low
3.22
1.30
3.57
2.44
2.81
2.34
2.85
1.25
2.03
2.50
3.28
2.06
2.14
1.18
High
10.00
4.75
8.39
16.07
49.64
8.93
7.50
8.10
6.43
9.64
5.71
6.43
8.00
9.29
(Microstomus pacificus)
140
-------
TABLE 6-1
(Continued)
Species
Sole, English
(Parophrys vetulus)
Sole, English
(Parophrys vetulus)
Sole, Petrale
(Eopsetta jordani)
Sole, Petrale
(Eopsetta jordani)
Sole, Rex
(Glyptocephalus zachirus)
Sole, Rex
(Glyptocephalus zachirus)
Spot
(Leiostomus xanthurus)
Sturgeon, Green
(Acipenser medirostris)
Tuna, Bigeye
(Thunnus obesus)
Tuna, Bluefin
(Thunnus thynnus)
Tuna, Skipjack
(Euthynnus pelamis)
Tuna, Skipjack
(Euthynnus pelamis)
Tuna, Yellowfin
(Thunnus albacares)
Tuna, Yellowfin
(Thunnus albacares)
Weakfish
Area6
C
w
C
w
C
w
s
w
H
C
C
H
C
H
A
Samples
analyzed0
24
40
18
36
10
37
19
10
16
10
13
16
22
15
18
Mean
4.59
4.42
3.60
4.32
3.89
3.63
4.99
4.33
3.29
6.92
10.39
4.82
6.41
3.44
4.67
Zinc, ppm
Standard
1.53
1.06
0.87
0.96
2.36
1.27
0.91
1.36
1.07
2.87
3.77
1.72
2.21
1.34
1.96
Low
1.44
2.38
2.13
2.86
1.44
1.79
3.21
2.86
0.50
3.13
4.31
2.13
2.40
2.31
2.19
High
9.19
6.22
5.88
6.61
9.82
7.00
6.43
7.06
4.69
14.28
15.55
7.31
13.19
6.78
10.36
(Cynoscion regalis)
141
-------
TABLE 6-1
(Continued)
Species
Weakfish
(Cynoscion regalis)
MOLLUSKS
Abalone, Green
(Haliotis fulgens)
Abalone, Red
(Haliotis rufescens)
Clam, Butter
(Saxidomus giganteus)
Clam, Hard
(Mercenaria mercenaria)
Clam, Hard
(Mercenaria mercenaria)
Clam, Razor
(Siliqua patula)
Clam, Razor
(Siliqua patula)
Oyster, Eastern
(Crassostrea virginica)
Oyster, Eastern
(Crassostrea virginica)
Oyster, Eastern
(Crassostrea virginica)
Oyster, Pacific
(Crassostrea gigas)
Oyster, Pacific
(Crassostrea gigas)
Scallop, Calico
Area
S
C
C
W
A
S
K
W
A
G
S
C
W
S
Samples
analyzed
10
8
10
27
37
9
9
26
23
36
31
19
37
20
Mean
5.23
18.11
8.11
8.04
22.53
9.37
10.76
11.88
271.08
156.87
250.29
100.90
228.72
7.09
Zinc, ppm
Standard
Deviation
1.16
4.86
2.56
5.71
14.44
1.80
1.65
9.39
176.32
129.33
68.81
34.00
126.64
5.82
Low
3.75
10.00
4.14
0.78
7.00
6.56
9.37
0.85
38.13
21.28
9.81
48.12
31.24
0.71
High
7.78
26.07
13.93
15.72
58.93
13.37
14.32
26.92
1046.88
455.00
682.14
175.00
912.50
17.14
(Argopecten gibbus)
142
-------
TABLE 6-1
(Continued)
Species
Squid, Pacific
(Loligo opalescens)
Squid, Short-finned
(Illex illecebrosus)
CRUSTACEANS
Crab, Blue
(Callinectes sapidus)
Crab, Blue
(Callinectes sapidus)
Crab, Dungeness
(Cancer magister)
Crab, King
(Paralithodes camtschaticus)
Lobster, American
(Homarus americanus)
Lobster, Spiny
(Panulirus argus)
Shrimp, Brown
(Penaeus aztecus)
Shrimp, Pink
(Penaeus duorarum)
Shrimp, White
(Penaeus setiferus)
Shrimp, White
(Penaeus setiferus)
Area6
C
A
6
S
W
K
A
S
G
6
G
S
Samples Q
analyzed
21
15
15
11
28
10
16
18
27
15
19
29
Mean
6.98
16.41
34.99
34.76
47.98
57.24
22.54
21.99
12.63
9.46
6.77
8.07
Zinc, ppm
Standard
Deviation
5.84
4.91
32.19
8.95
12.55
38.09
6.76
16.43
4.85
1.34
5.93
4.91
Low
1.33
8.06
12.50
23.00
24.38
16.31
8.81
4.69
4.93
7.36
0.83
0.50
High
18.13
35.71
115.62
50.00
68.57
118.70
35.65
83.75
31.07
11.50
20.62
15.00
from 1975 microconstituent resource survey, National Marine Fisheries Service,
College Park, Md. (unpublished).
Areas: A = Atlantic Coast, Newfoundland to Chesapeake Bay
S = Atlantic Coast, Cape Hatteras to Florida
G = Gulf of Mexico
C - Pacific Coast, California
W = Pacific Coast, British Columbia to Oregon
K = Pacific Coast, Alaska
H = Hawaiian coastal waters
A11 samples are raw edible flesh from individual fish, except when noted.
143
-------
species ranged from 3-24 ppm. As mentioned, oysters are exceptionally high
in zinc among mollusks; their mean for values shown in Table 6-1 is 202 ppm,
as compared to a mean value of 12.5 ppm for clams. Crustaceans contained
slightly more zinc than finfish; the mean value for crustaceans was 12.5 ppm,
and means for individual species ranged from 7-57 ppm.
Such results are consistent with measurements of the concentrations of
1626
zinc in shellfish from coastal areas of Scotland, and amounts of zinc
found in finfish sampled from coastal and deepwater areas around England and
1266
Wales. The evidence indicates that fish regulate the concentrations of
zinc in muscle tissues and that zinc levels in adult finfish do not vary
greatly with age.
Results are also similar for amounts of zinc reported for processed fish
and crustaceans. For example, Gormican found the following zinc concentrations
in processed samples: canned and salted crab, 36 ppm; uncooked frozen
haddock, 3 ppm; canned sockeye salmon, 11 ppm; canned and salted shrimp, 19 ppm;
549
uncooked, frozen sole, 3 ppm; and water-packed canned tuna, 4 ppm. However,
Takino reported that the average zinc content in the muscle of 34 species of
1591
fish was only about 1 ppm, with a range of zero to 2 ppm. These values
are about tenfold lower than the others reported in previous literature on fin-
fish. The low values and very narrow ranges makes the validity of Takino's
research suspect.
Mollusks. The subject of most investigations on zinc in marine animals
has been mollusks. Mollusks are able to concentrate certain trace metals up
1301
to many thousands of times that level found in the environment. Fringle et al.
have published extensive data on the uptake of zinc and other trace elements
by several estuarine mollusks. They collected samples of shellfish at 100
144
-------
stations from Atlantic and Pacific waters and reported the following zinc
contents: Eastern oysters accumulated about 1,428 ppm zinc on a wet weight
basis, with a range between 180-4,120 ppm; amounts reported for the Pacific
oyster ranged between 86-344 ppm wet weight. The mean zinc content of soft-
shell clams was 17 ppm wet weight, and the range was 9-28 ppm. The northern
quahog had a mean of 21 ppm wet weight and a range of 12-40 ppm.
Zinc levels in shellfish along the Atlantic Coast varied from about
10-40 ppm in the case of hard- and soft-shell clams to a range between
1301
180-4,100 ppm for the Eastern oyster. But zinc levels in the Pacific
oyster only reached 90-350 ppm. The high levels of zinc found in Eastern
oysters suggests a possible genetic difference between Atlantic and Pacific
oysters for the physiologic role of zinc.
Studies on the uptake and content of zinc in soft shell
clams showed that clams reached concentrations of 27 ppm in 50 days when
1487
exposed to 0.2 ppm zinc at 20 C. This uptake represented a 17 ppm increase
in zinc over a period of 50 days, or 0.35 ppm zinc/kg/day. It was also
found that the soft-shell clam apparently concentrates copper preferentially
1487
over zinc under identical experimental conditions.
The significance of chemically polluted estuarine waters for shellfish
and, by implication, public health, prompted Shuster and Pringle to
obtain comprehensive data on the levels of certain chemicals in shellfish
and study the uptake of trace elements by oysters. Extensive data on zinc
in the Eastern oyster (Crassostrea virginica) were compiled; they are
summarized in Tables 6-2 and 6-3.
145
-------
TABLE 6-2
Zinc Content in American Oysters from Atlantic Coast Waters
Source of Data
Grassestrea.virginica
Zinc Content, ppm wet weight
Range
Mean
1301
Pringle et al. (Maine
through North Carolina, 1965-1967)
1009a
McFarren et al.
(New Hampshire through North
Carolina, 1960)
522
Galtsoff
(Long Island Sound, 1933-1935)
275
Chipman et al.
CConnecticut through Georgia}
204-4,120
310-4,000
710-2,760
740-1,332
1,404
1,641
1,468
1,018
1487
Shuster and Pringle also studied oysters exposed to 0.1 and 0.2
ppm zinc and produced the data set forth in Table 6-3. Zinc accumulation
increased with time over the 20-wk period, and oysters exposed to 0.2 ppm
exhibited somewhat higher concentrations than those exposed
to 0.1 ppm zinc. The highest levels observed in oysters exposed to 0.2 ppm zinc,
however, were comparable to the naturally high concentrations noted in Table 6-3.
146
-------
TABLE 6-3
Zinc Accumulated by the Eastern Oyster
in Two 20-Week Periods'2
Crassostrea -vlrgioiea
mean levels zinc, ppm wet weight
fc u.i ppm exp(
We
. 0.1 ppm exposure 0.2 ppm exposure
iCIfc
1967 1968 1967 1968
1.036 1,708 1,036 1,708
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
1,456
1,561
1,538
1,831
1,956
1,736
1,666
1,732
1,911
1,570
2,035
2,189
2,059
2,139
2,212
2,118
2,382
1,970
2,206
2,708
2,065
2,186
1,936
2,095
2,293
1,996
1,613
2,229
2,135
1,877
1,918
2,251
2,366
2,229
2,859
2,475
2,224
3,314
2,340
2,560
1,381
1,470
1,519
1,761
2,234
1,811
2,055
1,869
2,037
1,767
2,269
2,660
2,733
2,445
2,740
2,667
3,033
3,528
2,976
3,813
2,265
1,856
2,030
1,474
2,267
2,200
2,451
2,307
2,343
2,531
2,413
2,642
2,222
3,051
2,912
3,233
2,869
3,159
3,743
3,185
Data from Shuster and Pringle.
The mean levels of zinc in the oyster tissue are shown for each weekly
sampling interval at two different zinc levels in experimental seawater
systems. The experiment, first performed in 1967, was repeated in 1968;
trends were similar, although the initial zinc content of the oysters
was higher in 1968. The sample was about 200 oysters harvested from
Narragansett Bay, Rhode Island.
147
-------
Oysters lose zinc rapidly if they are transplanted into water with low
768
zinc content. A linear decrease from about 1,000 ppm to about 100 ppm over
a 4-mo period was observed. Copper in oysters did not decrease until at least
2 wks after transplanting, whereas zinc started to disappear immediately after
the change.
745
Huggett et al. established a high positive correlation between levels
of zinc and levels of copper and cadmium in oysters from the Chesapeake Bay.
Based on average values for these elements, they showed that a concentration
gradient existed in oysters from all river systems emptying into the bay,
and that in the oysters, each metal increased in concentration with proximity
to fresh water. The concentration of heavy metals by oysters was shown to
follow a predictable pattern: therefore, the oyster is usable as an index
for measuring and identifying unnatural amounts of metal taken up.
Although the high content of zinc in oysters has been well documented,
little information is available on the biologic availability of zinc in
1478
oysters to consumers of seafood. Shah et al. reported that when rats
were fed oyster-supplemented diets containing 60 pg zinc/g diet, the apparent
absorption of zinc from oysters was 16.3%, an amount not significantly
different from the corresponding value of 18.5% for zinc carbonate
ingested by rats. Dietary levels of zinc up to 2,000 ppm from either
source had no significant effect on body weight, hemoglobin, hematocrit and
liver cytochrome oxidase activity after 8 wks. The concentration of zinc
in various tissues was not affected by the source of zinc; however, the
dietary level markedly affected the concentration of zinc in all
tissues examined
148
-------
except muscle and hair. A homeostatic control of the intestinal absorption
of zinc was demonstrated when the fraction of dietary zinc deposited in rat
femurs was found to decrease markedly with increasing dietary concentrations.
The homeostatic mechanism that regulated absorption of zinc appeared to be
overcome at a dietary level between 275-1,550 yg/g because the skeletal load
of zinc increased at a dietary level of 1,550 ug/g but not at 275 ^ig/g or less.1478
Crustaceans. The zinc content of crustaceans has been studied less than that
of moHusks. The zinc content of several whole decapod crustaceans is rather
186
consistent and ranged from 20-50 yg/g (Table 6-4). The overall similarity
among the values from different animals suggested that the concentrations of
zinc and copper are probably regulated in all decapods. In Homarus, Carcinus,
Cancer and Maja, zinc is found mainly in the blood plasma, rather than in
189
the blood cells, where most of it is bound to proteins in the plasma. The
concentration of zinc in the blood is regulated at different values in different
species of crustaceans, as listed in Table 6-5. Variations between individuals
of the same species appear to be related to the amount of protein in the blood.
•
As Table 6-5 shows, wide differences exist among samples of muscle from
different species. Evidence indicates that t' s sine content of a muscle may
be related to the speed with which it can contract. For example, the muscles
that contract quickly in Homarus contain about 15 Vg/g zinc, whereas muscles
/ 191
that contract slowly contain about 100 pg/g.
An indication of the permeability of slow and fast muscles in crustaceans
was obtained by measuring concentrations of zinc-65 in lobsters kept in sea
186
water to which the isotope was added. The concentrations of zinc-65
(expressed as muscle:blood ratios) in slow muscle were about twice those of
149
-------
TABT.E 6-4
Mean Concentrations of Zinc and Copper in Whole
Decapod Crustaceans'2
Species
Shrimp and Prawns
Palaemon serratus
Palaemon squilla
Palaemonetes variant
Crangon vulgaris
Lobsters and Crayfish
Homarus vulgaris
Austropotamobius pallipes
Galathea squamifera
Porcellana platycheles
Number
of
Animals
6
3
3
6
1
2
7
3
Concentration, Ug/g
Solid
Content, %
27.0
29.7
24.9
28.6
34.9
28.6
32.7
44.5
wet weight
Zinc
21
30
20
34
23
24
18
54
tissue
Copper
30
31
32
32
33°
17
29
27
Hermit Crab
Eupagurus bernhardus
30.2
282
25
Crabs
Corystes cassivelaunus 1
Atelecyclus septemdentatus 3
Cancer pagurus 3
Portunus puber 3
Portunus depurator 3
Carcinus maenas 2
Xantho incisus 1
Pilumnus hirtellus^ 7
Maja squinado 1
36.8
39.6
33.5
33.9
31.6
28.6
34.6
36.6
33.8
39
32
27
27
21
22
26
49
21
22
10
20
21
18
22
20
28
25
a Data from Bryan.186
" Females with extenu
° Mean of 10 animals.
eggs,
150
-------
fast muscle, suggesting that slow muscle is more permeable to zinc. The nature
of the differences in concentrations of zinc between the two types of muscle
is not known. This feature, however, suggests that lobster, and presumably
other decapod crustaceans, would be suitable organisms for determining how
zinc functions in the muscles of crustaceans.
Concentrations of zinc in the hepatopancreas vary from species to species,
but usually lie within the range of 30-90 yg/g (see Table 6-5). In Homarus,
dietary zinc is rapidly absorbed and the concentration increases in the blood
and hepatopancreas. It is thought that the zinc stored in the hepatopancreas
of Homarus is not removed in the feces. Rather, it is lost across the body
193
surface or gradually lost into the blood and then excreted in the urine.
The amount of zinc which can be absorbed directly from seawater across
the body surface of decapod crustaceans increases if the concentration in the
water is raised. Therefore, species living in fairly clean water will absorb
less zinc from the water than species living in a polluted water where the
concentration is higher. However, variation in the dietary intake is probably
186
more important.
Freshwater Animals
Data on the content of zinc in freshwater animals are limited, but
it seems that the levels in freshwater fish are not markedly different from
amounts found in marine animals.
The concentration of zinc and 12 other elements was measured in a limited
study of dressed fish from nonindustrialized and heavily industrialized fresh-
1648
water areas around the Great Lakes. All samples were composite, consisting
of at least 2.25 kg or 3 fish. No major differences were detected between
151
-------
TABLE 6-5
Mean Concentrations of Zinc in Body Fluids and Tissues of Decapod Crustaceans—
Ul
10
Zinc, jig/g fresh weight
., b
Species—
Palaemon serratus
Palaemonetes varians
Crangon vulgaris
Falinurus vulgaris
Homarus vulgaris
Austropotamobius pallipea pallipes
Galathea squamifera
Eupagurus bernhardus
Corystes cassivelaunus
Atelecyclus septemdentatus
Cancer pagurus
Portunus puber
Portunus depurator
Carcinus maenas
Pilumnus hirtellus
Haja squinado
Number
of
Animals
6
6
6
2
4
6
5
3
5
3
5
5
3
11
3
3
Blood
38
87
23
3.1
7.4
0.9
0.25
11.6
11.0
9.5
49
7
1.8
36
12.3
2.4
Leg
Muscles
66
60
36
52
61
64
28
15
44
63
Abdominal
Muscles
10
14
14
20
15
12
10
24
Stomach
Fluid
48
0.7
41
47
26
12
6
15
15
13
18
92
31
Hepato-
pancreas
64
65
78
97
34
109
49
69
50
88
45
42
24
56
169
71
Urine
MHM^MM^^B
0.6
2.2
0.02
1.4
0.6
0.2
0.3
0.4
0.3
-------
TABLE 6-5 (continued)
Zinc, jig/g fresh Weight
Species—
Palaemon serratus
Palaemonetes varians
Crangon vulgaris
Palinurus vulgaris
Homarus vulgaris
Austropotamobius pallipes pallipes
Galathea squamifera
Eupagurus bernhardus
Cory s tea cassivelaunus
Atelecyclus sept end entatus
Cancer pagurus
Portunus puber
Portunus depurator
Carcinus maenas
Pilumnus hirtellus
Maja squinado
Soata from Bryan.186 Samples
Number
of
Animals
6
6
6
2
4
6
5
3
5
3
5
5
3
11
3
3
are from
Excretory
Organs
20
20
7
23
16
29
29
10
8
19
15
English waters.
Gills
20
15
8
27
69
45
27
42
23
25
26
60
10
Shell
16
5
8
9
28*
5
7
3
13
3
3
17
5
Ol« *l 1
Vas
Deferens
13
14
27
27
30
16
20
23
23
16
. not heavily
External
Ovary Eggs
24
82 107
50
26
56
87
45
calcified.
-------
the zinc levels of fish from nonindustralized and industralized areas. Lake
whitefish and northern pike from the nonindustralized Moose Lake area had 14
and 19 ppm zinc, respectively. Their counterparts in parts of Lake Ontario
and Lake St. Pierre affected by industry had incorporated 12 ppm and 19 ppm
zinc. Northern pike, rainbow smelt, and yellow perch around industralized
1648
areas of Lake Erie had absorbed 10, 20, and 12 ppm zinc, respectively.
Lucas et al. also investigated the concentration of zinc and other
954
trace elements in fish from the Great Lakes. They studied the content
of zinc and 14 other trace elements in 40 liver samples from 10 species of
fish from Lakes Michigan, Superior, and Erie. The levels of zinc varied
littie.between species and the range was 11-48 ppm. This range is somewhat
higher than the range (3-24 ppm) reported in Table 6-1 for mean concentrations
of zinc in marine samples.
154
-------
Concentration of Radioactive Zinc
Levels of zinc-65 have been studied in marine organisms mainly for
1215
their relation to discharges from nuclear reactors. Pearcy and Osterberg
investigated the major y-emitting radioisotopes found in the livers of
albacore (Thunnus alalunga) along the west coast of North America from 1962-1965.
The zinc-65 content of albacore livers from southern California and Baja California.
ranged from 10-100 pCi/g, which was about 10% of the levels found in samples
off Oregon and Washington.
The highest levels of zinc-65 were found in samples taken off northern
Oregon and Washington, and those levels increased markedly during the summer.
The data suggest that albacore rapidly accumulate zinc-65 after they migrate
into Oregon waters. It was concluded that radioactivity from the Columbia
River significantly affects the zinc-65 content of migratory albacore tuna.
Zinc-65 content and specific activity in albacore tended to be higher in
fish taken nearer the mouth of the Columbia River and in those taken in the
1215
summer and fall months.
The levels of zinc-65 in benthic invertebrates of the Oregon coast
231
have been reviewed by Carey. Zinc-65 and other artificial radioisotopes
in the marine environment off the northwest coast of the United States come
from activation of naturally occurring stable elements and from corrosion of
reactor parts within the Atomic Energy Commission (AEC) reactor at Hanford,
Washington. Marine benthic fauna concentrate zinc-65 and the concentration
232
changes markedly with depth. Rapid decreases generally occur within the
first 400 m of depth.
155
-------
1290
Preston reported that the rate of discharge of zinc-65 by a nuclear
power station in Essex, England, is based on the amount of the metal that
accumulates in oysters in the surrounding area. The rate of discharge of
zinc-65 is restricted specifically because of its high concentration in oyster
flesh. Assuming maximum human consumption of oyster flesh of 75 g/day over
the whole year, the calculated permissible level for oyster flesh was calculated
to be about 2,900 pCi/g. Because it had been shown that the concentration
factors (concentration of zinc-65 accumulated in organism tissue) for zinc
concentration of zinc-65 in seawater 275
tended to increase as the level of zinc in the seawater decreased, Preston
was able to suggest that the accumulation of zinc-65 in oysters could be
1290
limited by adding inactive zinc to seawater.
Studies on the stable zinc concentration in oysters before opening
the Essex power plant reported a mean of 367 ppm, with a range of 308-419 ppm,
whereas when the power plant was operating the mean was 834 ppm, with a range
of 388-1,230 ppm. These data indicated that zinc levels had risen, perhaps
caused by the zinc in the pipework of the power station's condenser cooling
water system.
156
-------
ZINC METABOLISM IN AQUATIC ANIMALS
Mo Husks
Oysters. The accumulation of zinc by various members of the family Ostreidae
received attention over half a century ago, 36»70^ ^ut that interest was
generated primarily by the undesirable phenomenon of "greening" in commercial
oysters, the outcome of excessive tissue concentrations of metallic elements,
521 522
particularly copper. ' More recently, the accumulation by oysters of zinc-65
from nuclear fallout475'1097'1797 or reactor effluents1290'1397'1477'1738
renewed interest in studying zinc metabolism in oysters and other organisms.
It has been found that oysters may concentrate zinc to levels greater than
275 1290 1797
100,000 times the ambient concentrations in seawater, ' ' and an
inverse relationship was demonstrated between this concentration factor and
the zinc concentration in seawater, suggesting a limiting or regulatory
275,1290
process. This mechanism may exist because of the limited number of
1223
zinc-binding sites on structural proteins. Curiously, replotting the
data reported reveals a strong linear log-log correlation between "equilibrium
zinc concentrations" in oysters and seawater, a correlation that might be
275,1290
expected from a simple adsorption mechanism.
Zinc is not strongly localized in particular organs or tissues of
312 768 1797
oysters; ' ' nor are there great differences in the molar ratios of
312
calcium, magnesium, sodium or potassium compared to zinc in any tissues.
External tissues such as mantle, gills, and labial palps tend to contain
more zinc than do internal tissues; adductor muscle tissue usually contains
157
-------
the least. In all tissues, zinc commonly is found with nuclear and cell debris
and with cell sap or cytoplasm, rather than being localized in either microsomal
312,1361,1800
or mitochondrial fractions. However, in none of these studies
was the relative concentration of zinc determined in any of the subcellular
fractions.
Nearly all soluble zinc in supernatant preparations from homogenized
oyster tissues is bound to proteins of high molecular weight, as experiments
1361,1800
with gel-diffusion chromatography have shown. Dialysis of whole
312,1361 1800
oyster tissue homogenates or soluble extracts, however, removes
about 95% of the zinc. Therefore, the zinc may be very weakly bound and
readily dissociated from the protein moieties. Based on the observation that
removing 95% of the zinc present had no effect on the activity of alkaline
1800
phosphatase, Wolfe concluded that most of the zinc in oysters was
superfluous to the biochemical or nutritional requirements of the organism.
Similarly, calculations of the zinc requirements of the oyster based on
enzyme-specific activities and zinc stoichiometry recorded for zinc enzymes
purified from other biologic systems amount to a small fraction (about 0.1%)
312 1224
of the total zinc present in oysters. ' Of the enzymes demonstrated
to be zinc metalloenzymes in other systems, only four (carbonic anhydrase,
alkaline phosphatase, carboxypeptidase A, and malic dehydrogenase) have
312 1800
been detected in Ostrea edulis or Crassostrea virginica. Alpha-
D-Mannosidase, a zinc metalloenzyme previously described for a gastropod
1520 312
mollusk (the limpet), was also abundant in Ostrea edulis. The zinc
metalloenzymes alcohol dehydrogenase, lactic dehydrogenase, glutamic
dehydrogenase, and carboxypeptidase B were not detected in whole tissue
homogenates or subcellular fractions from either species of oyster.
158
-------
Two hypotheses—both related to the metabolism of calcium—have been
offered to explain the high zinc concentrations found in oysters. The first
postulates that zinc may be assimilated from the environment coincidentally
with calcium by a relatively nonspecific ion transport mechanism to satisfy the
organism's large calcium requirements for shell deposition.1800 Moreover, at
the mantle and shell interface, effective ionic discrimination occurs, such
that calcium is deposited in the shell and zinc remains loosely bound to
available sites in the soft tissues. According to the second hypothesis, zinc
would have to be accumulated selectively and actively to overcome the com-
petitive action of calcium and maintain the vital functions dependent upon zinc,
especially the activities of carbonic anhydrase and alkaline phosphatase,
312
which are necessary for shell formation.
This possible zinc-calcium interaction also may be studied by considering
research on the dependency of relative ion composition of oyster tissues on
745 746
salinity and relative turnover rates of zinc and calcium. ' Because
calcium content of seawater is directly correlated with salinity and zinc
content is not, significant differences in zinc concentration or turnover
as a function of salinity would be expected for either of the mechanisms
proposed above.
To satisfy the first hypothesis, the zinc concentration would have
to increase at reduced salinities, because the calcium-assimilative transport
system presumably would have to work harder to maintain sufficient calcium
reserves for shell deposition. Conversely, if the second hypothesis is
correct, reduced zinc turnover and concentration in oysters at lower salinities
would be expected because of the decreased competition from environmental
159
-------
calcium. To date, only one experimental study has been conducted in which
the effects of salinity were separated from the influences of other significant
environmental variables. Duke et al. studied zinc-65 accumulation by
Crassostrea virginica and other estuarine organisms in a multifactorial
experiment in which salinity, stable zinc, pH and temperature were each
tested at two different levels for accumulation over 15 days.
The effect of salinity, tested at 25 and 30 ppt, was significantly
inverse: that is, zinc-65 accumulation was about 10% lower at the higher
salinity. An inverse salinity effect was also noted on the contents of
1383
manganese and magnesium plus strontium in shells, implying that the
oyster accepts less preferred ions for shell construction when subjected
to lower environmental calcium concentrations.
Copper and zinc were analyzed in oysters (Crassostrea virginica)
from various stations in the Newport River in North Carolina, and the
Rappahannock River in Virginia. Higher concentrations of copper and zinc were
found in animals living in fresher waters as had been shown previously for oysters
746
in the James, York, and Rappahannock estuaries in Virginia. For example,
in the Newport River estuary, oysters living at about 12 ppt salinity contained
a mean of 320 ppm zinc wet weight, whereas oysters at 32 ppt contained about
130 ppm zinc. The concentrations of zinc in unfiltered water samples from
339
this same estuary were constant, irrespective of salinity, implying that
the concentration gradient found in the oysters does not result from a
similar gradient in the water.
160
-------
The biologic turnover of zinc by oysters and other aquatic organisms
is summarized in Table 6-6. The most reliable data on zinc turnover in
oysters are from Seymour's studies conducted in the natural environment
for two years.1476 These data, summarized in Table 6-6, were recently analyzed
o / *7
again by Cutshall, who showed that both the accumulation and loss data
were described by simple first-order exponentials with similar rate constants.
The effective half-life for zinc-65 turnover was 135 days, and .the corresponding
biologic half-life was 300 days. Thus, once accumulated by oysters, zinc
is retained very effectively.
161
-------
TABLE 6-6
Zinc Turnover in Aquatic Organisms
Species
Biologic Half-Life
Experimental Conditions
Mo Husks
Mytilus edulisa
Mytilus californianus
Crassostrea gigas°
Anodonta nuttalliana
Littorina irrorat*^
Cruatacea
Homarua vulgar is"
Mala squinado*
AustropotamobiusJ
pallipes pallipes
Euphausla pacifica?
Fundulus heteroclitus"
k
Pleuronectes platessa
Micropogon undulatus
3.3-3.9 days
76-3.5 days
300 days
650 days
40 days Q 15 C
25 days @ 25 C
23 days @ 30 C
60-270 days
38 days
30-38 days
140 days
140 days @ 3 C
100 days @ 7 C
90 days @ 12 C
75 days @ 10 C
58 days @ 20 C
35 days @ 30 C
295-313 days
6.5 days
Up to 20-day zinc-65 uptake; 70-day loss in
laboratory; animals unfed
Columbia River zinc-65; life-long accumulation,
one year of loss in natural environment
Columbia.River zinc-65; life-long accumulation;
2 years of loss in natural environment
' Single injection of zinc-65; loss to synthetic
running pondwater
Unspecified, but turnovers correlated positively
with zinc concentration of seawater
Injection of zinc-65; loss into seawater contain-
ing 100 ug/1 zinc; periodic feeding
Single feeding of zinc-65; loss into unlabeled
freshwater; stable zinc in food increases turnover
15-day feeding period with «inc-65 labeled Artemia
nauplii; 5-mo Loss period
6-10 day zinc-65; accumulation from seawater; 29-
day loss period; data obtained with and without
sediment were combined
267-day zinc-65; accumulation from seawater; 91-
day loss period
Single intraperitoneal injection of zinc-65; loss
to clean seawater
aDerived from van Weers.1674
Derived from Young and Folson.1814
cDerived from Seymour.1476
'Tterived from Bryan e_t al.^^
^Derived from Bryan; 186 see also Table 6-5.
•'Derived from Bryan; *'2 see also Table 6-5.
^Derived from Fowler et al.502
^Derived from Cross e£ al.
^Derived from Mishuma and Odia.1069
^Derived from Shulman et. al.1*86
^Derived from Pentreath. 1222
^Derived from Baptist et^ al.
'"Derived from Harrison.
637
162
-------
Marine mussela and scallops. Mytilus has been used extensively as an
indicator of radioactive contamination because of its local abundance and worldwide
distribution; consequently, considerable attention has been focused on zinc
metabolism in this organism. Values for zinc turnover have been obtained for
, ,, . _, T 1222,1477,1674 899,1477,1814
Mytilus edulis L.. Mytilus californianus. and
826
Mytilus galloprovincialis. and they are listed for the first two species
826
in Table 6-6. Keckes et al. demonstrated conclusively that the experimental
determination of short-term turnover rates using zinc-65 and resolution of
turnover into differing rate constants depended heavily upon the duration of
exposure of the organism to zinc-65. From these observations, it is evident
that many mechanisms interact to produce zinc turnover, and that long exposure
periods are necessary for valid measures of elemental turnover in radioisotopic
347
loss experiments. This relationship is discussed theoretically by Cutshall;
it is evident specifically in comparisons of the data from experimental
826,1674
exposures of various short durations, and in data from lifelong exposure
to zinc-65, which suggested only a single long-lived component when zinc-65
was lost by Mytilus.
Unlike oysters, where zinc is generally found in most tissues, zinc is
strongly localized in certain tissues of Mytilus. Tissue concentrations of
zinc in Mytilus edulis aoteanus have been reported in the order of visceral
168 1223
mass > gills > gonads > mantle > muscle = foot = shell. Pentreath
found visceral mass > adductor muscle > gonad > mantle = foot = gill. The
distribution of zinc-65 fallout was examined in Mytilus californianus, where
half the total zinc-65 was found in the kidney tissues constituting about 1% of
the total weigh; an additional 19% of the isotope was discerned in the digestive
163
-------
glands representing less than 4% of the weight of Mytilus. Only 4% of the
zinc-65 was in the shells, which constitute about 70% of the weight. Presumably,
168 1222
the "visceral mass" measurements just noted ' include kidney tissue.
The localization of zinc in the kidneys was confirmed by autoradiography of
1674
zinc-65 accumulated experimentally. However, in a related species of
Mytiltdae, Modiolus modiolus, rather uniform zinc concentrations were found in
gonad, mantle and gills, gut, and digestive gland, and they were 2-2.8 times
1467
higher than that in muscle tissue. Although it is possible that zinc
metabolism varies from species to species, it is more likely that greater
care should have been exercised during dissection to demonstrate the specific
tissue distribution of zinc.
Scallops (Pectenidae) also show strong tissue localizations of metals,
including zinc. Again, kidney tissue exhibits highest concentrations of zinc,
10-100 times the concentration for the total soft parts, in Pecten novae-
168 190
zelandiae, Pecten maximus L., and Chlamys opercularis L. High zinc
concentrations in the kidneys usually are accompanied by high concentrations
of manganese and iron168,190,1422,1467 Some of thft data190 show similar
relationships between kidneys and the various other tissues which also hold
true for cdbalt, copper, nickel, and lead, which are summarized in Table 6-7.
Although metal concentrations were usually highest in kidneys, the digestive
gland frequently contained a significant portion of the total metal present
in the scallop, because the kidney is such a small organ. Seasonal fluctua-
tions were evident in the kidney and digestive gland: zinc and manganese
concentrations were maximal in late autumn and winter and minimal during
164
-------
TABLE 6-7
Relative Content of Metals in Different Scallop Tissues
.a
Metal, %b
Aluminum
Cadmium
Chromium
Cobalt
Copper
i-*
gj Iron
Lead
Manganese
Nickel
Silver
Zinc
Gills
—
—
—
6.7
5.3
5.7
3.1
3.1
7.2
—
3.0
Pecten
Mantle
—
—
—
8.8
4.4
7.4
5.4
1.2
7.9
—
3.6
maximus L.
Digestive
Gland
33.9
89.9
46.9
50.3
60.9
71.9
19.0
1.4
45.9
63.7
16.2
Chlamys opercularis L.
Kidneys
0.7
1.7
1.7
23.9
1.5
0.6
52.3
92.5
20.0
1.5
51.8
Gills Mantle
— —
— —
—
11.5 8.7
7.9 4.6
9.7 8.4
2.1 1.8
5.7 1.7
9.1 6.1
—
7.4 5.1
Digestive
Gland
7.4
41.5
18.1
26.0
17.3
61.4
10.2
1.6
24.5
62.4
2.6
Kidneys
0.8
7.5
3.0
36.2
57.6
2.3
78.8
89.0
43.0
3.4
75.1
CL ^ „ 190
Data from Bryan.
^Percentage of total metal in soft parts, including fluid.
-------
190
spring, summer and early autumn. Bryan suggested that seasonal variations
may be caused by the reproductive cycle, temperature, availability of food,
and land drainage; he surmised that elemental changes in the kidney were
independent of the reproductive state of the gonads, and that changes in
phytoplankton productivity were probably more important than either temperature
or runoff. Generally^ periods of highest productivity coincided with lowest
metal concentrations in scallop tissues. Bryan further reasoned that the
greater availability of phytoplanktonic food would increase the metabolic
rate of the scallop and hence the excretion of waste products; reduce the
amount of metal left available in the water; and diminish the metal concentra-
tion per phytoplankton cell, thereby decreasing metal intake of the scallop
despite the additional food ingestion.
Freshwater bivalves. Freshwater bivalves are well known for their concentration
of manganese, ' generally accompanied by a lower concentration of
6^7 A^R ftAfi
zinc. ' ' As in the scallops, these metals are unevenly distributed
among the tissues of unionid bivalves. Anodonta nuttaliiana Lea has a mass
of tissue in which approximately 25% of the dry weight is composed of calcium.
This calcareous tissue contains metal-rich granules 2 ym in size, and is
present as an elongated white area on the surface of the mantle near the
C."1~J
attachment of the gills. Radionuclides of manganese, cobalt, zinc and
lead are rapidly accumulated to high levels in this tissue, whereas accumulated
radionuclides of scandium, chromium, iron, europium, tantalum, and mercury were
f."3Q
found in the organs of the digestive system. After 147 days of accumulation,
the gonad and foot of the clam appeared to have equilibrated with both zinc-65
and manganese-54, but the mantle and calcareous tissue were still accumulating
isotopes at an exponential rate.
166
-------
The biologic half-time for turnover in the large, long-lived pool
638
in Anodonta was about 1,300 days for manganese and 650 days for zinc.
Of the total zinc-65 accumulated in 35 days, about 35% was in the calcareous
tissue, whereas the proportion of stable zinc in this tissue was 42% in small
clams (in which average wet weight of tissue was 15 g) , and about 58% in large
637
clams (where wet weight averaged 45 g or more).
1214
In similar studies with Anodonta californiensis. Pauley and Nakatani
found that zinc-65 was associated predominantly with mantle and gill tissue,
but they did not distinguish calcareous tissue. Nor did their autoradiographic
637
techniques identify the granules observed by Harrison. Instead the auto-
radiography showed highest localizations of zinc-65 in the base and tip of the
outer mantle epithelium, the tips of the gill epithelial cells, and the
epithelial cells of the kidney. It seems probable that Pauley and Nakatani
simply included the calcareous tissue either with gills or mantle tissue, and
they probably did not section the organism through this region. A
similar judgment may be applied to the distribution of zinc-65 in tissue of
1024
Margaritifera margaritifera. In studies of whole animals, rapid equilibra-
183,648,1024
tion with zinc-65 has been noted for several species of unionids.
637
But the slow turnover and high concentrations of zinc in the calcareous tissue,
probably thwarted the achievement of true equilibration, and some of the biologic
648,1024 f s
half-lives obtained are probably not representative of zinc turnover
in these species.
167
-------
Freshwater gastropods. In a freshwater prosobranch, Viviparus malleatus, zinc-65
was accumulated to much higher concentrations in soft parts than in the shell,
183
but tissue distribution was not studied. Unborn embryonic snails contained
a higher concentration of zinc-65 than did the adults collected at the same
time from the experimental pond, suggesting that zinc mobilization into the
gonads and internally developing embryos is rapid compared to turnover in some
other organs of the adult snail. Stable zinc was not analyzed in this study;
183
therefore, dynamics of zinc turnover were not well defined.
Calcium accumulation and turnover were studied in the pulmonate
1678
Lymnaea stagnalis. Calcium was pumped against a concentration gradient by
this gastropod from low concentrations in the water ( < 1 rag calcium/1). The
calcium transport system was directly affected by temperature; the rate of
calcium uptake showed a Qin* of ^"^ between 6 C and 16 C and 3.0 between
16 C and 30 C. At least two rate components were resolved for calcium exchange
between Lymnaea and the medium. Although zinc was not measured in this study,
it is mentioned here because of the implied relationship in oysters between
calcium metabolism and the bioaccumulation of metals like zinc and manganese,
which was discussed in the section on oysters.
Crustaceans
Zinc-65 was assimilated with about 56% efficiency from ingested food
(zinc-65-labeled Artemia) by the gammarid amphipod Anonyx and retention of
338
the isotope was affected significantly by feeding and deprivation.
Amphipods starved during the 3-wk loss period exhibited a mean biologic half-life
Increase in rate of chemical reaction for each 10 C increase in temperature.
168
-------
for zinc of about 100 days, compared to about 35 days for animals on a brine
shrimp diet. Similar effects of controlling the food allowance on zinc-65
turnover were observed in marine Idothea1160 and crabs.192 Turnover of zinc
by Anonyx was also directly related to temperature; biologic half-life for
338
zinc turnover was about 92 days at 12 C and about 150 days at 3 C.
Similar effects of temperature were also noted with the isopod Idothea,
with the euphausids Euphausia pacifica and Thysanaessa spinifera,503 and
with the crab Callinectes sapidus. ^3-*9
Autoradiography of crustacean tissues indicates that zinc-65 is located
primarily in the interstitial spaces between muscle fibers, in the eye mainly
between the rhabdoms and cystalline cones of adjacent ommatidia, within and
on the interior surface of the exoskeleton, regardless of whether the
340 503
isotope was accumulated directly from water ' or assimilated from labeled
food. In euphausids, the prawn Pasiphaea pacifica and shrimp Pandalus
tended to concentrate zinc-65 accumulated from water more in the exoskeleton
and less in muscle than when the isotope was accumulated from food. In all
cases, however, the exoskeletons contained 30-66% of the accumulated zinc-65.
The turnover of zinc by growing crustaceans was not readily studied in
short-term zinc-65 loss experiments because a mean of 41% of the body burden
of zinc-65 was lost with the molted exoskeletons after zinc-65 accumulation from
503
water. In long-term loss experiments after ingestion of zinc-65-labeled
food, however, only 1% of the body burden was excreted with each molted
502
exoskeleton. This finding corresponded to the earlier radioautographic
observations that zinc-65 accumulated from water tended to be associated
169
-------
with interstitial spaces in the calcified cuticle of the exoskeleton, whereas
ingested zinc-65 was more localized on the underside of these cuticular
340,501,503
layers .
501
Fowler et al. speculated that the localization of zinc in euphausid
eyes might be associated with melanin pigments in the distal and proximal
screening pigment cells that closely surround the cones and rhabdoms of
814
euphausid eyes. Zinc also may be required by the enzyme retinene dehy-
drogenase, which catalyzes the oxidation of vitamin A alcohol (retinol)
to vitamin A aldehyde (retinene) . Zinc is necessary to
1664
many other nicotinamide adenine dinucleotide (NAD) -dependent enzymes.
Based on the above mentioned research on zinc-65 accumulation and
turnover in euphausids, and studies of zinc-65 accumulation by Euphausia
pacifica under conditions simulating those which euphausids would encounter
if they migrated vertically in and out of a zinc-65-labeled mixed surface
1499
layer of the ocean at different seasons in temperate latitudes, the
1501
turnover of zinc by the single euphausid Hegany ct iphanes norvegica
1500
as well as by total populations of this organism in the Mediterranean region
was estimated. Flux of zinc through Meganyctiphanes norvegica was presumed
to follow a linear model where:
Ke = Pe + Xe»
-L'
*e = Qf. pf+ Qm pm + Qc. p c. + ^ Px' and
°-i» Qfi » Qf » Qm» Qc» and QX are respective zinc concentrations
170
-------
in ingested food (i), new tissue added in growth C.6), feces (f_), molts (m),
dead carcasses (c), and nonviable eggs (x); and p± p6, pf, pm, PC, and px are
the rates of ingestion (i) and growth (6 ), and the respective rates of
production of f eces (f), molts (m), dead carcasses (c) , and nonviable eggs (x).
For adult animals feeding and defecating 12 h/day, zinc ingestion rate (K )
was estimated as 51-130 yg zinc ingested/g dry weight/day, depending on
whether maximal or minimal values of the other concentrations and rates
were used. The zinc concentration in the food of Meganyctiphanes norvegica
would have to be 400-450 yg zinc/g dry weight to satisfy the elimination
and growth rates of the model. Because zinc measured in Arte^-fa fed to
Meganyctiphanes norvegica in the laboratory (417 + 103 yg zinc/g dry weight)
and in natural plankton mixtures consisting of 95% small copepods plus phytoplank-
ton, flagellates,and detritus (570 + 113 yg zinc/g dry weight) hewed closely
to the calculated estimate, the zinc budget for Meganyctiphanes norvegica was
believed to be described adequately. Fecal pellet deposition represented
over 90% of the total zinc flux from the organism. For the Ligurian Sea,
the entire pool of ionic zinc in the water would be circulated through the
Meganyctiphanes norvegica population in 500-1,200 yr, depending on the dietary
1500
availability of zinc. Furthermore, zinc would be effectively transported
downward from the surface waters by sinking fecal pellets, molts, and carcasses
generated by the vertically-migrating euphausid population during daylight
hours. Daily net transport of 36-98% of the pool of body zinc in Meganyctiphanes
norveeiea would occur to a depth of 500 m, depending on whether food availability
was marginal or sufficient, and at least 6% of the body zinc pool would reach
2,500 m daily.
171
-------
1183
Osterberg et al. attempted to correlate the distribution of several
radionuclides including zinc-65 with the exposed surface areas of macroplanktonic
organisms, and concluded that surface adsorption played a relatively insignificant
503
role in the bioaccumulation of the nuclides. Yet Fowler et al. found that
weight-specific uptake and elimination of zinc-65 were statistically similar
between live and formalin-preserved euphausids over a range of temperatures
and zinc-65 concentrations. In euphausids which apparently do not regulate zinc
concentrations in tissue, the turnover and steady-state concentrations
of zinc probably depend entirely on the number and affinities of internal
502
binding sites for the metal. A similar hypothesis also was suggested for
zinc concentration and turnover in oysters.
In large decapod Crustacea (crabs, lobsters, and crayfish), zinc
concentrations in the body are controlled by mechanisms regulating zinc absorp-
1 Of.
tion through the gills and zinc loss in the urine and across the body surface. '
191-193
The following discussion is derived from Bryan's reviews of zinc
metabolism in decapods.
The zinc concentration in blood of lobsters (Homarus vulgaris) from
waters containing about 5 yg zinc/1 is similar to the value of 5.6 yg zinc/g
376
given for human blood. Whereas in human blood about 90% of the zinc is found
in the erythrocytes and leukocytes, in lobster blood 93% of the zinc is in
the serum. As in vertebrates, this zinc appears to be bound to blood protein^
which in the lobster is principally the copper protein, hemocyanin. Normal
hematic zinc concentration is highest in lobsters that have more hemocyanin,
measured by high contents of copper and solids in the blood.
In normal male lobsters, the highest concentration of zinc is found
in the hepatopancreas: about 25 ug/g. High zinc concentrations were not
172
-------
found to be associated with the male reproductive system in the lobster (fhe
vas deferens contains about 13 yg/g). But the highest zinc concentrations in
normal female lobsters were found in the ovary, which contains up to 50 pg/g.
No other obvious differences between the sexes were observed. Zinc concentra-
tions in normal lobsters are relatively low compared to vertebrates, but are
very similar to the zinc concentrations of 20 yg/g found in whole shrimp
136,1209
and crabs.
193
Bryan has showed that zinc concentrations in most lobster tissues
are quite accurately controlled. Except in the hepatopancreas, variations
produced by zinc injections were rectified quite rapidly. In muscle tissue
and gonads no changes were induced by zinc injections, suggesting that these
organs are either almost impermeable to zinc or that zinc regulation is particularly
good.
The process of zinc regulation in the freshwater crayfish (Austropota-
mobius pallipes) differs from that in the marine lobster Homarus because
most crayfish tissues contain less zinc than those of the lobster, but the
concentrations in the hepatopancreas and stomach fluid are much higher.
Concentrations in the main abdominal flexor muscles are similar in the two
species, however, and zinc concentrations in muscle are fairly constant under
different conditions. In both species, muscle is the tissue responding least
to changes in the blood concentration of zinc, although zinc exchange does
193
occur.
Crayfish blood contains less zinc than the blood of any marine
decapod crustaceans that have been examined, although in estuarine species
blood zinc was higher than in marine species. The blood concentrations of
zinc do not increase in crayfish, despite high concentrations of blood protein
and copper which tend to increase amounts of zinc in Homarus. Zinc is bound
173
-------
to proteins in the blood, but binding to the variable hemocyanin component may
be less important for crayfish.
Low zinc concentrations for crayfish have been found consistently in
the excretory organs and urine. The excretory system appears to be unimportant
in regulating zinc, although it certainly prevents zinc loss. The permeability
of the body surface to zinc is low and therefore zinc losses are small. Similarly,
very little zinc is absorbed across the body surface from solution. In contrast,
Homarus absorbs and loses zinc across the body surface, apparently in a controlled
manner. Urinary losses in Homarus are also closely controlled, yet they can
192
be appreciable.
Both species are likely to obtain excess zinc from food, and it is the
major source of zinc for the crayfish. Excess zinc is absorbed rapidly from
the stomach fluid in Homarus, partly by the hepatopancreas, and concentrations
will increase in all tissues except muscle, gonads and exoskeleton. Uptake
from the stomach fluid of the crayfish is slower, and all the zinc is absorbed
by the hepatopancreas before affecting the other tissues. If zinc penetrates
directly from the stomach of the crayfish into the blood, it is removed so
rapidly by the hepatopancreas that no obvious change in blood concentration
is seen. When excess zinc was injected into crayfish blood, all the excess
zinc.was absorbed by the hepatopancreas in a few days and some of it was
transferred to the stomach fluid. When excess zinc was injected into Homarus,
some was removed in the urine, some was lost across the body surface, and
as in the crayfish, an appreciable amount was removed by the hepatopancreas.
Whereas in the crayfish excess zinc in the hepatopancreas and stomach fluid
174
-------
was lost in the feces, in Homa.rus excess zinc in the hepatopancreas was
eventually lost by blood, excretory organs, or body surface.192
The amount of zinc absorbed directly from sea water across the body
surface of decapod crustacea varies with the zinc concentration in the water.
Hence species living in fairly clean water away from the coast will absorb
less zinc from the water than species living in lightly polluted estuaries.
Variation in the dietary intake of these metals is, however, probably much
more important. Animals which feed on worms from sediments may take in
considerable zinc and copper. Although the intake of metals may be highly
variable, the concentrations of zinc and copper in the majority of species
lie between 20 and 35 yg/g. Therefore, it seems likely that zinc regulation
occurs in all species of decapod crustaceans.
Zinc is bound so tightly by the proteins in the blood that even in waters
containing only a few yg/1 of zinc, the concentration gradient for unbound
zinc points toward penetration from the water to the blood through the gills.
The amount of zinc absorbed in this way increases as the external concentration
is increased, indicating that the proteins in the blood to which zinc binds
189
are not normally saturated. So far, no unequivocal evidence has been
obtained to show whether or not absorption via the gills is an active or
controlled process.
Decapod crustaceans probably receive more zinc than they require,
and thus processes for removing extra zinc probably are more important than
measures for its absorption. The removal of zinc from the body can occur
through the feces, urine, or across the body surface. Losses in the feces
depend on whether the animal is feeding and how much zinc is in the stomach
fluid. In Homarus, for example, the zinc concentration in stomach fluid is
175
-------
usually less than 1 ug/g and losses In the feces are very small. In freshwater
192
crayfish, almost all losses of zinc occur in the feces.
The ability to regulate the concentration of zinc—and perhaps the
concentration of copper—in the body may give decapod crustaceans a degree of
protection in regions where metal pollution is found. The concentrations of
zinc and copper in muscle tissue are unlikely to vary, but the hepatopancreas
°f Cancer pagurus (which is consumed by humans) would probably yield higher
186
concentrations of metal in polluted regions.
Polychaetes
in the Newport River estuary 339
In a study of manganese, iron, and zinc/in North Carolina, Cross et al.
found that three polychaete worms (Glycera americana, Diopatra cuprea and
Amphitrite ornata) showed similar concentrations of metals regardless of large
differences in metal concentrations of the sediments in which the worms were
burrowing and feeding. Therefore they suggested that these species may regulate
195
body concentrations of trace metals. Bryan and Hummerstone examined Nereis
diversicolor, a species closely related to Glycera, and found that whereas
the concentration of copper was roughly proportional to that of the surrounding
sediment) zinc appeared to be regulated independently of the zinc level in
194
the sediment. It subsequently was found that zinc in Nereis varied by a
factor of only 2.7 although they were living in sediments where zinc concentra-
tions varied by a factor of about 30. In these same samples, concentrations
of cadmium in the worms were roughly proportional to those in the sediments,
indicating that cadmium—like topper—was probably not regulated, whereas
zinc was. In experiments to induce toxicosis, worms from sediments high in
176
-------
zinc were more resistant to zinc than were normal worms. This adaptation of
Nereis was ascribed to a reduced permeability to zinc and more effective excretion.194
Fish
Theoretical considerations of limited experimental evidence show that
fish must obtain zinc from their dietary intakes rather
726 1222
than through exchange with dissolved zinc in the aqueous medium. ' This
conclusion was deduced from the observation that turnover rates of larger body
pools of zinc cannot be sustained by inflow rates of zinc from water alone.
Such a conclusion again points to the shortcomings of turnover rates estimated
from short periods of exposure to radioactive zinc. Estimates of zinc turnover
in fish are listed in Table 6-6, along with certain aspects of the experimental
approach. Longer turnover times are obtained naturally when the fish are
1332a,1679,1680
exposed for long periods to zinc-65 in the natural environment.
Similarly, the use of short-term exposures of fish to experimental dissolved
zinc concentrations in tests for acute toxicosis is of questionable significance.
This practice usually provides estimates of lethal levels that can cause death
through tissue hypoxia from direct gill damage or precipitation of gill secretions.
Under these experimental circumstances, the organisms die before their body pools
have equilibrated with the environmental level of zinc, and the significant
lethal or sublethal physiologic effects of longer term exposure tend not to be
studied.
Zinc concentrations in whole fish are a function of age in certain
337
species, at least in juvenile stages. In croakers (Micropogon undulatus),
bay anchovies (Anchoa mitchilli) and menhadens (Brevoortia tyrannus), zinc
177
-------
concentrations decreased as an exponential function of weight over the range
of 0.01-5 g dry weight for the whole fish. In adult bathyal-demersal morids
(Antimora rostrata), zinc concentration in white muscle also decreased slightly
with size between 200-1,400 g total live weight, but zinc concentration was
independent of fish size in muscle of bluefish (Pomatomus saltatrix) between
I/ "I
400-4,500 g. In freshwater carp, the initial zinc concentration in roe
decreased with the age of the female, although after gastrulation the zinc
1389
concentrations in the larval fish were similar regardless of the parent's age.
Tissue distribution of zinc-65 in yearling trout was observed after oral
administration of a single dose of radioisotope contained in a force-fed
gelatin capsule. Initial high blood concentrations of zinc-65 declined
exponentially after 1 day, whereas the isotope was retained for 2-3 days in
kidney, spleen, and liver and continued to increase in gills through the fifth
day. Muscle, bone, and eye tissues appeared to have equilibrated with zinc-65
by the fifth day and were retaining the isotope more effectively than the other
tissues, albeit at a lower concentration than gills, kidneys, or spleen. This
experiment suggests that zinc elimination from fish may occur both through
urine and gills.
The daily flux of manganese, iron, copper, and zinc was estimated for
populations of Atlantic menhaden, spot and pinfish for the summer months in
342
the Newport River estuary in North Carolina. Assimilation efficiencies of
these metals by fish were highly variable and dependent on the trace metal
concentration in inorganic content of ingested materials. Except for zinc in
menhaden and pinfish, assimilation efficiencies were less than 10%. Because
a significant fraction of the trace metal ingested is not assimilated, defecation
178
-------
of unassimilated trace metals by these fish may be a major biologic process
in cycling trace metals in highly productive coastal plain estuaries.
By comparing analyses of manganese, iron, and zinc in estuarine organisms,
sediments and water, Wolfe et al. 1801 estimated the fluxes of these metals through
and within the Newport River estuary and a descriptive model was constructed
for the annual movement of zinc in that estuary.1798 Zinc appears to be
retained very effectively in coastal plain estuaries: sediment and detritus
reservoirs are large, and efficient recycling occurs through the living organisms.
This model was subsequently examined-'-7" for sensitivity of the major zinc
flows around the detritus compartment toward the other conditions, either
measured or assumed, used in the development of the model. The model was most
sensitive to phytoplankton incorporation of zinc, microbial zinc assimilation
within the detritus compartment, and macrofaunal standing crop, production, and
assimilation of zinc. The model was insensitive to changes in Spartina productivity,
phytoplankton standing crop, zooplankton standing crop, and productivity and
estuarine flushing rate. It is still poorly understood how zinc adsorbed on
sediments or incorporated in detritus becomes remineralized and available again
for bioaccumulation.
Metabolism of Radioactive Zinc
Zinc-65 introduced into the Columbia River by the Hanford nuclear installa-
tions was accumulated by various organisms in the adjacent North Pacific Ocean.
Studies of zinc-65 specific activities in fish and prey organisms have produced
estimates of intake rates of zinc-65 in the predator fish species and hypotheses
1216,1679,1680
of mechanisms of zinc-65 transport in the ocean. As mentioned,
179
-------
content of zinc-65 decreased markedly from 1965 to 1971 when the reactors were
shut down. Specific activities decreased faster in small individuals of the
flounder Lyopsetta exilis than in large individuals, indicating more thorough
labeling of longer-lived pools of zinc in larger fish. Specific activity
also decreased with increasing depth and with increasing body size for both
Lyopsetta exilis and the rockfish Sebastolobus.
From extensive time series of zinc-65 specific activities in Lyopsetta exilis
1680
and prey organisms of this species, Vanderploeg used the model
£ = a (F(t) - S) - S X,
at
where j[ = zinc-65-specific activity of the predator; a = rate of zinc input
as fraction of zinc body burden in the fish; J_(t) - zinc-65-specific activity
of the prey; and X = physical decay constant of zinc-65. He calculated the
daily rate of input of zinc as a fraction of the body burden of zinc in the
fish. Values of 0.0027 day" and 0.0026 day" were obtained for fish of 22 g
and 35 g wet weight.
Zinc-65 from the Columbia River was apparently in a form more available
to the Lyopsetta exilis food chain than stable zinc in seawater was to it.
1679
Vanderploeg hypothesized that several mechanisms could contribute to this
conclusion. For example, some soluble zinc-65 in the river is rapidly
accumulated by plankton and other small particles. A portion of the zinc-65
446
entering the ocean as suspended particulate material is desorbed and might
also be sorbed to particles of marine origin. Herbivorous zooplankton feed
on the particles, transform them into fecal pellets, and they, as well as
settling particulate matter, deliver high specific activity zinc-65 to the
180
-------
continental shelf. Distribution of zinc-65 specific activity in the pelagic
food web and in fauna at different depths and distances downstream from the
river mouth was consistent with these hypotheses. Vanderploeg1679 found that
zinc-65-specific activities in suspended particulate were greater than in the
soluble form. His observation is consistent with the existence of stable
unreactive complexes of zinc in seawater. Bernhard and Zattera observed
that phytoplankton could accumulate zinc-65 and stable zinc differentially
1249
if they were available in different physicochemlcal forms. Piro et al.
found that ionic zinc added to seawater equilibrated with the ionic and
particulate zinc already present, but that the amount of complexed zinc remained
unchanged. Zinc-65, when added in the form of an ethylenediaatinetetraacetic
acid (EDTA) complex, however, was distributed among all forms
in the same proportion as the stable element. Consequently, the presence of
complexed stable zinc in the ocean would promote biologic accumulation of
zinc-65 in the ionic form from organisms which selectively accumulate ionic or
particulate zinc.
Correlation of Zinc Turnover with Respiration
Retention of zinc-65 has been proposed and used with varying degrees
of success as an indirect measure of respiratory metabolism in a variety of
organisms.1069*1159'1160 A very high positive correlation was demonstrated
between zinc-65 turnover and oxygen consumption in young plaice, when
417
temperature was the only experimental variable. Similar observations
have been obtained with mice. ' Shulman et al. found no effect
of varying food intake on the turnover of zinc-65 in Menidia menidia and
181
-------
Fundulus heteroclitus; nor was any effect of varying dietary zinc concentration
727
on zinc-65 turnover found in Lagodon rhomboides L. For zinc-65 turnover
to be a valid measure of respiration in the natural environment, both feeding
rate and temperature must respond similarly and consistently to environmental
changes affecting either function. Thus both feeding rate and temperature—
which affect respiration—must be correlated to zinc-65 excretion rate, as has
been demonstrated for temperature; and respiration must be correlated with
environmental zinc concentration, or the turnover of zinc by the experimental
organism must be regulated independently of environmental zinc.
In euphausid shrimp, respiration was poorly correlated to zinc-65
503
accumulation rates as either a function of temperature or individual dry weights.
Turnover of zinc in euphausids was not metabolically controlled and was likely
a surface phenomenon, suggesting a strong interdependence on both temperature
and zinc concentration. Thus, the utility of zinc-65 loss as a measure of
respiration under natural conditions may be restricted to vertebrates. For
fishysuch a remote technique is especially desirable in natural conditions to
avoid the excessive respiration produced by the stress of being confined in
experimental respirometers.
TOXICITY OF ZINC TO AQUATIC ORGANISMS
How zinc and other heavy metals are toxic to fish has been studied since
921
the mid-1920's 39^» 1^90and the subject recently has been reviewed. Acute
heavy metal toxicosis in fish has been attributed to the coagulation or
precipitation of mucus on the gills and/or to cytologic damage to the gills.
The physiologic mechanism of death from either cause is related to a breakdown
182
-------
208
in gas exchange at the gills. Burton et al. studied acute zinc toxicosis
in rainbow trout (Salmo gairdnerii) and was able to support the hypothesis
that modification of the gas exchange process at the gills creates hypoxia at
the tissue level. Tissue hypoxia appeared to be the major physiologic change
preceding death once the gas exchange process at the gills was no longer
sufficient to supply the oxygen requirements of the fish.
Daphnia magna has been used as a representative of common and abundant
zooplankton to study acute and chronic toxicosis produced by several metals.112
In 48-h tests for acute toxicosis, the median lethal concentration (LC ) for
zinc was 100 yg/1 when no food was provided and 280 yg/1 when food was added
to the water. In 3-wk studies for chronic toxicosis, reproductive impairment
was found to be a more sensitive measure of toxicosis than survival. The 16%
reproductive-impairment concentration was used as the criterion, as values
below this level could not be detected from the controls because of variability
within groups. The 16% reproductive-impairment concentration for zinc
was 70 yg/1. Because the organic material added as food was found to alter
the potential for toxicosis, the results may not be directly applicable for
any particular water.
Waller et al. evaluated data from two studies on fathead minnows
(Pimephales promelas) populations from three lakes. They concluded that the
maximum concentration of zinc to which fish could be continuously exposed
should not exceed 1/100 of the 96-h median tolerance limit (TL5Q)—a concentration
that caused a 50% reduction in the mean number of eggs laid per female fathead
minnow in a laboratory study.
The Environmental Protection Agency has suggested that bluegills be used
1529 1009
as a warm water species in bioassays. McDonald and Heimstra showed that
183
-------
bluegills (Lepomis machrochirus) are very agressive and set up dominance-
submission relations when confined in aquaria. They found that dominant fish
survived an exposure to 32 mg/1 zinc longer than submissive fish. A shelter
placed in each compartment was found to reduce the number of agressive encounters
between fish and reduced the response difference. The results indicate that
dominance-submission relations may be a variable for the outcomes of bioassays
with fish.
216
Cairns et al. studied the effects of pH, solubility, and temperature
on the ability to produce acute zinc toxicosis in the bluegill sunfish (Lepomis
machrochirus). Bluegill sunfish were exposed for 96 h to water-soluble zinc
sulfate and water-insoluble zinc phosphate at two tem-
perature ranges (21-24 C and 7-9 C) and two pH ranges (5.7-7.0 and 7.3-8.8). Control
fish were maintained in water containing no zinc. No bluegills died in water
containing particulate zinc in amounts comparable to the amounts of soluble
zinc (13.5, 18.0, 24.0, and 32.0 mg zinc, which produced mortalities of 90-100%.
Bluegill mortalities in concentrations of soluble zinc ranging from 10-32 pg
zinc/1 were zero to 10% at the high pH, whereas at the low pH, where ionic
zinc would more readily dissociate from the zinc sulfate, mortality was 100%.
Bluegills acclimated to the low temperatures died at a much slower rate, and
the time-to-death of the first fish was considerably delayed in comparison
to bluegills at the warmer temperature. At the concentration of zinc tested
(32 yg zinc/1), the acclimation temperature had no effect on the percentage
of fish surviving at the end of the 96-h test period. The acclimation
temperature did, however, affect the percent survival of the bluegills at
24 and 48 h.
184
-------
The toxicity of zinc to the larvae of crab (Carclnus maenas) during periods
309
up to 64 h has been examined. At a concentration of 1 ppm zinc, the median
effective time for 50% mortality (ET ) was 47 h. For higher concentrations of
zinc, the log of the concentration and the ET were related linearly. The level
of 1 ppm of zinc is about 100 times the concentration found in natural seawater.
For periods longer than 47 h, however, the LC would be cnnsiderably lower
and much closer to the level of zinc found in natural seawater. Thus, elevation
of zinc levels in confined water to values substantially above natural levels
could have a severely deleterious effect on survival of larvae. The toxicity
of several metals, including zinc, to oyster embryos in synthetic seawater at
218
25% salinity and 26 C, also has been tested. For 48-h exposures, 0.075 ppm
zinc produced no mortality, and 0.5 ppm zinc produced 100% mortality; the
estimated LC (48 h) was 0.31 ppm zinc.
185
-------
CHAPTER 7
ZINC IN HUMANS
The roles of zinc in the normal physiology and biochemistry of mammalian
. . . . .. , 362c,504,609-614,1044, 1274,1275,1285,1286,1640
systems have been reviewed widely. * ' ' ' ' ' ' '
The purpose of this chapter is to describe major aspects of the role of zinc
in humans.
ZINC CONCENTRATION IN BLOOD. URINE. AND FECES
Zinc is found in every human tissue and tissue fluid, although concen-
trations vary in different fluids and tissues. Zinc is present in nuclear,
mitochondria!^ and supernatant fractions of all cells that have been examined
by ultracentrifugation. Total body zinc for a hypothetical 70 kg man may
*
be estimated to be 2.3 g, making it the most prevalent trace metal in tissue.
Table 7-1 compares the estimated zinc concentrations of several tissues in a
hypothetical 70 kg man. Clearly, the major amount of zinc in the total body
resides in muscle and bone (approximately 90%), although the highest con-
centrations of zinc is found in tissues from the reproductive tract.
Blood
For convenience, mean serum zinc concentration in humans may be con-
sidered to be approximately lOOyig/dl (see Table 7-2 for the ranges involved)
and is the same in healthy men and women. ' Reports that differences in
935
plasma zinc occur between mean and women have appeared, but they have not
been careful^ substantiated. Deviations from the mean of 100^ug/dl may be indicative
"e
Iron, which exists in higher concentration, is found primarily in blood.
For a human weighing about 70 kg, the total concentrations of trace metals
are estimated as follows: iron, 4.0 g; zinc, 2.3 g; manganese, 0.2 g;
copper, 0.1 g; and all others combined, ^Q.l g.
186
-------
TABLE 7-1
Tissue
Adrenal
Aorta
Bladder
Blood
Bone
Brain
Gastrointestinal tract
Heart
Kidney
Liver
Lung
Muscle
Ovary
Prostate
Skin
Spleen
Testes
Thyroid
WHOLE BODY
in Some Human
Zinc,
Ug/g wet wta
6
26
22
1
66
13
21
27
48
27
14
48
12
87
6
19
13
25
33
Tissues
Zinc,
mg/organ
0.9
2.6
4.4
6.0
660-0
18.0
25.2
8.7
1-9.8
40.5
16.6
1,420.0
0.3
1.7
30.0
3.8
0.8
0.4
--
Zinc, % of total
body (70 kg man)
—
0.1
0.2
0.3
28.5
0.8
1.1
0.4
0.9
1.8
0.7
62.2
—
0.7
1.0
0.2
—
—
__
TOTAL
2,259.7
98.9
Measurements from Forssen,491 Soman et al.,1522b Tipton and Cook1620 and
Tipton et al.1621
187
-------
of disease or environmental conditions, and such changes will be discussed in
Chapters 8, 10, and 11. Estimates of zinc in the serum, plasma, and other fluids
of normal adults are set forth in Table 7-2.
Measurements of serum zinc are influenced by many factors. Different
techniques of measurement may produce varying results. Sample contamination
(a major problem, particularly if glass syringes or collection tubes or rubber
or cork stoppers are employed), hemolysis, and the addition of various agents
to the sample tend to make values higher. Serum zinc is about 16% higher than
205
plasma zinc, the higher percentage reflecting differences in zinc liberated
from platelets204,482 as weii as differences in volume, hemolysis and other
AQO
unidentified factors. Circadian variation also affects blood zinc levels:
693 932
values in the afternoon are higher than in the early morning. ' Zinc
levels in serum for 2-3 h following ingestion of food have been found to be
lower than those measured during the fasting state. '
Zinc is also an important constituent of red blood cells, representing
approximately 10 times the amount of zinc found in serum. Erythroblasts mea-
sured by histochemical techniques do not appear to contain zinc, suggesting that
1588
carbonic anhydrase may be found only in erythrocytes. Systematic chemical
analyses have not yet been performed. Zinc also is present in young and mature
1588 377
reticulum cells. Leukocytes contain more zinc than erythrocytes. Granu-
locytes, particularly eosinophils and basophils, are relatively rich in zinc
q
(14.2 yg zinc/10 cells). Zinc has been incorporated into metamyelocytes, and
1588
the content of the metal increases as granulocytes mature. The content of
zinc in peripheral blood granulocytes was found to be about 30% higher than in
1588
bone marrow granulocytes. Most zinc in leukocytes is protein-bound and can
1660
be isolated in a purified form. The function of zinc in leukocytes has not
been systematically investigated; however, leukocytes contain the zinc-
dependent enzymes alkaline phosphatase and peptidase, indicating that some zinc
1588,1660
in leukocytes may be associated with the activities of these enzymes.
188
-------
TABLE 7-2
Authors
Prasad et al.1277
Butt et al.213
Gofman et al.542
Parr and Taylor1210
Kahn et al.809
Prasad et al . 1283
Helwig et al. 661
Parker et al.1208
Davies360
Hackley £t al.595a
970
Mahanand and Houck
Withers et al. 1794
Woodbury et al.1804
Meret and Henkin1031
Number of persons
19
37 men
45 women
39
6
97
14
64
23
30 men
30 women
96 men
97 women
7
25
11
82
45 women
37 men
Mean value ± SD
Serum or plasma,
102 ± 13
157
159
98 ± 5
85 ± 7
84 ± 30
104 + 14
91 ± 17
90 ± 10
95 ± 13
96 ± 10
96 ± 13
94 ± 29
110 ± 21
• 85 ± 15
92 ± 18
90 ± 20
94 ± 18
Range
Vi g zinc/dl
80-99
76-125
72-120
86-102
63-147
189
-------
TABLE 7-2 continued
Authors
Number of persons
Mean value ± SD
Range
Dennes et al."'
Brune et al. *-°*-
58
Auerbach
Kleinman jet al. 852
Whole blood, yg zinc/ 100 ml
28
7
30
15
560 ±
680 ±
734 ±
545 ±
20
8
186
18
408-1170
Talbot and Ross
1592
Dennes et al.
Prasad et al.
377
1277
Valberg et al.
1651
Auerbach
58
Rosner and Gorfien
1369
Red blood cells
30
15
57
30
23
11.8 ± 1.8yg/g blood 8-14.9
0.97 ± 0.03yg
zinc/109 cells
12.5 ± 1.2yg/ml blood 11-14.8
10.6 yg/g blood 7.7-14
1.6 ± 3.3yg x 9.6-25
10^ erythrocytes
12 ± 2yg/1010
erythrocytes 8.6-16.1
Leukocytes
Dennes et al.
377
30
Frischauf et al.
510
14 ± 1.9 yg
zinc/10-9 cells
5 yg/g dry weight
Szmigielski and
Litwinl588
Units
Granulocytes 10
bone marrow metamyelocytes
juvenile cell
polymorphonuclear cell
peripheral blood
94 ± 8.21
112 ± 7
139 ± 14
178 ± 12
190
-------
TABLE 7-2 continued
Authors
Number of
persons
1667
Vallee £t al.
Prasad et al.
Prasad et al.
Helwig et al.
Meret and Henkin1031
1283
1283
661
Urine, pg zinc/24 h
14
5
5
62
82
(45 women)
(37 men)
Mean value ± SD
457 ± 120
658 ± 206
658 ± 202
525 ± 254
353 ± 207
347 ± 322
360 ± 128
Range
273-660
145-1,256
141-179
191
-------
Despite the numerous sources of zinc in blood, total circulating hematic
zinc represents less than 0.5% of the total body zinc. This amount contrasts
with circulating copper in the blood, which represents more than 6% of total
body copper but is consistent with the relative percentage of circulating
hematic manganese.
Concentrations of zinc in blood plasma vary considerably during the first
/1QO fld£0
two years of life, as shown in Figure 7-1. At birth, plasma zinc levels
approximate those of normal adults, but within the first few days to a week
of life, zinc levels decrease to less than one half of the means shown by
C0O
normal adults. °^ Zinc concentrations in infant plasma appear to remain at
lower than adult levels for the first five or six months of life; then values
rise again to the normal adult range, albeit at its lower end. An abrupt
fall in serum zinc levels occurring at about one year of age was observed in
f O O
one group of normal infants. Before 15 months of age, human plasma zinc
values remain normal except when influenced by physiologic or pathophysiologic processes,
Such changes may be characteristic only of some infants; although clearcut patterns
of change have been observed in American infants, few fluctuations have been measured
in Western European babies.
In adult life, the circadian variation found in plasma or serum zinc is
significant.204'6603'693'932'17153 It is generally agreed that zinc levels
in serum are at a minimum at 6 a.m., with the maximum occurring at 10 p.m. After
10 p.m. ». values drop rapidly and consistently. They rise rapidly after the nadir at
GOO
6 a.m., and remain relatively stable over the next 8 h. J The pattern of
circadian variation is somewhat similar to the circadian pattern observed in
\
the secretion of adrenal corticosteroids, although the specific timing of the
nadir and zenith is displaced by approximately 4-5 h.
192
-------
i Plasma Zinc
T (MiiSEM)
0246
Days
6 9 12
Months
TIME
18 21
FIGURE 7-1 Total plasma zinc concentrations during the first two
years of life.. Hatched area indicates normal adult
levels (mean+1 S.D.). During the first week of life
and at 2, 3, and 12 months of age, plasma zinc
concentration was significantly below the lower limit
of the normal adult range. All points represent
determinations from three or more infants. Where not
indicated, all the SEM's were too small to be visible
on the graph. Reproduced from Henkin et al.
193
-------
MACROMOLECULAR-LIGANDED MICROMOLECULAR-LIGANDED
TIGHT
LOOSE
Zfl I °^Mocroglobulinj"d^'
32%
Albumin
66%
lAmino ocidsl
(histidine
cysteine)
2%
FIGURE 7-2 Bindings of serum zinc. Reproduced from Henkin.
668
194
-------
Zinc in serum is always bound to some ligand, as illustrated in figure 7-2.
Approximately 43% of the zinc circulating in blood serum is bound to al-
. . 537,1205,1692,1802
bumin, presumably to one of the histidine
CQf.
moieties of this molecule. However, it has been suggested that zinc binds to
the free carboxyl and not to the imidazoles of the histidine
residues of albumin.1226 It is chemically possible for each of the 16 histidine
moieties of albumin to bind one zinc molecule,-^ but albumin in human serum is
normally undersaturated in zinc.^37 The zinc-albumin complex has an association
constant of approximately 10° and has been called the major macromolecular
zinc ligand.537'-668 This albumin-bound zinc is in equilibrium with amino acid
zinc complexes that comprise about 1-2% of the circulating zinc. * This
latter group of ligands, called micromolecular zinc ligands, are almost exclusively
some form of the amino acids histidine and cysteine. ' These amino acid^zinc
ligands are available for transport to all tissues, including body organs, red blood
cells, and brain. Histidine and cysteine easily cross the blood-brain barrier.
537,668
Porphyrin-bound zinc may serve as another micromolecular zinc ligand, although
it may not be of particular quantitative importance in normal humans. Small
concentrations of zinc-porphyrin complexes have been measured in normal subjects
and larger concentrations are present in patients with various types of porphyria.
Approximately one-third of the zinc carried in blood serum is bound to an
a2-macroglobulin.^6»D68» 1205,1226 This macromolecular zinc ligand, a zinc-
protein complex with an association constant greater than 10 , is not in
equilibrium with the albumin-zinc complex or the amino acid zinc complexes.537,668
Zinc is incorporated into this o^-macroglobulin only in the liver and the com-
plex is metabolized only there.
Zinc in serum may be divided functionally into diffusible or nondiffusible
fractions. Diffusible serum zinc is composed of the amino acid-bound zinc and
195
-------
682
the freely exchangeable or more readily removable zinc from albumin.
682
This zinc has been quantitatively determined by ultrafiltration. The nondiffu-
sible zinc has been distinguished by ultraf iltration to represent ligands with a
molecular weight greater than 50,000. These distinctions have clinical value in
the analysis of abnormal zinc metabolism, as noted in Chapter 10.
The distribution of zinc in human serum has also been studied by
measuring the partitioning of zinc between the two major macromolecular zinc
COT
ligands in serum. J/ As mentioned, albumin-bound zinc is both the major
and the more dynamic of the two. However, it is possible to measure the
a^-macroglobulin component of human serum,1^20a thereby obtaining an estimate
of its relative importance. Whereas concentrations of albumin-bound zinc and
total serum zinc were highly correlated (p<.01) as were the concentrations
of albumin and albumin-bound zinc, c^-macroglobulin-bound zinc was not
significantly correlated with either total serum zinc or with the serum
concentration of c^-macroglobulin.
Urine
Approximately 500 yg/24 h appears in the urine of healthy -subjects.
This zinc content reflects changes in nutrition, physiology disease, and the
environment. In normal subjects, urinary zinc is composed of zinc pri-
marily bound to amino acids and secondarily to porphyrins. Zinc in urine
apparently is not correlated with any of the known variables that determine
the behavior of serum zinc. Whereas there is a clear circadian variation
in serum zinc, none has been observed in urinary zinc excretion.^ This
inability to pinpoint a circadian variation in urinary zinc contrasts with
the discovery of a circadian variation in serum and urine copper^2 an^ may
indicate some basic differences in the manner by which the metals are handled by
196
-------
the kidney. The unhindered passage of zinc-amino acid complexes in normal
blood across the renal glomerulus would result in a calculated filtered load
of 2 mg of zinc in a normal 24-h glomerular filtrate of 18351. Thus, the
major part of the normal filtered load of amino acid-complexed zinc (and
presumably, porphyrin-complexed zinc) must be reabsorbed by the kidney, al-
though the exact nature and characteristics of the reabsorption have not
been well studied.
Feces
Zinc in feces represents the major source of zinc lost from the body.
The zinc content of feces varies with the zinc content of the diet, and,
as such, varies from 5-10 mg daily, and roughly 70-80% of ingested zinc is found
in the stool. Zinc in feces also is composed of zinc secreted from the
gastrointestinal tract and the bile. Although reabsorption has not been
well studied in man, estimates from the rat suggest that zinc is secreted
362b
into the gut lumen and that one-third of this zinc normally is reabsorbed.
Zinc in meconium of normal babies varies from 39-569 Vg/g dry weight, with a mean
of 230 iig/g dry weight. The amount of zinc in meconium was found to be approxi-
mately three times the amount of iron or copper and 10 times the amount of
Q/r «y
manganese. Infants with cystic fibrosis of the pancreas, meconium ileus,
and intestinal obstruction were reported to exhibit lower mean levels of zinc
O /• T/»
in meconium than did normals.
Other Body Fluids and Tissues
Zinc in sweat is about 115 pg/100 ml. Under conditions of extreme heat,
zinc losses through this route have been reported to increase to as much as
97 ntA „ 309a,720a,1288,1585a,1816b 1355 U7A
2-3 mg/day. These and other 1JD-5»-L^/^ data
indicate that the zinc content of sweat can be greater than that of serum.
197
-------
Zinc concentration in human milk was found to be higher than that of
any other trace element. 1101'16/t° The level of zinc in human milk varies
with time after delivery. In colostrum the zinc level was found to be
3-5 times the level in milk found 1 wk after parturition, after which
675 996
the level fell slowly over the next 6 months. ' It is evident that
zinc levels in human milk are carefully controlled, and maternal levels of
prolactin have been suggested as an important factor in such control.
An average zinc level in human milk 4-6 mo after parturition is 50 yug/100
ml. In milk from cows delivered to market in the U.S., zinc content varied from
1.3-12.4 ug/100 ml with an average of 4.4 ug/100 ml. ' '
As would be expected, cows' milk stored in galvanized cans was higher in
zinc, varying from 55-154 ^ig/100 ml. " Zinc content of cows' milk is re-
portedly constant all year, but it will vary somewhat with geographic
1101 other factors affecting bovine metabolism.
location, diet, and / The importance of the zinc content of milk as an
essential nutrient is suggested by a report of zinc deficiency in newborn
/ 1 O
mice nourished by a mother lactating for a fairly long time after parturition.
Small amounts of zinc also appear in. the cerebrospinal fluid (CSF) .
Zinc has been found in various secretions of the gastrointestinal tract, in-
eluding saliva, 674, 678, 684a, 1087 gastrlc juicej1532,1569 gall biadder bile,1569 and
duodenal and pancreatic secretions. Zinc is also an important constituent
of cerumen. Its concentration is over 1,800 yg/g, roughly 300 yg/g greater than
the amount of calcium found, 50 times greater than the concentration of
151
copper, and 5 times greater than the concentration of magnesium.
198
-------
Zinc has been found in sebum excreted from human skin.208*'374'1557 By
measuring excretion rates of sebum, 344b,1558a tne amount of sebum may be
z±nc content estimated. In male weanling rats made zinc-
deficient, sebaceous gland hypertrophy was observed despite depressed pituitary
and testicular function. From these observations, hypotheses were advanced that
disorders of sebum production such as acne might be related to disorders of
zinc metabolism. ' Patients with acne treated with oral zinc sulfate
were said to show decreased sebum production and remission of acne. '
208a
These findings have not been reproduced.
In 1940, Eggleton estimated the zinc and copper content of tissues of
Chinese subjects by a colorometric technique. Using emission spectroscopy,
Tipton and Cook estimated the concentration of 24 trace elements, including
zinc, in normal human tissues from 150 adults who died quickly as a result of a,n
accident (Table 7-3) . Many of these values subsequently were confirmed by atomic
1451
absorption spectrophotometry. Although limited sampling made direct comparisons
difficult, zinc in tissues of Mideastern and Chinese subjects was higher than in
1621
their counterparts in the United States and Africa.
Forssen carried out similar studies in Finnish and British people who
died accidentally or by violence. ^ His results, which are set forth in
Table 7-4, are generally similar to Tipton and Cook's, whose findings in American
subjects are provided in Table 7-3. No differences in zinc concentrations
were observed between men or women and little difference was noted with age,
except that zinc levels in the ovary appeared to decrease with age in healthy
.
British women. Zinc in prostate is also known to increase with age, usually
in association with prostatic hyper trophy. 1451
Highest concentrations of zinc were found in human prostate and hair, followed
by muscle and liver, and kidney and pancreas; the rest of the metal was distributed
199
-------
TABLE 7-3
Concentrations of Zinc in Normal Tissue of Adult Americans
a
Organ, or Tissue
Adrenal
Aorta
Brain
Diaphragm
Esophagus
Heart
Intestine, duodenum
Kidney
Larynx
Liver
Lung
Muscle
Ovary
Pancreas
Prostate
Spleen
Skin
Stomach
Testis
Thyroid
Urinary bladder
Uterus
Number
of
Samples
15
104
129
91
67
140
68
145
48
150
141
137
16
138
50
142
21
130
71
21
112
32
jjg Zinc/g Tissue
Median
1600
1800
760
4600
2600
2700
2400
4500
1200
3500
1300
4300
1700
2300
7500
L300
780
2400
1400
2700
3000
1900
Ash
80%
Range
1300-2000
1100-2700
520-1100
3300-7200
1900-4500
2000-3700
1700-3400
3200-7400
680-2300
2200-5900
880-1800
2900-6800
840-2700
1500-3500
3200-19,000
1000-1900
550-1000
1700-3400
900-2200
1700-4000
1800-4900
1300-3400
ug Zinc/g
Median
17.6
68.4
46.36
156.4
98.8
108
93.6
207
111.6
129.5
62.4
180.6
74.8
80.5
337.5
65
12.26
76.8
79.8
99.9
69
81.7
Dry Weight
80%
Range
14.3-22
41.8-102.6
31.72-67.1
112.2-244.8
72.2-171
80-148
66.3-132.0
147.2-340.4
63.24-213.9
81.4-218.3
42.24-86.4
121.3-285.6
36.96-118.8
52.5-122.5
144-855
50-95
9.35-17
54.4-108.8
51.3-125.4
62.9-148
41.4-127.7
55.9-146.2
°Data from Tipton and Cook.1620
-------
TABLE 7-4
Concentrations of Zinc in Normal Tissue of Adults in
Brain
Parietal lobe (grey)
Parietal lobe Cwhite)
Hypothalamus
Cardiovascular System
Aorta
Myocardium
Vena cava
Respiratory System
Larynx
Trachea
Lung (upper lobe)
Lung (middle lobe)
Lung CLower lobe)
Digestive System
Esophagus
Stomach
Duodenum
Jejunum
Ileum
Cecum
Sigmoid colon
Rectum
Pancreas
Liver
Gall bladder
Urinary System
Kidney
Bladder
Reproductive System
Testes
Prostate
Ovary
Uterus
Vagina
Breast
Endocrine System
Adrenal
Thyroid
Lymphomyeloid Tissue
Spleen
Lymph nodes
Thymus
Bone marrow
Musculoske.letal System
Skeletal muscle
Articular cartilage (knee)
Costal cartilage
Bone (rib)
Integument and Fat
Skin (midventral)
Fat (under midventral skin)
Hair
Ash 7. of
Median
0.093
0.052
0.066
0.146
0.256
0.174
0.044
0.056
0.094
0.093
0.110
0.220
0.186
0.210
0.220
0.220
0.189
0.241
0.224
0.307
0.552
0.097
0.306
0.250
0.118
0.908
0.103
0.271
0.118
0.030
0.180
0.245
0.144
0.064
0.097
0.017
0.575
0.016
0.014
0.015
0.073
0.132
0.784
Dry Weight
Ran^e
0.040
0.034
0.043
0.031
0.151
0.069
0.015
0.023
0.036
0.017
0.012
0.022
0.068
0.124
0.157
0.139
0.094
0.140
0.144
0.133
0.254
0.030
0.118
0.132
0.082
0.303
0.066
0.066
0.072
0.022
0.130
0.062
0.068
0.015
0.081
0.012
0.355
0.005
0.011
0.011
0.015
0.061
0.080
0.131
- 0.076
- 0.131
- 0.255
- 0.435
- 0.255
- 0.123
- 0.103
- 0.188
- 0.216
- 0.152
- 0.300
0.362
- 0.346
- 0.316
- 0.318
- 0.253
- 0.315
- 0.339
- 0.780
- 1.900
- 0.186
- 0.560
- 0.942
- 0.159
- 2.870
- 0.139
- 0.298
- 0.159
- 0.038
- 0.210
- 0.513
i 0.211
- 0.123
- 0.108
- 0.040
- 0.930
- 0.051
- 0.040
- 0.028
- 0.117
- 0.174
- 3.600
Number of
.. Samples_
20
20
16
18
20
14
19
18
19
19
19
18
18
18
18
17
18
18
18
19
20
12
19
15
12
12
4
5
5
4
16
17
20
6
7
20
19
19
19
20
13
8
6
^Adapted from Forssen.
201
-------
491 the
rather uniformly through several tissues. In the endocrine system,/ zinc
content of the thyroid is greater than that of the adrenal, testes, or
491
ovary. Discrepancies among zinc values in different studies differ primarily
in the zinc content of muscle, which varies considerably.
Differences in measurements of zinc content of the same tissues and organs
commonly occur, and result in part from the diversity of methods used (emission
•spectroscopy, atomic absorption spectrophotometry, X-ray fluorescence, etc.) to
analyze these tissues. Results have been expressed as jig zinc/g tissue ash,
ug zinc/g dry wt,^ug zinc/g wet wt, and ug/zinc/g nitrogen. Because soft
tissue loses water easily during sample preparation, results calculated on a
wet weight basis must be evaluated carefully. Losses and contamination during
preparation for analysis can also make it difficult to specify the zinc level in a given
tissue with certainty. In addition, the source of the human tissues analyzed is
factors
dependent upon such diverse / as age, sex, physical and clinical well being
at the time of demise, drugs used, and etiology and timing of death. For example,
tissues analyzed from chronically ill patients had more variable metal levels
than were found in patients who died suddenly and unexpectedly.491,1232 yari-
also
ability / was reported in tissues according to geographic locations. These
data are presented in Table 7-5.
Tissues of rats, ' a horses, and other animals have been analyzed
for zinc.
/ Zinc levels in several tissues of ruminants, rats, and humans are compared
in Table 7-6.
SPECIFIC ORGAN SYSTEMS
The Reproductive System
Human semen is unique in its h&gh content of about 10-35 mg zinc/100 dl.
A positive correlation seems to exist between number of spermatozoa and the
202
-------
TABLE 7-5
Concentrations of Zinc in Samples of Sine
Human Organs Reported from Diverse Geographic Areasc
Orean
Aorta
Brain
Kidney
Liver
Lung
upper lobe
middle lobe
lower lobe
Myocardium
Pancreas
Spleen.
Testes
Organ
Adrenal
Aorta
Brain
gray matter
white matter
Breast
Esophagus
Intestine
duodenum
jejunum
ileum
sigmoid colon
rectum
Kidney
Larynx
Liver
Lung
upper lobe
middle lobe
lower lobe
Muscle (skeletal)
Myocardium
Pancreas
Prostate
Skin
Spleen
Stomach
Test is
Ovary
Thymus
Thyroid
Trachea
Ur. bladder
Uterus
Finland* U.S. A?
0.1466 0.180
0.093 0.074
0.306 0.450
0.552 0.340
0.094 ]
0.093 \ 0.130
0.110 J
0.256 0.250
0.307 0.220
0.144 0.130
0.118 0.130
Finland0
0.180
0.146
0.093 I
0.052 J
0.030
0.220
0.210 )
0.119 (
0.220 }
0.241 1
0.224^
0.306
0.044
0.552
0.094 1
0.093 \
0.110J
0.575
0.256
0.307
0.908
0.073
0.144
0.186
0.118 I
0.103J
0.097
0.245
0.056
0.250
0.271
Area Sampled
Africa** Near East
0.150 0.200
0.068 0.078
0.310 0.330
0.400 0.380
0.120 0.160
0.200 0.220
0.230 0.190
0.130 0.150
0.190 0.170
Indiad
0.290
0.103
0.102
0.086
—
0.227
0.310
0.120
0.331
0.219
0.128
0.107
0.200
0.126
0.209
0.100
0.094
0.220
0.103
0.206
"™
Far East**
0.250
0.092
0.600
0.720
0.160
0.280
0.280
0.170
0.160
U.S.A.6
0.200
0.180
0.076
— *™
0.260
0.240 )
0.220 /
0.260 V
0.260
0.330J
0.450
0.120
0.350
0.130
0.430
0.270
0.230
0.750
0.078
0.130
0.240
{0.140
0.170
0.270
0.089
0.300
0.190
f Adapted from Forssen. 91
'' Median values given as^Z in tissue ash.
^ Original study by Forssen.491 i522b
" Original study by Soman et al.
e Original study by Tipton et al.
203
-------
TABLE 7-6
Zinc Concentrations in Human and Mammalian Organs'2
Organ
Zinc,
in ash
Adrenal (sheep)
Aorta (human)
Bone (calf)
Bone (goat)
Bone (rat)
Bone (human rib)
Heart (calf)
Heart (goat)
Heart (sheep)
Heart (human)
Kidney (calf)
Kidney (goat)
Kidney (sheep)
Kidney (human)
Liver (calf)
Liver (cattle)
Liver (goat)
Liver (sheep)
Liver (human)
Liver (human)
Lung (human)
Muscle (calf)
Muscle (goat)
Muscle (sheep)
Mus cle (human)
Pancreas (cattle)
Fancreas (sheep)
Spleen (calf)
Spleen (goat)
Spleen (sheep)
Skin (human)
epidermis
dermis
Testicle (calf)
Testicle (goat)
0.520
0.228
0.024
0.020
0.036
0*026-0.037
0.187
0.195
0.136
0.117-0.280
0.146
0.170
0.182
0.511-0.517
0.300
0.01-0.1
0.248
0.346
0.238-0.690
0-1.78
0.23
0.206
0.360
0.200
0.900-1.270
0.01-0.03
0.225
0.145
0.163
0.178
0.580
0.087
0.013
0.016
a
Data
from Forssen.*91
204
-------
,. . 1/1 iiai been considered
level of zinc in semen. -^1,1191 zlnc in gpem hag / an index of
male fertility, H" although this concept is far from proved. Zinc is sperm is
removable by incubation with chelators,739 and it may retard the oxidation of
sulfhydryl groups normally present in sperm. ^24 -phe zinc in human semen
973a
is approximately 8-10 times as great as the zinc in uterine cervical mucus.
Zinc concentration may vary within the normal human prostate gland,
although it contains a higher zinc content than any other soft tissue in the
109,419 491 937 993
human body. ' ' * ' It is of interest that the ventral prostate of
rat contains little zinc compared to the posterior portion. * These proportions
may relate to histologic and physiologic differences observed between these areas.
although
The function of prostatic zinc is unknown, /it may supply zinc to spermatozoa.
On a dry weight basis, prostatic fluid has been reported to be 2-5 times richer
964
in zinc than spermatozoa. The release of zinc has been associated with the
activiation of starfish spermatozoa.514*1075 Cytochemical evidence revealing the
has been used to
influence of zinc in mitosis / explain its presence. ^ Some investigators
143
have tried to correlate zinc in seminal fluid with a zinc-containing protein.
Later studies indicated that zinc could be partitioned between high and low
molecular weight ligands, although a specific protein could not be identified .
Studies of boar semen have been repeated in the dog; the zinc in epididymal
1 OQO
sperm was loosely bound in both cases. However, zinc in rat sperm is less
1393
available for ethylenediaminetetraacetic aci4 (EDTA) extraction. In women,
adding zinc to the cervipal mucus in the uterus inhibits sperm penetration although
copper is a more effective contraceptive agent.
Zinc levels in endometrium are lowest in the late proliferative phase
(days 10-12 of the menstrual cycle); they gradually increase thereafter, reaching
205
-------
SQfi SQ7
their highest levels during the late secretory phase (days 25-27). ' The
zinc concentration of cervical mucus of normal menstruating females has also
been studied. In menstruating women, zinc in dry cervical mucus varied between
14 and 939 yg/g.866b The highest zinc values were found before or just after
ovulation (794-939 wg/g dry mucus). A cyclic pattern of zinc concentration
was noted in women, 66a,871a an(j an observable midcycle peak was measured
in the monkey. a Sharp peaks in zinc in cervical mucus unrelated to menses
were thought to come from intercourse, since an average ejaculate contains
about 14 mg zinc/100 dl, or about 8-10 times as much zinc as does cervical mucus.
Kidney
Few data are available on the levels of zinc in kidney other than
those reported in studies on tissues in general. In rat kidney, zinc in the
cortex was found to be five times higher than in the medulla, 1^83 with the
highest concentrations occurring in cytoplasm of the cells of the juxtaglomerular
apparatus, 1482a ^& ciliary edge of the proximal part of the nephron, the
ascending part of Henle's loop, and the cytoplasm of the cells in the distal part
of the nephron. After unilateral nephrectomy, zinc rises over the first few
postoperative days, but after the compensatory hypertrophy occurs the level
reverts to normal. ^' In rats with alloxan diabetes, renal zinc concentration
decreased markedly, 1482a ^eTea8 z±nc levels were observed to return toward
normal after insulin administration. A correlation between zinc content and
alkaline phosphatase activity was reported for the renal tubules.
Sensory Systems
The concentration of zinc in the human eye is high, although not as
964
high as in the prostate. Human eye tissues may not be as richly endowed
with zinc as are some animal eye tissues.1751'1753'1754 The presence of zinc
206
-------
in the eye suggests some- physiologic role for this metal in the eye or in
1752
the visual process. Indeed, studies with metas table zinc-69 suggest that
zinc may be important in uveal and retinal metabolism,1177 and zinc in the
human eye may be related to carbonic 'anhydrase or retinal reductase activity.886
The lens is rich in carbonic anhydrase,66 which may account for the high level
of zinc in this tissue. Rats treated with N-methyl-N-nitrosourea
developed retinal atrophy and cataracts associated with increases in
877
zinc content of the eye. A zinc-containing protein has been isolated from
332
the tapetum lucidum of the cat, and the zinc concentration is among the highest
found in cat tissues. Zinc has been found in various fluids of the eye, further
implicating this metal in some aspect of the visual process.
Zinc has also been found in or near the taste bud681 and in saliva.
Its absence from the saliva is associated with loss of taste acuity and the
appearance of pathologic changes in taste buds. Zinc in saliva has been
with
found in gustin, a metalloprotein that may be involved /providing nutrition to
the taste buds. In addition, zinc in saliva has been claimed to reflect zinc
678
nutrition in the body. Because this fluid is easily obtainable, its measure-
ment may be useful in determining some aspects of zinc status in the body.
Brain
Zinc is uniformly present in the human brain. It is the fourth most
prevalent element in the brain, preceded only by sodium, potassium, and
333a, 344, 393, 1424
magnesium. Twice as much zinc is present in the brain as copper.
The gray matter of human brain tissue contains about twice the amount of zinc
AQl
found in the white matter. In 21 brain samples, gray and white matter had
257
78 and 33.7 yg zinc/g dry weight respectively. Recently, brain regions
in experimental animals627*11^3 and humans *1>738 were analyzed for zinc content;
zinc was found primarily in the hippocampus, hypothalamus , and mossy fibers
207
-------
of the cerebellum. Zinc in the cortex was greater than that in the medulla. ^
Loss of zinc from the brain has btten associated with several neurologic symptoms.
Zinc has been found in the basal ganglia " as well as in various pituitary
fractions. ° Radioactive zinc accumulates in high concentration in steer
, , 456
pituitary.
Heart
In isolated heart mitochondria zinc alone or with £-chloromercuribenzene
sulfonate increased the accumulation of magnesium; it was suggested that zinc
T67
altered the permeability of the mitochondrial membrane. Zinc also initiated a
vigorous uptake of potassium. Addition of zinc under carefully defined experi-
mental conditions was sufficient to induce the transport of potassium by heart
....... 163,164
mit ochondr ia .
Lung
Little is known about the concentration of zinc in the lung. Few functional
studies have been performed concerning changes in zinc levels during physiologic
or pathologic changes in pulmonary parenchyma. One report notes that during
the acute phase of tuberculosis, zinc levels in lung decrease. (In patients with
chronic forms of the disease, zinc levels increase in lung parenchyma. )
Zinc also accumulates in the damaged pleura of patients with acute tuberculosis.
Pancreas
Zinc is found mainly in the islets of Langerhans in the cytoplasm of the
a-and 0-cells. In other parts of the pancreas, the concentration of zinc is
very low. Small doses of alloxan have been shown to increase the zinc
content in the pancreas, whereas high doses made zinc disappear from the 3-cells; in
Thus
the a-cells zinc content increased. /zinc may pl&y a role in the function of
208
-------
cells that elaborate insulin and secrete glucagon.1482 Although insulin is
biologically active when zinc is absent, the metal has been shown to prolong
the physiologic action of insulin and adrenocorticotropic hormone (ACTH)
•
preparations, apparently by retarding their rate of absorption.1274 The relationships
between zinc and insulin, and glucagon are thought to be similar. The role
of zinc in ACTH action has not been clearly characterized.
Liver
Very few studies of zinc content in liver of normal humans have been carried
out except those in which zinc levels in several tissues were studied.
Zinc levels in liver have been studied most often in patients with hepatic
cirrhosis, in which condition zinc levels are consistently lower than in normal
149a,1680a,1680b
subjects. From studies of viral hepatitis, it has been found
that the production of macromolecular zinc ligands synthesized in the liver is
683
reduced, and the concentration of amino acids found in blood is increased.
This shift in equilibrium allows zinc bound to micromolecular ligands to increase,
683,1420a
with subsequent hyperzincuria and total body zinc loss.
Zinc also has been studied in the liver of patients with various types of
carcinomas.8^3* Mean normal adult values of liver zinc are 279 yg/g dry
weight; zinc levels in patients with cancer ranged from 147-421 yg/g dry weight.
The meaning of such a wide range is unclear, although Addink has suggested some
relation between high levels of hematic and hepatic zinc. Olson et al.
reported low levels of zinc in patients with hepatic tumors. Zinc in
i
normal liver was calculated to be 38 yg/g wet weight, zinc in the uninvolved
portion of carcinomatous liver was 80 yg/g wet weight, and the liver tumors
contained 18 yg/g wet weight.
209
-------
Muscle^
The concentration of zinc in muscle represents the largest zinc
pool in man. Zinc levels reported in muscle have been quite variable. ' *
1451,1620,1621 Only a small fraction of the total zinc in muscle is free.
The role of zinc in muscle contraction has been investigated but its action
is unclear. Zinc binding to glycerol-extracted muscle exerts a strong relaxing
effect, probably through interaction with sulfhydryl groups. In small con-
centrations, zinc produces a marked potentiation of the muscle twitch. At
higher concentrations^ zinc reversibly blocks transmission at the neuromuscular
1402a,1402b
junction. The amplitude of the contraction of skeletal muscle was
785
reduced in the presence of 0.5 mM zinc and increased with 0.01 mM. The
1122
amplitude of cardiac muscle contraction was not enhanced by zinc, sug-
gesting that the role of zinc is different in skeletal and cardiac muscles.
Hair
Analyses of zinc in hair under some conditions can correlate with the zinc
level of serum. Because hair is readily available, it has been commonly used
as an index of tissue zinc. However, hair represents "history" as its growth
is reflected in its length from the hair shaft. Awareness of this phenomenon
is necessary to evaluate any changes in zinc levels along the hair shaft. Zinc
1704
fluctuates greatly close to the scalp. Hair zinc varies in hair of different
colors—the highest amounts are in black hair and the lowest in blonde hair—
723,1450 apparentiy related to the high level of zinc in hair melanosomes and
melanoproteins. ' Zinc may be active in melanogenesis.
Harrison et al. measured the zinc content in hair of 122 normal individuals.
The mean dry weight was 178.7 ug/g, the median 171.3 ug/g,
642
and the range was 81-314 Vg/g* These values agree reasonably well with other
. 411
measurements.
It has been claimed that hair zinc levels reflect body zinc levels, but by
themselves they are not particularly good indices. Rats fed zinc-deficient diets
1 ^27
exhibit lower than normal zinc levels in hair. Similar low levels of zinc
210
-------
in hair have been associated with zinc abnormalities in man,617'1557'1558 but
such relationships are more firmly established in animals than in humans.855
Hair zinc levels are particularly inaccurate in reflecting zinc stores in
prepubescent children. " Lower than normal levels of hair zinc have also been
found in pregnant and lactating women.620 Hair zinc varies systematically from
infancy through the first four years of life615 and demonstrates changes that
generally follow those of serum. At birth, hair zinc levels are similar to
those found in adults, but they decrease over the next year of life. Adult
levels are not attained again until four years of age. Fluctuations in hair
zinc are illustrated in Figure 7-3.
New methods for evaluating the composition of hair take into account the
incorporation of metals into the hair matrix during mitosis. Metals also may
and incorporated into the
be adsorbed onto the hair surface or utilized metabolically/ structure of the
hair shaft. Thus zinc concentration in the cold- and hot-water soluble and
Insoluble fractions can be determined.
Skin
whole
Although/ skin does not contain a high concentration of zinc (ranging from 20-1,000
1690
yg/g dry tissue), '5>1 its importance in relationship to
the physiologic function of this surface barrier has been emphasized. a> *
307c,1075a,1232b ^Laa^s in z±nc concentration in relation to several dermatologic
disease states in man have been investigated by biopsy techniques and neutron
activation analysis. Despite the marked variability inherent in tissue sampling,
handling ,
/and analysis, lower zinc levels in skin of patients with scleroderma, basal cell
O C / J
carcinoma, and various forms of skin cancer were found.
Application of these findings to clinical dermatology has not been continued
211
-------
180
I160
o.
| OH
ut
^100
o
u
O 80
5 60
ul
5
40
0 0.25-1 1-2 2-3 3-4 4-5
(25> cm cm «ji ut> mi
S-7
(141 •
7-K>
137)
AGE-YEARS
MJ-t3
120
13-17
1111
17-40
tin
FIGURE 7-3 Mean concentrations or zinc in hair of
subjects from infancy to age 40. SEM indicated on
graph; number of subjects in each age group is given
in parentheses. Reproduced from Hambidge et al. ^'
212
-------
enthusiastically, although the role of metals, particularly copper and zinc,
in collagen formation has been investigated in vitro. 729a The effect of zinc
on healing surgical wounds is discussed in Chapter 10.
Bones, Tendon, Nails, and Teeth
Zinc is always found in bones and represents a major body pool of
Zinc.17»651»909»1399 Although the function of zinc in bone is not clear, it
may aid in calcification as an organic constituent of bone or as an activator of
the calcification process itself. Its ability to exchange with bone calcium
1399,1399a
in vitro is well known. Even when bone is demineralized, zinc is found
91,1527
in small but constant amounts in the insoluble organic fraction of bone
t o o ^ RO7
and tendon. * Thus, zinc could be bound in some manner to the collagen
1 527
matrix and not merely present in the interstitial fluids. Indeed, Hsu
729
has postulated a specific role for zinc in collagen metabolism, perhaps in
the cross linking of the collagen matrix.
1437
Zinc is present in human rib cartilage. Concentrations vary with
age; it is lowest under 2.5 years of age and after that it rises. Dwarfs
did not show any significant differences from normal subjects.
In teeth, zinc concentration is greatest in the top layer of the enamel,
and it decreases in the underlying layers. The deposition of zinc in the tooth
enamel probably takes place at the same time fluoride is laid down. Analyses
of human enamel showed that zinc was present in all enamel samples but varied
1144
widely, ranging from 58-1,550 ppm.
Pregnancy and Fertility
Changes in zinc metabolism during pregnancy have been studied extensively
and reviewed in other portions of this phapter. However, several
213
-------
investigators have attempted to assess the relationship between zinc metabolism
and pregnancy by measuring changes in blood and/or hair zinc during pregnancy.
Many changes noted in zinc metabolism during pregnancy are the result of the
changing hormonal state, and the shifts in zinc among the various body pools
1 primarily
are related /to the complex interaction between maternal, placental, and perhaps
fetal secretion of several hormones. The dynamics of hormonal interactions have
not been fully characterized so that many of the changes in zinc metabolism
must still be described as observations determined over time* Changes in blood
zinc during pregnancy have been described,613.«-6»669,676,797,1164a as have
changes in hair * and blood zinc in the fetus and in the amniotic fluid.669'676
That zinc is retained during pregnancy primarily in the placenta has been hypothesized
by Sandstead,1404 based upon data collected by others.767»1770a
A zinc coating has been used either alone or with copper as an effective
agent in the control of reproduction following the insertion.of a wire, loop,
or T-bar device into the cervix in rabbits and rats.1021*1022 Although the
specific role of zinc and/or copper in this process has not been characterized
as a chemical event, Insertion of zinc and copper wires into the uterus of pregnant
and estrous rabbits appeaxad to increase uterine motility, thereby affecting fertili-
1021
ty. This may be associated with the release of metal ions from the impregnating
1022 1601a,1821b
wire. Zinc and copper also may Interfere with Implantation of the bias tula.
Of further interest is that following the insertion of a copper-T device into the cervix
1163a
in 16 women there was a much greater increase in endometrlal zinc than copper.
Changes with. Age
Human embryos and fetuses. Zinc has been found in first trimester fetuses,
260
and Increases sevenfold between the thirty-first and thirty-fifth day of gestation.
214
-------
Livers from younger fetuses contain more zinc than do livers from older ones
and all fetal values are proportionately higher than adult levels.
Plasma zinc in human fetuses has been reported to be three times that
QC
of adults. Human amniotic fluid also contains zinc; the level may
fluctuate' during pregnancy.5713
Zinc-65 rapidly and easily crosses the placenta and is taken up by fe-
tuses of several species,455'580'629*630'1605 the ease apparently increasing as
gestation age increases. Some evidence suggests that this increased uptake
is dependent upon the fetal liver.1605 Retrograde transfer of zinc from fetus
1605
to mother has also been demonstrated. Zinc-65 has been found in /even
when the only source is the male sire, ultimately from the spermatozoa in
580
which zinc-65 has equilibrated with naturally occurring zinc.
Infancy and childhood. High (>200 yg/g wet tissue) concentrations of zinc
813
have been found in the liver of newborns, which persist until about the fourth
month of life. Then levels fall, and the lowest values are reached at age 10;
813
after age 10, hepatic zinc increases gradually until adult values are attained.
The zinc content of many brain regions of newboms was lower than that found
641
in adult brains. Zinc contents of infant organs are shown in Table 7-7.
Table 7-8 lists zinc values for some body fluids of children at different
zinc
ages. Whole blood/levels of a newborn child are about 29% the amount found in
852
whole blood of a healthy adult. One reason for the small amount may be
discovered in the lower content of zinc and carbonic anhydrase in the red blood
i
cells of the newborn. By the end of the first year of life, the zinc level
957
doubles, and it reaches normal adult values by the second year of life.
Zinc levels in blood and hair of infants and children with several physiologic
and pathologic conditions have been reviewed recently.
215
-------
TABLE 7-7
Concentration of Zinc in Tissues of Infants6
Organ or tissue
Liver
Kidney
Spleen
Pancreas
Thymus
Brain
Adrenal gland
Age
4 h
6 h
13 d
2 yr
2 yr
6 h
13 d
6 h.
13 d
2 yr
6 h.
6 h
6 h
6 h
Sex
Female
Female
Male
Male
Male
Female
Male
Female
Male
Male
Female
Female
Female
Female
Zinc, ^ig/g wet
weight of tissue
228
105
172
80
33
28
21
15
12
24
58
16.7
7.2
10.7
1210
from Parr and Taylor.
216
-------
TABLE 7-8
Zinc Content of Body Fluids in Children
Number of
Authors
970
Mahanand and Houck
Hellwege660
'
Henkin*69
682
Henkin et al.
852
Kleiman et al.
Children
43
13
8
10
9
15
22
16
15
10
76
26
Age of Children
Blood serum (S) or plasma (F), yg
1-7 yr
Abortions
Newborn
1-12 mo
1-2.5 yr
2.5-5 yr
5-12 yr
Over 12 yr
Newborn
21 months
Whole blood, yg zinc/100 ml
Full term newborn
Premature infants
Mean value,
SD
zinc/ 100
108 +
115 +
111 -»-
111 ••-
112 +
111 +
114 t-
109 ±
83 +
90 i
177 ±
147 +
ml
15
22
16
14
15
17
18
11
11
16
6
7
Range
74-120
70-160
78-144
83-139
82-142
77-145
78-150
86-142
70-100
Mahanand and Houck
970
Urine, yg zinc/100 ml
1-7 yr
42
33-55
-------
Changes in organ systems. Changes In zinc with age in several organs have
been systematically studied. A staple way to convey this information is through
the presentation of figur.ee in which changes in concentrations of cine in the
kidney (Figure 7-4), aorta, brain, and heart (Figure 7-5), liver and lung
(Figure 7-6), and pancreas and spleen (Figure 7-7) a*« charted.
ZINC ABSORPTION
Routes
Zinc is absorbed across several physiologically active membranes, including
the gut mucosa, alveolocapillary membrane, and tissue and organ membranes.
Absorption across the gut mucosa occurs after oral administration. Absorption
across the tissue and organ membranes normally follows gastrointestinal absorption
or parenteral administration. Zinc most commonly enters the body through ingestion
of food and drink. Inhalation of the metal and its absorption across the alveoloctp-
illary membrane present a specialised type of absorption, because the amount of cine
available via this route is usually small except among sine workers where metal fums
fever or other forms of zinc inhalation may cause clinical problems.* Absorption
across tissue and organ membranes is a natural consequence of all forms of
presentation. In addition, zinc may be absorbed across the unbroken epithelial
membrane of the skin as well as across the broken epithelial membrane after
burns or wounds.
Mechanisms
Orally administered zinc is absorbed at several loci in the gastrointestinal
tract. Earliest evidence of absorption appears in the stomach as soon as 15
min after ingestion. Although the major site of zinc absorption appears to be
*See Chapter 11.
218
-------
pom
o»h
, 7000
6000
5000
4000
3000
2000
1000
0
II
X
S * -S^ .Zn
••* ' *^.9 Jo--i9 20-29 30-39 40-49 50-59 60-69 70-79 «0->
Agt In Vtor»
FIGURE 7-4 Mean concentrations of zinc in the
kidney in 221 subjects from the United
States. Reproduced from Schroeder
219
-------
ash
4000
3000^
" ''•'.. ..••' *'•. (war!
\ •-,.,.-
\
\
\ s,--2-^
2000 \ , -•-^!
\ X » "^"^ o
V-'- °""^-^ o
0 "** •• aorta
1000-"^--^ ^^ ^ I.._«....jr.-."
'i~ !£, 1"9 1(W9 20"29 30~39 A0"49 50"59 60"69 70"79 fl°*
" _ ***" Ag« in Years
FIGURE 7-5 Mean concentrations of zinc from 121 aortas, 149
brains, and 180 hearts of subjects from the Di-i';
States. The early decline of zinc in the aorta
was the result of measurements from two subjects.
1 / d
Reproduced from Schroeder et a_l»
220
-------
PP"».
ash I
«
6000
5000
4000
3000
2000
1000
0
f\
\
\
\
\
\
s"'
e
N,
\
'N*
•
•
" "*~"^
.lung
M*. CHO M 10.19 20.29 30.35
Age in V*ar*
FIGURE 7^-6 Mean concentrations of zinc in 231 livers and
188 lungs of subjects from the United States.
The low zinc levels in six older infants may
reflect the diseases of which they died. Re-
produced from Schroede,i; .et al.
221
-------
ppm
o»n
3000
2000
1000
'poncraos
\ • •
\
\
\ ^-o o o o -o r spleen
„o
V^9 10-19 20^29 30-39 40-49 50-59 60-69 70-79 80->
Age In Years
FIGUKE 7-7 Zinc In pancreas and spleen in 167 pancreases
and 183 spleens of subjects from the United
States. Specimens of pancreas from infants
were not available. Reproduced from Schroeder e)t al.
222
-------
in the second portion of the duodenum, absorption appears to occur at other
portions of the small and large intestine.6723'1035'1036
The specific manner by which zinc is transported from gut lumen across
the gut mucosa or from the mucosa across the gut serosa is not known. However,
a consideration of the molecular characteristics of zinc would indicate that
987
the formation of a tetrahedral quadradentate ligand with a small organic
molecule is the preferred absorption complex. Our knowledge of this process
is limited. The initial event in absorption probably is the formation
of a low molecular weight organic zinc chelate; the presence of a zinc-protein
complex in the gut has been suggested by several investigators. '*
The soluble organozinc complex may be passively absorbed across the plasma
1398
membrane of the mucosa of the intestinal villi following first-order kinetics.
Although not established in gut mucosa, this type of kinetics has been suggested
in studies of zinc uptake into fish liver slices, where metabolic inhibitors
such as dinitrophenol, potassium cyanide, or iodoacetic acid
did not influence transport.1328 gome workers have questioned this
passive first-order kinetic process, ^28 jjut their data—in which an active
process is suggested—are not entirely convincing in view of the informa-
tion supporting the other opinion. The remainder of the ingested zinc forms an
insoluble, nondiffusible complex transported and carried in the intestinal
products and excreted in the feces. Fecal zinc is composed in large part of
all the unabsorbed zinc. However, some absorbed zinc is also found in feces;
this amount includes zinc excreted in the bile, in the pancreatic fluids found
in the second portion of the duodenum, and fluids composed of back diffusion
products derived from gut serosal to mucosal transport. In the rat the serosal
to mucosal zinc transport can account for as much as one-third of the total zinc
content of the feces. The feces of a man with an average zinc intake of 15 mg
daily contains between 10-14 mg zinc.
223
-------
Many agents affect zinc absorption positively or negatively at the initial
stage of absorption. Agents that inhibit formation of the low molecular weight
organic zinc complex and thereby decrease absorption include phytate (from
bread and other sources),1157'1328 and soy protein. Other agents that influence
absorption (but to a lesser extent due to a smaller oral intake) are peanut,
sesame,906 cottonseed, saf flower, and other meals,1141 other plant proteins,1517
sphalerites and franklinites,416 arginine,716 calcium,365'4893 particularly
691
when phosphate in high and phosphates, particularly when calcium is high,
other metals,1669 particularly cadmium,40 and food itself.3'822'1421 Oral intake
of exogenous zinc even 45 min after a light meal may decrease zinc
absorption.* In the rat, a lowered protein intake was associated with
decreased zinc absorption, but a low protein intake has been associated with
increased zinc uptake in man.
Other factors also influence the manner by which zinc is transported
across the intestinal mucosa. They include temperature (as temperature decreases,
1398
transport decreases), concentration of zinc in the external medium (as zinc
concentration increases, absorption increases), and efflux of zinc from the
tissue itself (e.g., loosely bound zinc will exchange with circulating zinc and
become mobilized and metabolized) . Concentration of zinc in tissue must be
taken into account because it varies inversely to the amount of absorbable
zinc. Other factors include the physicochemical characteristics of
154
zinc in relationship to the state of the mucosa, dietary components not
previously mentioned (for example, food additives or other substances that act
on zinc such as oxidizing or reducing compounds), alcohol, and the microbial
C.OQ
characteristics of the gut. The age of the subject may affect absorption.
^Substances which limit zinc availability by limiting solubility of the
zinc itself by complexation with the organozinc complex may be species-
dependent.
224
-------
Adding EDTA and other synthetic chelators to food counteracts the negative
effects of phytate and calcium365'691'741 to some extent,1508 and other naturally
occurring complexing agents produce similar effects.1157 In states of zinc
deficiency, rats and dogs exhibit impaired intestinal absorption of zinc, and
it may be considered an anomalous physiologic condition.
Clinical problems of gastrointestinal absorption, including malabsorptive
processes of several types, will decrease metal absorption. Zinc can be lost
during several disorders, such as excessive renal loss of trace metal because of
intrinsic kidney disease or changes in tubular or glomerular function; defective
metal binding in plasma of a congenital, acquired, or iatrogenic type; or probably
through Inborn errors of metabolism, since some aminoacidemias and aminoacidurias
1495,1496
are associated with excessive loss of metals.
A few agents appear to increase zinc absorption in animals by providing or
aiding in the fomation of the low molecular weight organic zinc complex; these
537,904,1141 875
include histidine, cysteine, and methionine; EDTA; and several
96
zinc-sugar complexes. Vitamin C also has been reported to
increase zinc absorption with the intake of food.882 Rats raised in a germ-free
environment appear to utilize zinc better than rats raised in an environment with
1509
normal bacterial flora. 1345c
When zinc-65 is administered orally, plasma levels peak 2-4 h later;
1368,1532,1534,1535,1537
about 50% of the total dose was found in the feces within 15
days after oral administration. The average net absorption of zinc-65 varied
between 30-70%, but Spencer observed approximately 50% absorption of the isotope.1532
She also noted that low zinc-65 levels in plasma reflected low absorption, where-
1532
as higher plasma levels reflected greater absorption. Although high calcium
intake decreased zinc absorption in rat, sheep, and other animals, this did
1538
not appear to be the case in humans. Dietary protein does influence zinc
absorption in humans. Subjects on a low protein intake had higher zinc-65
plasma levels and lower fecal zinc-65 levels than individuals on a normal
225
-------
Human
protein intake.1534'1535'1537 /subjects on a high protein intake exhibited the
expected converse results.
After intravenous administration of a single dose of zinc-65, human liver
11 ^7
has been shown to accumulate and retain zinc, perhaps because of metallothlonein
1S38
in liver. More than 2 mo after the dose, the liver contained about one-fourth
the level found the first day after administration. Zinc is also found in the
kidney, spleen, and intestinal mucosa, and zinc levels in lung are similar to
1532
those found in the gut. Pancreas, adrenals, and thyroid show relatively
high uptake of zinc-65, whereas prostate, the organ with the highest zinc
1532
concentration, is rather low in activity. In muscle, the turnover of zinc-65
is quite slow" the highest activity is found in cardiac muscle. Bone
exhibits low uptake but prolonged retention. Bile concentration of zinc-65 also
is low.1532
Differences in human blood, fecal, and tissue levels are a function of the
route of administration of zinc-65. Following oral zinc-65, the isotope may
be found in blood within 15-20 min with peak levels found within 2-4 h. After
oral administration, plasma or serum zinc-65 levels are higher than in whole
blood. Levels in red blood cells increased for the first 5 days after intravenous
injection of a single dose of zinc-65 and decreased thereafter; after 28 days
zinc-65 in red blood cells was 2-3 times higher than that in plasma. No matter
what the route of administration of the isotope was, most of it was found in
of administration.
the feces^although the amount differed according to route/ Following oral
administration of a single dose, 66% of the isotope was found in the feces within
the first 3 days. After 21 days, 70% of the isotope was found in the feces, 2%
in the urine, and 28% remained in the body. Most of the gastrointestinal
absorption occurred within 4 h of oral administration. These studies were carried
out in older adult males. Thus, little or no information about distribution
in young adults or male-female differences is available.
Zinc balance has been studied in only a few small groups of children.
In 36 7-10-yr-old girls in 3 different areas in the United States, daily
226
-------
437
zinc intake was found to be from 4.6-9.3 rag, with a mean of 6 mg. Protein
sources in food were of animal origin in 74% of the subjects. The average
fecal excretion was 3.9 mg/24 h, and urinary excretion was 274 + 69 yg/24 h.
Values for fecal zinc correlated with zinc intake. In 2-yr-old children given
1707
4.34 mg zinc, 53% of the metal was retained. In 8-9-yr-old children given
1707
10.9 mg zinc, 3.8% of the metal was retained.
Studies of oral absorption of zinc-65 in human subjects provide values
1345c
ranging from 25-90%. Absorption values found by Richmond et_ al. for four
people given zinc-65 chloride orally and calculated by the method of
la,672a 672a 515a
Aamodt e£ al. were 34.8, 47.5, 66.9, and 88.8%. Furchner and Richmond
also reported an effect of stable zinc intake on absorption of zinc-65 in one
1534
human subject. Spencer e£ al. reported absorption of orally administered
zinc-65 in five subjects. They reported values ranging from 27% to 43.9% with
an average of 37.5%. The zinc-65 chloride was given as a single, daily dose of
15-25 yCi with breakfast. The values obtained by Aamodt e£ al. indicate a vari-
ation of 19-100%. These values are not inconsistent with the values reported by
Richmond et^ al. and it is probable that binding of zinc-65 to food proteins could
1534
account for the lower values reported by Spencer et al. Comparison of the
1534 1345c
values from the studies of Spencer and Richmond et al. with those of
la,672a with taste and smell dysfunction
Aamodt e£ al. suggest that the patients/they studied fall into
three groups—a low absorption group with values between 19-37%, an intermediate
absorption group with values between 60-75%, and a high absorption group with
values close to 100%. Because only 11 patients were studied by Aamodt et_ al.,
the separation of absorption values into three groups was somewhat arbitrary, and
the positive results by T-test, comparing the high, low, and middle
group, only suggested that these groups were independent.
227
-------
The long-range component for the 11 patients observed by Aamodt et al. who
received oral zinc-65 ranged from 250 to 498 days, except for one patient with a
congenital loss of taste, who had very shortened half-times of 143 and 167 days
after oral and intravenous administrations, respectively. The combination of low
absorption and rapid loss of ingested zinc might be expected to lead to chronic
zinc deficiency unless supplemented by a high zinc diet or exogenous zinc.
Two patients studied following oral administration of zinc-65 were found to
have an unusual retention pattern characterized by a sharp decrease in the second
component of biologic retention. One patient's half-time changed from 288 to
la,672a
145 days and the other from 350 days to 177 days. The period of shortened
half-time corresponded with the period in which each of these patients had been
receiving exogenous stable zinc (100 mg/day). These results imply a washout effect
of orally administered, stable zinc. Some evidence for this type of effect
323
had been previously reported.
Retention of intravenously administered zinc-65 by normal human subjects
follows a two-component exponential pattern. The rapid component consists of
20% of the injected activity and is lost with a biologic half-time of 8 days.
The slow component includes the remaining 80% of the activity and is lost with
la 1534
a biologic half-time of 300 days. Spencer et £l. reported that two pa-
tients lost 25% of intravenously injected zinc-65 with an average half-time of
12.5 days and 75% of the injected activity with a half-time of 322 days. Newton
1137
and Holmes observed that following accidental inhalation of zinc-65 by a
human subject, 27% of the inhaled activity was retained with a half-time of 18
1345c
days and 73% was retained with a half-time of 453 days.* Richmond et al.
reported that the slow biologic retention component was 418 days for oral admin-
istration to four human subjects. Figure 7-8 compares retention of zinc-65 in
man and other species.
*In this study there was a greater uptake and retention of zinc-65 by soft tis-
sues, including liver, than by bone.
228
-------
1-0
O 10 20 X> 40 50 60 70 80 SO"1' ?(» ' WO
FIGURE 7-8 Retention of orally administered zinc-65 by four
mammalian species. Reproduced from Richmond et al.
1345c
229
-------
The retention pattern of a patient with cystinuria who took D-penicillamine
was different from that previously observed in normal
human subjects. This patient's zinc-65 retention after intravenous injection
required three exponential components, reflecting a very rapid initial loss:
21.9% of the activity was lost with a biologic half-time of 1.5 days, 17.6X with
la
a half-time of 15.9 days, and 60.5% with a half-time of 235 days. Increased
excretion of zinc-65 during penicillamine treatment has also been reported by Ekberg
direct
et. al • 2a and it is probable that the increased removal was a /effect of D-
penicillamine. Moreover, urinary excretion was substantially increased during
the first 6 days of the study. The ratio of urinary to fecal excretion was 4,2:1
compared to a ratio of 0.2:1 for 4 patients, 2 with Wilson's disease and 2 het-
672a
erozygous for Wilson's disease, neither or whom wore treated with D-penicillamine.
A urine to feces ratio on the order of 0.25:1 UOX of daily excretion in urine) developed
1137 1534
following inhalation of zinc-65. Spencer et_al. reported variable ratios for
urinary to fecal excretion from 0.75:1 to 4:1 in 8 terminal cancer partients, with an
average value of 0.49. The variations between urinary to fecal excretion ratios and the
limited amount of data available make it difficult to conclude that D-penicillamine
1534
increases urinary zinc excretion independent of its effect on zinc absorption.
After intravenous administration of rinc-65j activity decreased in whole blood
and plasma during the first 24 h, followed by increased activity in whole blood
and continued loss from plasma. Red blood cell activity increases during the
first 5-10 days after injection and then decreases at about the same rate as
the plasma thereafter. The biologic half-time is 26 days. Plasma activity
initially decreases with a half-time of 3.8 min and more than 95% of the activity
is removed by these short half-time components. The data for whole blood of
la,672a
normal subjects were resolved into four exponential components. Initially,
95% of the injected activity was removed with a half-time of 6.5 min, 2.5% of
the activity was lost with a half-time of 80 min, 1.5% of the activity was lost
230
-------
with a half-time of 2 days, and the remaining IX was lost with a half-time of
about 25 days. These values are consistent with data on whole blood reported for
*1277,1278
normal subjects and growth-retarded, hypogonadal Egyptian boys, and for
1488a
patients with neoplastic diseases. What is striking about the disappearance
of zinc-65 from plasma is the extremely rapid loss of nearly all of the activity
within the first few minutes after injection.
of
After intravenous administration/zinc-65, most of the isotope is rapidly
la,672a
taken up in liver. The rapid uptake phase, completed within the first 20 min,
is followed by a slower rate of uptake continuing for 5 h, with maximum activity
found between 5 and 24 h. Zinc-65 activity in liver then decreased slowly over
the next 100 days. Activity measured over the thigh decreased initially, prob-
ably reflecting lost activity in the blood; it then increased during the first
40-50 days after injection as the activity in the liver decreased. The apparent
uptake of zinc by the thigh area following release from the liver may imply a
and/or
biochemical conversion / release of the injected zinc to a form stored by
muscle or bone.
Although many studies of zinc absorption have been reported, a great deal
about the dynamics of zinc absorption 504,
of uncertainty/still exists. Zinc is still considered to be absorbed poorly
614,1652 92
and variably.
i
% •
Data from several zinc-65 absorption studies lead to the following conclusions:
1. Retention (R) of intravenously injected zinc-65 bound to plasma follows a
two-exponential component pattern in normal subjects; average retention
was:
-(0.693/6.36)t -(0.693/266.5)t
R - 16.2 e +83.8 e
*See Chapter 10.
231
-------
2. The cystinurln patient treated with D-penicillamine had a markedly dif-
ferent retention pattern, characterised by three exponential elements:
-(0.693/1.5)t -(0.693/15.9)t -(0.693/235)t
R - 21.9 e +17.6 e +60.5 e
The major effect of D-penicillamine was rapid removal of 22% of the
injected activity with a half-time of 1.5 daye. Th* similarity of the
remaining retention pattern to those reported by other investigators and
by studies of normal volunteers suggest that the effect of the drug was
to remove zinc-65 that otherwise would have been part of the long com-
ponent of retention.
1*.
3. Following Injection of zinc-65, activity in whole blood of all subjects
515a,672a,1345c,1534
decreased rapidly during the first 24 h, then In-
creased to a maximum value between 5 and 10 days after injection, after
which it decreased. Activity in plasma quickly decreased to less than
2% one day after injection and then more slowly for the rest of the study
period. Red blood cell activity increased for the first 5-10 days after
injection, then decreased at a rate similar to that of the plasma there-
given D-panlcillamine
after. Although the patient with cystinuria/always had less activity in
cells and plasma than the normal volunteers, no significant dif-
ferences were observed.
4. Measurements made over the liver area after intravenous zinc administra-
in
tion/patients with taste and smell dysfunction showed a pattern of rapid
uptake by the liver, followed by a much slower loss. Maximum liver uptake
reached 55-70X of the injected activity between 5 and 24 h after injec-
tion. Hepatic retention of injected zinc-65 was described by three
exponential components* Normal volunteers showed the following retention
pattern:
232
-------
-(0.693/0.60)t -(0.693/7.5)t -(0.693/70.0)t
R • 22 0 +48 c +30 e
The cyatinuria patient's pattern was vary different} the absolut« magni-
tude of tha loea from har livar was represented as:
-(0.693/2.65)t -(0.693/15.0)t -(0.693/250)t
R " 59.1 e +28.0 a +12.9 o
5. Activity over tha thigh initially dacreaaed, probably reflecting losa of
activity from tha blood. Activity incraaaed after injection for 40-50
daya. Thla pattern suggests that cine la taken up by bone or muscle
once it is released by the liver.
mean
6. The/long retention component for patients with taste and smell dysfunc-
tion receiving oral rinc-65 waa approximately 300 daya with a range of
254-498 daya.
Zinc Binding in Humana
Zinc complexes of low molecular weight found in serum are thought to be
transferred paaaivaly across tissue membranes and bound at various concentrations
in different tissues, according to the concentration of tiasue-binding cine pro-
teins at specific sites in tha tissues. Although metallothionein represents an
807,1304 806
important binding protein in kidney and liver, other specific tissue-
binding proteins may be present. Studies with zinc-65 suggest that liver, pancreas,
844,1715
and kidney retain high levels of this isotope within 96 h of injection,
but liver and bone have been shown to contain higher concentratlona of the isotope
ovar longer timee. Whether these results indicate different binding pro-
teins with different association constanta or different concentrations of binding
proteins with similar association constanta la unknown. Thia diatinction is
particularly obvious in the binding of copper to different tiasuaa in patients
with trichopo liodystrophy (Hankea'a disease) and my be important in normal and
pathologic atates of zinc metabolism as well.
233
-------
PHYSIOLOGIC FUNCTIONS OF ZINC
Zinc affects the function of several organ systems. Because zinc is an
integral component of DNA polymerase, any rapidly dividing cellular system has
an important requirement for zinc. However, several systems requiring zinc
metalloenzymes or metalloproteins have a requirement for zinc. In addition, some
systems are influenced by the presence of zinc in a manner not readily apparent
from their molecular characteristics. This section will attempt to outline the
role of zinc in each major system and specify how zinc acts in it.
Zinc directly affects the activities of carbohydrates, lipids, and proteins.
Yet in terms of nutritive factors, carbohydrates, lipids, and proteins can affect
zinc concentration and thereby change organ function and behavior. Although the
role of zinc as a cof actor in enzymes has been well studied,* that zinc is an activator
or transducer at cell membranes in intercellular or intracellular systems
285
only recently been suggested. Indeed, systematic studies of zinc effects
thwarted until reliable techniques were developed in which concentrations of
easily
zinc present in the system under study could be/ measured and used to assist in the
investigation of metabolic function. The discovery of zinc as a membrane stabilizer,
a participant in electron transfer processes, and a major participant in enzyme-
substrate interactions, all indicate the important role zinc plays in several
285
physiologic systems.
The Endocrine System
There are many zinc-hormone interactions in which zinc influences hormonal
activity at several levels of action, including synthesis, secretion, target- organ
binding, and function. Similarly, there are hormone-zinc interactions in which
hormones influence the absorption, distribution, and excretion of zinc, its trans-
port in blood, and its action at several levels of organization.
*See Chapter 9.
234
-------
Zinc-hormone interactions. Zinc has been shown to influence the endocrine
system at each of its major levels. Thus, increasing or decreasing the concen-
tration of zinc has been shown to influence hormonal secretion in the hypothalamic-
pituitary axis, the anterior pituitary-target gland axis, hormonal synthesis
within target glands, and activities at several peripheral organs and target tis-
sues, including hormone binding and utilization.
A surfeit or deficiency of zinc has been shown to influence the endocrine
system. Excess zinc will affect the endocrine system, but much less than will
1403a,1816c 9?2b 575b 518a,1206a 888a
lead, mercury, iron, cadmium, nickel, and
257a,790a
copper.
It is more common to find that zinc deficiency affects the endocrine system
adversely. It is noteworthy that zinc may influence the same or different
hormonal systems independently of the mechanism of how the deficiency was produced;
that is, from reduced dietary intake or greater body loss.
*
Other factors may influence dietary intake of zinc. For example,
zinc deficiencies in animals and humans have been reported to be associated
382a,1174,1277,1640,1642
with hypogonadism. This condition has been linked
with decreased testicular weight and size, decreased number of spermatozoa, and
a late-stage spermatogenic arrest, yet Leydig cells have remained intact. In rats
are depressed in 575b
with severe zinc deficiency, growth and development / the testes, epididymis,
423a,488a,1044b
accessory sex organs and pituitary. It has been reported that
1044b
complete testicular atrophy induced in zinc-depleted rats is irreversible,
whereas in other studies gonadotropin administration was associated with
gonadal maturation. Zinc supplements have been reported to reverse the signs of
the deficiency in all organs, but neither the testes nor the epididymis regain
1044b
their normal size, function or zinc concentration. Similar effects have been
*Zinc-deficient diets and their effects are discussed in Chapter 8.
235
-------
1054a 1053
reported for cattle and goats. However, there was little or no differ-
ence in the synthesis of testosterone or dehydroepiandrosterone following the in-
cubation of testicular tissue with hydrogen-3 cholesterol from zinc-deficient or
1272a
pair-fed control rats. These latter findings indicate that the mechanisms
involved in this process are not in the testis itself. These results are consistent
with those investigators who suggested that the effects of zinc deficiency on the
release of pituitary gonadotropins are the major phsiologic factors influencing the
1044c
changes observed. Indeed, lower levels of plasma and pituitary luteotrophic
hormone (LH) were found in zinc-deficient rats than in controls although levels of
544
hypothalamic LHRH were similar. Responsiveness to exogenously administered LH
and FSH was greater in zinc~deficient than in restricted-fed control rats although
a comparison of pituitary or plasma LH or FSH levels between these two groups of
rats was not reported.
The influence of zinc on pituitary function has been studied for many years. In
1939,
/zinc was shown to enhance pituitary activity when a crude extract from the
posterior pituitary was injected into frogs and the water-retentive property of
150
the extract was prolonged by adding zinc salts. The action of what might
presently be considered to be the pituitary follicle-stimulating hormone (FSH)
and luteinizing hormone (LH) was enhanced after zinc salts were added to anterior
pituitary extracts and then injected into sexually immature or hypophysectiomized
116,117,997a
rats. More recently, incubating zinc with bovine pituitary extracts
has been associated with increases in circulating growth hormone (GH), thyroid-
stimulating hormone (TSH), LH, FSH, and ACTH, although the amounts of zinc re-
quired to produce these effects were larger than would be necessary for copper or
888b
nickel to produce them.
236
-------
The specific role of zinc in insulin action has not been clearly defined.
When present in the insulin molecule, it is well known that it delays its phsio-
1459
logic action and prolongs hypoglycemic activity. Although native insulin
does not contain zinc and zinc-free insulin is fully potent, the metal has been
1459
useful in the crystallization of the insulin molecule.
Hormone-zinc interactions. Just as zinc influences the endocrine system,
changes in the concentrations of various hormones affect the concentrations of
several trace metals in blood, urine, and other tissues. Thus, increasing or
decreasing the concentration of trophic pituitary hormones has been associated
with fluctuations in body concentrations of zinc as well as changes in the secre-
tion of hormones from target glands. Differences in the hormonal secretion of
several target glands also have been associated with changes in zinc concentration.
Whether or not releasing hormones in the hypothalamus influence trace
metal concentrations is not yet known.
Changing levels of circulating hormones may alter the distribution of zinc
in various cellular compartments, promote alterations in zinc-ligand interactions,
or alter renal or hepatic handling of trace metals. These changes generally
accompany changes in the body concentration of zinc either because they produce
a shift to tissue compartments not normally associated with such zinc concentra-
tions or they may increase urinary and fecal loss, thereby bringing about total
body loss.
Effects of growth, hormone. Increased concentrations of circulating growth
hormone, which occur in patients with untreated acromegaly, have been associated
237
-------
667
with reduced serum zinc and greater urinary zinc excretion. Treating these
patients with surgical hypophysectomy or X-irradiation, procedures which lower
circulating growth hormone levels, also decreases urinary zinc excretion, in-
creases serum zinc concentration, and slightly decreases serum copper concentra-
667
tion.
Conversely, decreased concentration of circulating growth hormone, which
occuis in patients with untreated, isolated growth hormone deficiencies, has been
associated with elevated levels of serum zinc and decreased excretion of urinary
667
zinc. Treating these patients with exogenous growth hormone lowered the
elevated serum zinc level and elevated the previously reduced urinary zinc excretion,
To demonstrate these interrelationships further, changes in trace metal
metabolism were studied following the intravenous administration of arginine
hydrochloride to patients of short stature to evaluate
growth hormone release. ' In one subject with low plasma levels of growth
hormone, serum zinc concentration was elevated and excretion of urinary zinc was
depressed. Following arginine administration, growth hormone concentration in
blood increased, whereas serum zinc levels fell; urinary zinc excretion increased
in temporal relationship with the rise of growth hormone in plasma.
These results suggest that growth hormone levels in blood are directly
related to urine levels of zinc excretion. These changes may be related to the
manner by which growth hormone either directly and/or indirectly affects the
binding of zinc to its macromolecular and micromolecular ligands in blood, thereby
537,668
affecting the urinary excretion of zinc. However, the specific relation-
ships between total serum zinc concentration and plasma growth hormone levels are
there was
not entirely clear, because/no significant correlation between these two variables
were ggy
when the data obtained / plotted. The lack of significance may indicate that
668
blood growth hormone levels do not influence macromolecular liganded zinc as
238
-------
668
much as zinc bound to albumin or peptides. Therefore, total serum zinc may
only reflect some of the physiologic and biochemical changes that occur in
hormonal disorders.
Effects of progesterone. Little or no change in serum or urinary zinc occurs
after rats receive small amounts of progesterone. However, serum zinc concentra-
tions decrease following administration of large amounts of progesterone to
1415
rats. In women receiving various drugs to control fertility, serum zinc con-
613
centrations may be lowered, whereas serum copper concentrations are elevated.
The mechanisms by which these changes occur have not been well studied.
Effects of other gonadal hormones. In adult rats with gonads removed, serum
1415
zinc concentrations are uniformly low. They increase with the administration
gonadal
of the appropriate male or female/hormones. The pattern of change in serum zinc
599,
has been well studied in women and rats during their menstrual or estrus cycle,
1203,1415 599,676,797,839,1203,1548 1415
pregnancy, and during pseudopregnancy in rat.
Each condition demonstrated an inverse relationship between estrogen and progesterone
and serum zinc concentration. In these studies little influence of pituitary
415
gonadotropin on zinc metabolism in gonadectomized rats was observed, although
the influence of pituitary hormones on male hypogonadism in rats is well known.
Zinc, as noted, is present in the placenta of pregnant womenjand levels change
during pregnancy. In one study, at term, the zinc content of the
placenta was high, representing 1-2% of the total zinc content of the pregnant
1548 480
female. In another report, the zinc content of the placenta was highest
6-12 wk into the pregnancy and it decreased thereafter. The role of zinc in the
placenta is really not known. However, it may be related to its high level of
707 253
alkaline phosphatase, an enzyme that increases in serum as pregnancy proceeds.
239 .
-------
Changes in serum zinc have been studied during the menstrual cycle of normal
females with contradictory results. However, there was one indication that serum
zinc concentrations did exhibit changes during the menstrual cycle, with the
599
highest level coinciding with the time of ovulation. Whether any specific
relationship between ovulation and zinc metabolism exists is still unanswered.
In humans, the influence of gonadal steroids and their analogues has been
most clearly demonstrated in the effects of these agents on lowered serum zinc
611,613,1415
levels. No specific contraceptive effect was imputed to the changes
in serum zinc levels, but the possible importance of their coexistence should be
noted.
The exact role of zinc in testicular function is not known. In rats fed
80
zinc-deficient diets, testicular atrophy eventually developed. In zinc-deficent
rats, alterations in protein and nucleic acid metabolism in the testes were
found. Studies of the effects of gonadotropins and testosterone on zinc-65
uptake in male rats also have been carried out. Administration of FSH
decreased zinc-65 uptake in tissues of the reproductive tract as well as tissues
not involving gonadal function.
The nature of the synthetic defect in testosterone metabolism from zinc
deficiency has not been clearly established. It is clearly demonstrated that
sperm contain the highest concentration of zinc of any bodily tissue per milli-
1338
gram protein. High zinc concentrations have been found in the heads of
604*
individual spermatozoa by X-ray microprobe techniques, although this finding
511a
has been ascribed by later workers to contamination with seminal vesicle fluid.
Zinc is highly concentrated in ova of all species, but its purpose, except as
related in some general way to growth and development, is not clear.
Changes in the metabolism of pregnant women produced changes in zinc con-
767a,1404,1770a
tent of some tissues. In eclampsia of pregnancy, zinc levels
decreased to about 10% of their normal amounts. Infectious processes or neo-
plasia complicating pregnancy raised the content of placental zinc. It has been
240
-------
suggested that loosely bound zinc may affect the epithelial tissues of the
uterus. Some toxic states in pregnancy are associated with the loss of zinc by
1138
the placental epithelium.
Effects of jflsulin. Because of the high content of zinc in the islets of
Langerhans and the prolongation of action of insulin by zinc, a relationship
between zinc and diabetes has been sought. In patients with diabetes mellitus
without proteinuria, urinary excretion of zinc was reported to be above
normal. In patients with diabetes and proteinuria, urinary zinc excretion is
310,717
understandably higher than in normal subjects. Plasma zinc levels of
fasting diabetics have been reported to be higher than in normal subjects, and
diabetics of more than 10 years' duration have been reported to have plasma zinc
310
levels higher than those in the earlier stages of the disease. Whether or
not the extremely high values in diabetics of more than 10 years reflected pro-
longed therapy with zinc insulin was not determined.
However, in recent studies, no correlation could be obtained between plasma
361
levels of zinc, glucose, or glycosuria. Nevertheless, these investigators
367,717
demonstrated a striking fall in plasma zinc after loading with oral glucose.
On the basis of fat-free pancreas tissue, little difference in zinc content of
422
the pancreas from normals or diabetics was measured by some investigators,
although others estimated that the zinc content of the pancreas was one-half that
1460
of nondiabetics. These problems have not been resolved yet.
In oral and intravenous glucose tolerance tests, glucose rose as plasma
361,717
zinc concentration fell; however, the curves were similar for both normal
subjects and diabetic patients. In zinc-deficient rats, glucose intolerance was
intraperitoneal
observed after an / glucose load but not after an oral glucose load.
Thus, insulin has a biologic action independent of zinc. However, zinc
forms a physiologically active compound with insulin that delays its physiologic
241
-------
1459
action and greatly prolongs the hypoglycemia. Many studies have attempted
to elucidate the structure of zinc insulin. Imidazole groups are
165,1570
implicated in the binding of zinc to insulin. The ratio of zinc to the
1271
insulin monomer is approximately 0.5 g atom/mole; in actuality, there are 3
zinc atoms/insulin hexamer. Measuring the kinetics of photooxidation suggested
that the rate of photooxidation of zinc-free insulin was considerably faster than
1746
that of zinc insulin. Analyses by X-ray techniques have offered evidence of
the structure of crystallized insulin from different species. For example, bovine
391,632
insulin crystals are rhombohedral. Zinc is useful in the crystallization
of insulin: if the ash content of crystalline insulin is lowered it will not
1457a,1459
crystallize again unless a metal salt is added.
Effects of parathyroid hormone. Patients with untreated hyperparathyroidism
971a,971b
exhibited increased urinary zinc excretion. However, the mean concentration
of total serum zinc in these patients was normal. Following surgical treatment
of the hyperparathyroidism, urinary excretion of zinc decreased among patients
whose parathyroid hormone (PTH) levels reverted to normal; in patients with per-
971b
sistent hyperparathyroidism, no decrease in urinary zinc occurred after surgery.
These clinical results suggest that parathyroid hormone was directly related to
body zinc loss from urinary excretion. The results were partially confirmed by
studies in which purified parathyroid extract was administered intravenously to
two patients in whom urinary zinc excretion increased two- to threefold during
the second hour after infusion. However, their urinary cyclic adenosine 3':5f-
cyclic phosphate (AMP), which increased after the first 15 min, had returned to
971b
baseline levels.
There are several possible ways PTH could have produced these results. For
example, increased metal excretion observed in patients with untreated primary
242
-------
hyperparathyroidism or after PTH infusion could have been the result of an
aminoacidurla induced by the PTH excess. About 36% of the patients with primary
346a
hyperparathyroidism have been reported to show a generalized aminoaciduria,
and intravenous infusion of PTH increases the urinary excretion of histidine and
1491a
other araino acids by about 50% within 2 h. Since oral administration of
histidine has been shown to increase urinary excretion of zinc, these results
are consistent with the concept that the blood histidine strips the zinc normally
complexed with albumin, forming histidyl-zinc ligands that are readily excreted
668,679
in the urine. Another possible effect relates to the increased bone turnover
800a
commonly observed in patients with primary hyperparathyroidism. Since a
1640
large proportion of body stores of these metals is located in bone, the in-
creased urinary excretion of these metals in hyperparathyroidism may be an index
of bone turnover. The increased urinary excretion of copper and zinc observed in
667
acromegaly, a disease in which bone turnover may also be increased, may
also reflect this phenomenon.
Effects of adrenocortical steroid. Abnormalities of adrenocortical steroid
metabolism in man and animals have been linked to changes in copper and zinc
670
metabolism. Adrenalectomy or adrenocortical insufficiency from several causes,
including idiopathic Addison's disease and hypopituitarism, have been accompanied
by increased serum zinc concentration, decreased urinary zinc excretion and in-
670,677
creased retention of zinc in several tissues. Exogenous replacement of
adrenocortical steroids in both man and animals corrects these changes by
normalizing the serum concentration, urinary excretion and tissue retention of
479a,670
the metal.
1091b 932
Circadian changes in copper and zinc levels in blood have been
associated temporally with the circadian variation observed in the levels of
667
17 hydroxycortical steroids in blood and urine, although administration of
243
-------
exogenous carbohydrate-active steroids did not abolish the circadian variation
932
observed for these metals.
Conversely, elevated endogenous secretion of adrenocortical steroids, which
occurs in Gushing*s syndrome or in patients with adrenocortical carcinoma, has
been associated with decreased plasma zinc, increased urinary zinc excretion, and
670,677
decreased tissue retention of zinc. These changes are a consequence of
increased excretion of metals. Any treatment of these diseases that helped the
excessive endogenous adrenocortical steroid secretion brought about the return
of serum copper and zinc concentration to or toward normal, a reduction in urinary
670,677
excretion of zinc and an elevation in tissue levels of zinc.
Similar changes in patients with hypersecretion of adrenocortical steroids
were observed following oral administration of exogenous carbohydrate-active
670
steroids to normal volunteers. After five days of being dosed with 50 g
prednisolone daily, the subjects' serum zinc concentration decreased and urinary
zinc excretion increased.
These studies indicate an inverse relationship between adrenocortical steroid
levels in blood and serum zinc concentration, and a direct relationship between
blood levels of adrenocortical steroids and urinary excretion of zinc. These
changes may be related directly to some adrenocortical steroids affecting the
669,670
production of increased ultrafilterable zinc in serum. Such changes
previously were associated with decreases in serum zinc concentration and increases
670
in urinary zinc excretion. However, increased renal excretion of copper after
1748a
adrenal corticosteroid administration may be linked to changes in glomerular
filtration produced by adrenocortical steroids. These fluctuations may also affect
serum levels and urinary secretion of zinc and copper. It has recently been
demonstrated in mammalian cell cultures that zinc uptake was stimulated by the
1205
adrenocortical steroids hydrocortisone and psednisolone. The action of these
244
-------
steroids on zinc accumulation was thought to be caused by stabilization of cellu-
lar membranes,^since zinc protects membranes from fragmentation during subcellular
fractionation.
Prostate and Gonads
Zinc concentration in the prostate, seminal vesicles, and other associ-
ated structures is higher than in any other soft tissue of the body. Table 7-9
lists zinc concentrations in the male reproductive tract. In Sanyal and co-workers'
studies of rats, removal of zinc caused irreversible degeneration of the seminiferous
tubules of the testes. Degenerative changes were also found in the prostate and
seminal vesicles after zinc depletion, but they were reversed after administration
1412a
of androgens, chorionic gonadotropins, or zinc supplements.
Zinc concentration in prostate tissue undergoing various types of pathologic
changes has been reported to vary widely from normal. Verrilli et al. attempted
prostatic
to make a diagnosis of early/carcinoma and a differential diagnosis of prostatic
nodules by studying the differential uptake and distribution of zinc-65 throughout
prostatic tissue; their efforts were unsuccessful in making a selective differ-
entiation.1685
245
-------
TABLE 7-9
a
Zinc in the Male Reproductive Tract~
yg/g Fresh Tissue
Mean Range
Prostate gland
Lateral lobes
Posterior lobe
Epididymis
Testes
Ampulla of vas deferens
Seminal vesicles
Striated muscle
210
190
115
61
153
108
68
124-399
123-360
73-189
41-130
57-295
39-305
54-86
Number
of
Organs
11
10
9
9
12
17
10
a
"Data
from Lindholmer and Glauman.
937
246
-------
Zinc in prostatic cancer, studied by the assay of individual tissue sections,
1443
confirmed previous findings of lower than normal levels of zinc in this tissue.
However, a high zinc level in prostate has been observed in benign prostatic
1443
hyperplasia. Normal human prostatic fluid contains a high level of zinc, and
in cases of benign hyperplasia the most elevated zinc levels were in prostatic
fluid, an extracellular fluid, instead of within cells. The in vitro uptake of
zinc-65 was highest in cases of prostatic cancer and lowest in prostatic hyper-
plasia. An inverse relationship has been observed between zinc concentrations
594
and the in vivo and in vitro binding capacity of zinc-65 in different patho-
593
logic stages of prostatic tissue. Further evidence showed that zinc within
the prostate was bound to sulfonated-acid mucopolysaccharides: autoradiographs
indicated that zinc binding areas corresponded to areas of acid mucopoly-
593
saccharides.
A zinc-binding protein has been isolated from human benign hypertrophic
659
prostate tissue. Analysis of amino acids in this protein reveals a very high
molar ratio of histidine. The importance and specificity of this finding awaits
further investigation.
Because of the wide variation of zinc reported in prostate—ranging from
709,1443
30 to 2,315 yg/g dried tissue— several investigators attempted to cor-
relate zinc concentration in prostate by the histologically estimated ratio of zinc in
and 835a,948b
epithelium / stroma. However, the results were not very useful.
544a
Gonic e£ al. stated that zinc prostate concentration varied inversely with
prostatic
the amount of/fibromuscular stroma. In an attempt to establish some standard,
several investigators attempted to correlate levels of zinc and 3-glucuronidase
1091a,1091c,1143a
activity in prostatic tissue. Their results indicated a good
correlation between zinc concentrations and B-glucuronidase activity in noncar-
cinomatous tissue but not in carcinomatous tissue.
247
-------
Several investigators, using techniques correlating zinc levels with histo-
chemical enzyme activity, confirmed the extremely low levels of zinc in carcino-
593,1091c
matous prostatic tissue (40-50 ug/g tissue wet wt), and reached fairly
good agreement about the level of zinc in noncarcinomatous prostatic tissue
140a,1091c 835a,
(135-140 yg/g tissue) and in prostatic fluid (480-690 yg/g tissue).
948b,993,1091c,1766a
Since patients with benign prostatic hypertrophy exhibit elevated levels of
zinc in prostate, treatment with several chelating agents has been suggested as a
way of producing decreases in the symptoms associated with this condition and de-
593,948b,1008a,1443
creases in the size of the prostate. These results have not
been tested in controlled clinical trials, yet they
do appear encouraging. These effects could be explained as an effect of the
chelating agents on the noncarcinomatous prostate cells such that hyperplastic
areas of the prostate are shrunk.
Low levels of zinc in prostate have been observed not only in carcinoma of
the prostate, but in patients on zinc-deficient diets. Low levels of zinc also
have been observed in the prostatic fluid and testes of patients with zinc deficiency!
In recent years the incidence of hypogonadism among zinc-deficient subjects has
383,1174,1277
been noted, and the relatively high concentrations of zinc in the
prostate gland and in sperm have been related to the role that zinc plays in
interaction
gonadal function. However, this / has not been clearly defined. In a double-
blind study of zinc-deficient Egyptian dwarfs given either placebo or supplemental
zinc, no differences in growth or gonadal function could be demonstrated between
1410
the two treatments. Studies have been carried out in which gonadal weight
and zinc content of rats fed zinc-deficient diets have been compared with similar
measurements made in controls fed restricted amounts of food. Interpretation of
993
this study is difficult since 3-4 rats were grouped in each cage, thereby
248
-------
greatly increasing the variability of the results. Exogenous testosterone or
pituitary gonadatropins also have been given to rats fed zinc-deficient diets,
1045b
but as pair-fed controls were not studied simultaneously interpretation of
these results is limited. However, there is little question that decreased
testicular weight in zinc-deficient rats is a consistent feature associated with
zinc deficiency.
Differences in morphology between the testes of zinc-deficient and pair-fed
control rats indicate a true spermatogenic arrest in the zinc-deficient animals in
1272a
one study. Although destruction of the germinal epithelium was not observed,
1045b
which others had noted, decreased tubular size was apparent, as was the
absence or marked diminution in number of sperm. The presence of Leydig cells
1272a
in the testes of zinc-deficient rats was notedin one study. Vacuolated
1272a
nuclei of the spermatocytes were reported in this same study. Zinc deficiency
also has been observed to reduce the diameter of testicular tubules and to pro-
duce atrophy of the germinal epithelium up to the spermatogonia stage with
1045b
sloughing of cells into the tubular lumen. This latter change became more
apparent as the zinc deficiency was prolonged. It must be pointed out that
starvation, i.e., pair-feeding per se, apparently did not produce significant
1272a
histopathologic changes in the rat testes.
That zinc deficiency alters gonadal function is clear; however, the manner
by which it occurs is unclear. Either zinc is involved in some aspect of gonadal
function other than testosterone synthesis, e.g., in the synthesis or release of
pituitary LH or FSH, or the inanition produced by zinc deficiency impairs
gonadotropin release. Although little data are available to support either
hypothesis, previous studies suggest that zinc and gonadatropin function may be
interrelated. Exogenous administration of gonadotropins to zinc-deficient rats
1045b
increased the weight of epididymis but did not alter testicular zinc concentration.
• 249
-------
Although the results of these studies were quite variable and pair-fed rats were
not studied, some gonads of zinc-deficient rats appeared to mature once gonado-
tropins were administered. These animals' pituitaries were small in size and
more morphologic "gonadotrophs" were recognized when compared with those of
normally fed controls. In some ways the morphologic appearance of the testes
of zinc-deficient rats is similar to that of patients with hypogonadotropic
hypogonadism; i.e., a spermatogenic arrest accompanied by spennatogonia and
Leydig cells is observed in both conditions.
In addition to these effects of zinc deficiency on testicular function, zinc
1766
in high concentrations can be spermicidal.
Renal Function
Extrapolations from model studies suggest that the unhindered passage of
zinc-amino acid complexes in normal plasma across the renal glomerulus would re-
sult in a calculated filtered load of 2 mg zinc in a normal 24-h glomerular filtrate
537 537,668,1503,1551,1666
of 183 1. Observations of several investigators indicate
that under normal conditions about 0.5 mg zinc is excreted in urine daily. Thus, the
major part of the normal filtered load of amino acid-complexed zinc must be reabsorbed
by the kidney. Although earlier workers suggested that urinary zinc excretion was not
a function of intake, this notion may require further evaluation because many
recent studies have questioned it.
If plasma zinc shifts such that more or less zinc is complexed with micro-
668
molecular ligands, normal zinc transport and storage processes may be altered. An
increase in micromolecular zinc ligands (amino acid-zinc ligands) crossing the
renal glomerulus could lead to excessive loss of zinc through urine. Albuminuria
would lead to hyperzincuria as well.
250
-------
The manner by which zinc is reabsorbed by the kidney is not clear, although
analyses by laser microprobe and histochemical techniques have suggested that a
high concentration of zinc resides in the proximal tubule of the kidney, where
reabsorption may be expected to take place.668'1483 if reabsorption of zinc
occurs, then the role of metallothionein as an agent in the conservation of zinc
could be an important one. However, agents such as carbonic anhydrase and alka-
line phosphatase—both zinc-containing enzymes—have been shown to occur in high
541a
concentrations at the brush border of the renal tubule. Thus, although zinc loss
is controlled primarily through size discrimination at the renal glomerulus (which
inhibits the loss of macromolecular-liganded zinc moieties) , specific properties
of other substances and enzymes influence the tubular reabsorption of a very
large amount of zinc.
Schroeder has shown that kidneys of human subjects dying from hypertensive
complications had increased concentrations of cadmium or increased cadmium in
1444
relation to zinc. The systolic hypertension in rats caused by cadmium salts
may be reversed by injecting a chelate of zinc with a higher stability constant
430,1448
for cadmium than for zinc. It has not only been concluded that cadmium
plays some role in hypertension, but that the ratio of cadmium to zinc is very
important. In metallothionein, which contains both zinc and cadmium, these
two metals may interact with the same binding site and functionally replace each
other. Since the physiologic role of this protein is not understood, these data
cannot be specially related to renal pathology. All data suggest that, in the
1451
kidney, cadmium and zinc are closely related. In Figure 7-9, Schroeder et al.
have shown that the concentration of zinc and cadmium in kidneys changes in
amount and ratio in different decades of human life. The curves represent mean
concentrations of 221 subjects. The difference in peak levels in age groups
between 40 and 50 is statistically significant.
251
-------
Cd Zn ppm
ratio ash
07
06
05
0.,
0.3
0.2
01
00
7000
6000
5000
4000
3000
2000
1000
x — x
/ ^ <
•_• _^ .,
.,//- * ^-^ .Zn
^m t .0- X
y 'S / -•-•'"" 0'""--
\ S '
\ Si .•' ° *-.
^7' / •/' "••••"
s/y
o-x> * "V9 »-19 20-29 30-39 40-49 50-59 60-69 70-79 80->
Age in Years
FIGURE 7-9 Mean concentrations of cadmium and zinc and their ratios in
kidneys in 221 subjects from the United States. Reproduced
from Schroeder et a
252
-------
Zinc has been found in a large number of urinary calculi, and in some
patients with calcium oxalate renal calculi, hyperzincuria has been
430,1336
observed. Additional studies have suggested that only men with renal
294,343
calculi exhibit hyperzincuria. Hydrochlorothiazide has
been suggested as a useful agent for preventing calcific renal stones.1810'1811
1193
Administration of this drug also produces a significant hyperzincuria, but
evidently without significant cupruria. Zinc has been found to increase the
429
solubility of calcium oxalate in vitro, which may relate to the mechanism of
this effect. However, sodium cellulose phosphate, another
agent used in the treatment of some forms of renal stones, has no effect on renal
1192
excretion of zinc. Nephrocalcinosis was produced in rats fed a diet deficient
in magnesium and zinc, and these renal stones were more common in females than
1810
in males.
Muscle Function
Several studies over the past 20 years have attempted to establish a role
for zinc in muscle function. This function has not been clearly defined, but
zinc might be necessary to muscle contractility. Although not directly related,
examinations of cupric and zinc ions bound to crystalline sperm whale myoglobin
and studied by X-ray diffraction analysis indicated that the metals are bound by
histidine and probably by lysine and asparagine
V side chains with high affinity to myoglobin. Cann has suggested
227
that zinc ion alters the iron-imidazole linkages in myoglobins. The reaction
of zinc with myoglobin causes major changes in physical and chemical properties
of the protein. The most characteristic spectral change is a marked reduction
226,227
of Soret band intensity, which shifts the peak of absorption to 390 mm.
Whether metal ions other than the iron atom of theheme group
is physiologically involved in the function of myoglobin is not known.
253
-------
Collagen
Recent studies in rats indicate an important role for zinc in collagen
732
formation. For example, a primarily copper- and
733b
iron-dependent enzyme, lysyl oxidase, was shown to be zinc-dependent as well.
Cross linking per se appears to require zinc. Zinc deficiency appears to be
associated with a decreased proline . uptake in the epiphyses, a de-
creased transformation of proline to hydroxyproline, lowered sulfur-
923 1142
35 uptake and decreased epithelial shearing strength. ' Because zinc is
important for the activity of alkaline phosphatase, and alkaline phosphatase has
been related to the secretion of collagen and/or mucopolysaccharides, then a role
for zinc in this system may be reasonably surmised. Zinc also may inhibit
1083
the pyrophosphatase activity of alkaline phosphatase in humans.
Hematopoiesis and Red Blood and White Blood Cell Function
In vitro studies of zinc-65 transport into leukocytes and erythrocytes and
1041,1164
into erthyrocyte carbonic anhydrase have been carried out. Zinc-65
377
uptake in leukemic leukocytes has been found to be only half that of normal cells.
Carbonic anhydrase, first discovered in bovine erythrocytes, has been found
831
in several human tissues. Three forms have been found in human erythrocytes
33,901,1151
and named carbonic anhydrase: A, B, and C. These different forms are
similar in their role in catalyzing the reversible hydration of carbon dioxide,
in their molecular weights, and in the one atom of zinc in each molecule.
Studies of the mode of binding with different inhibitors, especially some
derivatives of sulfonamides, described the mode of action of the active site and
302
the differences between them. Carbonic anhydrase B has at least one histidine
1767,1768
molecule located in the active site region close to the metal. Using
t
zinc-65, it was shown by kinetics that immediate inactivation is caused by the
formation of an inert ternary complex and that the mobilization of zinc proceeds
235
slowly by formation of the apoenzyme. Infrared spectrophotometry revealed
that bound carbon dioxide attacked the hydroxyl coordinated to the zinc
1346,1347
ion. Other investigators studied the structure of the active site of this
254
-------
enzyme; they constructed a model that corresponded to the active site.388"390
Both human carbonic anhydrases B and C act as esterases on g- and £- nitrophenyl
acetates and the mechanism of the carbon dioxide hydration re-
1684
action may be extended to the esterase reaction. Chelating agents such as
0-phenanthroline at pH 4 and below may remove zinc and deactivate
414
the enzyme. The metal specificity of the enzyme seems to be independent of
1613
the substrate.
1365,1588,
Alkaline phosphatase also has been found in white blood cells of man,
1629
and its concentration has been shown to be abnormal in several blood
1365
dyscrasias. Reverse transcriptase, a metalloprotein containing two zinc
1255
atoms per mole of protein, has been isolated and a function in human leukemia
596
has been suggested.
Sensory Function*
The role of zinc in taste and smell has been documented in humans and
331,617,621,666,671,673,679,680,1004
animals. Zinc deficiency is associated with
loss of taste and smell acuity and the appearance of dysgeusia, dysosmia, and
621,666,671 666
pica in man. Zinc may be a component of the taste bud and in subjects with
678,1087
normal taste acuity it has been shown to be present in the saliva that
bathes the taste bud. Several causes of loss of taste acuity in humans are
678,1087
associated with decreases in salivary zinc concentration and the
671,680
appearance of pathologic changes in the taste buds. Zinc is an integral
part of a protein known to be secreted by the human parotid gland. The function
of this protein appears to be related to the maintenance of normal taste bud
674
form and function.
Zinc also may affect the visual system. Zinc has been found in high concen-
trations in the choroid of the eye. a This concentration has been caused by
*See also Chapter 10.
-------
localization of the metal Ln the tnpatum lucidum ceLluloaum, which la thought to b«
reaponaiblu for tha light reflection Mean in the ayes of carnivoraa. It ii thi
reduction of tha zinc contant of thia tiasue that haa baan auapactad to ba tha
affect of chlorambucil In reducing tha ophthaltnologic raflaotion
in canina tdpatum lucidum.
Ratinane reductnaa, tha Alcohol dehydroganaaa which convarta ratlnol to
retinal, In a Bine-dependent enzyme. Tha presence of thla enzyme in
126
tha retina partly nay explain the high laval of zinc in the retina. Thia
enzyme may nlao affect t ho olfactory syattm bacauaa vitamin A laveIn and hypoamia
672
arc aaaoclatad in aoveral pathologic atateii. Zinc lavela in tha olfactory
epithelium have not bean measured yet.
Liver
Several zinc-dependent enzymea have bean found in liver. Liver alcohol
dahydrogenaaea catalyze the snme reaction aa in tha retina, that ia, a revereible
oxidation and reduction of alcohols to tha corroaponding nldehyde* or katonea in
the preaance of nicotinamide adenlne dinuclaotide (NAD) or reduced nlcotinamidn
adanine nucleotide (NADH). They have different amlno acid compoaitiona and dif-
636
ferant total and aaaenLial aulfhydryl groupn. An early Mtudy reported that
1607
horae liver alcohol dehydrogenaae had two zinc atoma par mole of enzyme, but
a 13
in later work ueing/zinc-65 chloride tracer ua well aa nonrndioactiva
1173
methoda, four atom* of zinc per molo wore identified in thia protein.
Zinc porphyrlna are commonly found in liver, /inc protoporphyrina have been
1161
found in yeaat and zinc chelataae, an enzyme that atmlatti in incorporating
zinc into protoporphyrin haa been detected in different
1134,1292
extracta. Zinc farrochelataae aluo may act to incorporate ainc into
888
protoporphyrina. There La alao aoma aupport for the notion that zinc proto-
porphyrin and iron porphyrine are catalyzed by the anme enzyme, an both enzymai
256
-------
998
In the native state are cloeely associated with phoepholipide. In Saccharomyces
cerevialae several Bine porphyrins have bean isolated, suggesting that cine proto-
1292
porphyrini are praaant In Intact calls.
Pancreaa
Carboxypeptidaae haa baan puriflad from bovina and porcina pancreaa, with
•mall variations in properties. Bovina pancreatic carboxypaptldaaa A haa a-, (<-,
Y-, and 6- forma that diffar in solubility with raspact to tha amlno aclda of tha
884
N-tarminal raaidua and in reactivation of tha apoancyma by /.inc. Tha
76,1791
amino acid compoaition for moat typaa has baan datarminad. Tha cine atom
of tha activa alta of y- and 6- forma la bound to tha thlol group of tha
1248
cyataina raaidua and tha o-amino group of tha N-tarminal of aaparagina.
1793
In carboxypaptidasa B, ona cyatainyl raaidua ia tha ligand for nine. Many
studiaa hava baan conducted recently to evaluate tha active site and investigata
121,303,306
relatlonahips between atructure and function. Carboxypeptidaae B
1066
activity ia decreaaed in line-deficient rata.
Vitamins
Hau reported that pyridoxine-deficient rata have low tine content
721*
in plasma, liver, heart, and pancreaa. Conversely, Oershoff found that
pyridoxina and vitamin B. defioiencias increaaed the cino con-
tent of rat pancreaa, liver, and kidney. The greatest increaae in cine was
in the content of the pancreaa. In pyridoxina-deficient rata the decreaeed
availability of insulin was postulated aa the cause of the higher cine leveln.
IH vitro atudiee with adipoae tiasue isolated from theae rata indicated that xinc
inhibited the lipogenic activity of the adipoae tiaaue, suggesting that it might
3fl7
influence the control mechanisms related to inaulin activity.
257
-------
1512
A role for zinc in vitamin A metabolism has also been suggested,
but the relationship is unclear because of the complex interactions between zinc
and protein metabolism and between zinc and vitamin A. This proposed relation
may be more related to the availability of protein than to vitamin A itself—1505
were observed to
anorexic, zinc-deficient animals/take in less than normal amounts of protein.
Cardiovascular System
Little is known about the specific role of zinc in the function of any
particular aspect of the cardiovascular system, but important relationships have
been postulated. As already noted, an interaction between renal cadmium and zinc
1444,1448
has been suggested as an influence on some forms of hypertension. The
urinary excretion of cadmium was greater in 95 hypertensive women treated with
antihypertensive agents than in 107 normotensive women, but zinc excretion was
1013
the same in both groups. Carroll attempted to link the zinc concentration
in air and the incidence of cardiovascular disease—including hypertension—in
240
28 North American cities. His results are difficult to correlate with the
multiple or indirectly
/ clinical factors that directly/relate to the pathogenesis of the disease processes.
Some investigators have suggested that zinc deficiency may be a factor in
fiofi I o£i
the development of atherosclerosis * and that zinc therapy is useful in the
686
treatment of some of the symptoms of obliterative vascular disease. One
investigator noted that the zinc content of the aorta of atherosclerotic patients,
1703
particularly in the areas of the atherosclerotic plaque, was markedly decreased;
he also noted that these patients exhibited decreased zinc in plasma, liver,
myocardium, pancreas and hair. It has been suggested that accumulation of zinc
1133
in the renal cortex promotes aortic atheromata and hypertension. Another
474
group experimenting with hypertensive dogs could not find these large differences;
these studies were carried out in animals with a different set
of cardiovascular problems. Zinc-protamine-glucagon has been claimed to be useful
258
-------
in the relief of symptoms related to cardiac insufficiency,812 although the role
of zinc itself in this complex may not be critical.
After acute myocardial infarction, human serum zinc concentration falls. >17U
Dogs with experimentally produced myocardial infarctions had reduced zinc concen-
936
trations in the infarcted area. However, no differences in zinc were found
in heart muscle analyzed from patients who died from acute coronary thromboses
274
although magnesium levels were decreased. In addition, zinc was carefully
measured in the thoracic aorta of patients with graded levels of atherosclerosis
and differences between those subjects and patients not suffering from cardio-
1310
vascular disorders could not be measured. Such diverse results make it dif-
ficult to ascertain the specific influence of zinc in cardiovascular function.
Other Enzymatic Activities
Zinc complexes with adenosine diphosphate (ADP) to form
Zn.ADP, ZnADP, and Zn(ADP>2 in dilute solutions. Zinc markedly reduced the
conversion of adenosine triphosphate (ATP) to AMP in lipocytes
1784
when used alone or with magnesium. ATP-metal-norepinephrine
ternary complexes may be of major importance to the understanding of the biologic
298
phenomenon of binding or storage of catecholamines in vivo.
244,1231
Zinc has also been found to affect the activity of ATPase, acid
1728 287
phosphatase, alkaline 6-glycerophosphatase, phosphopyruvate hydratase,
1091
and yeast aldolase. Zinc inhibits the activity of several enzymes, including
158,690 1698
ribonuclease, allantoinases from bacterial and animal sources, ot-
1742 1272
oxoglutarate dehydrogenase, and sodium- and potassium-activated ATPase.
The character of the inhibition for each enzyme is specific.
1005
Superoxide dismutase, an enzyme important in oxygen metabolism, contains
645
two zinc atoms, perhaps as structural components of the enzyme. Zinc is bound
490
at the histidine site of this enzyme.
259
-------
Zinc-a2~glycoprotein, with a molecular weight of 43,000, is present in
1427
most body fluids and has been isolated from normal plasma. It is found
in low concentrations in normal serum and cerebrospinal fluid, and in higher
concentrations in urine, saliva and sweat. This glycoprotein appears to
be homogeneous, as shown by ultracentrifugation, electrophoresis, chromatography
and N-terminal-ami no acid analyses. In the low pH range, the protein appears
in several fractions even after the removal of sialic acid. The polymorphism
of zinc-(X2~glycoprotein may be an expression of genetically determined
1429
variants. The zinc-a2~glycoprotein should not be confused with the major
serum macromolecular binding protein of zinc-a2~niacroglobulin noted in the
beginning of this chapter.
260
-------
CHAPTER 8
ZINC IN THE DIET AND THE EFFECTS OF ZINC DEFICIENCY IN ANIMALS*
Zinc must be present in the diet of all animals, including man.
Moreover, it must be supplied In the diet almost continuously, because
animals have only small amounts of readily available zinc stored in the
body. This condition exists although the body contains a relatively
1640
large amount of zinc, particularly in bones, skin, and hair. However,
zinc in these tissues cannot normally be utilized by the animal to meet
its need for zinc.
LACK OF ZINC IN THE DIET
The primary effect of insufficient zinc in the diet is decreased
food intake and cessation of growth. The effect is quite rapid; growth
1195,1782
stops almost Immediately in rats and in about 2 wk in calves
1064
and lambs. Other signs of zinc deficiency observed in young growing
animals are listed in Table 8-1. It should be emphasized that the
additional symptoms described generally were observed in very young
animals that would have been in an active stage of growth. Somewhat
older animals will also stop growing soon after being placed on a zinc-
deficient diet but may not show other signs of deficiency. Six-to-nine-
month old calves, for example, failed to grow after being placed on the
1064
deficient diet, but showed no other clinical signs of deficiency.
Undoubtedly, adult animals are also affected by a lack of zinc in
the diet, but less information exists on the effects of a low zinc diet
*Work discussed in this chapter appeared in the literature prior to
1640
April 1975. Papers included in Underwood's comprehensive review
generally have not been covered here.
*See Chapter 10 for symptoms in'humans.
261
-------
TABLE 8-1
atoms of Zinc Deficiency in Young Animals
and Approximate Requirements for Zinc
Species
Ruminants
Cattle
Sheep
Deficiency Symptoms
Parakeratosis, increased bacteria
in mouth, stiffness of joints with
swelling of feet, horny over-
growths, excessive salivation
(transitory)1050
Salivation (transitory), increased
bacteria in mouth, parakeratosis
of tongue, '^ loosening of wool,
distortion of hoof walls and
horns10**
Goats
Swine
Poultry
Chicken
Similar to cattle
1114
Parakeratosis
1127
Poor feathering, shortening and
thickening of long bones of legs
and wings, hock enlargement,
reduced egg production and
hatchability, skeletal abnormali-
ties in embryos
Turkey
Japanese
quail
Laboratory Animals
Abnormal feathering,
nated gait1699
uncoordi-
Rat
ra-
Alopecia, dermal lesions, p
keratosis of the esophagus,
cyclic food intake, '^ impairment
of reproductive function1"40
Approximate Requirement^
Egg white: 10-14
Practical diet
Beef cattle: 10-3Q1114a
Dairy cattle: 401108
Egg white: 151064
(somewhat higher level
may be required for
normal testicular
development)1642
Casein diet: 14-20
Soy diet: 501114
,1156
1479
.1113
Casein diet: 12
Soy diet: 1911-
Practical dietj
Starting chick (0-8 wk):50
(1.5 times this amount may
be required during first
week to prevent abnormal
feather development)
72
Breeding hens: 65
Poults (0-8 wk): 70
251699
1113
Egg white or casein „„__ ,-„,,
diet: 12-13^6,1112,1197,1782
Soy diet: 191156
262
-------
TABLE 8-1 continued
Species
Mouse
Rabbit
Guinea pig
Monkey
Dog
Mink
Deficiency Symptoms
Alopecia1112
Sparse hair, reddening around
mouth, wet and matted hair on
lower Jaw and ruff, impair-
ment of reproductive function
in the female4*'1480
Rough coat, scaly dermatitis
Unkempt appearance, alopecia,
parakeratosis of tongue
Alopecia, hyperkeratinization,
acanthosis1^^ (symptoms and
requirement based on single
study in which a simple zinc
deficiency probably was not
present)
1001
Approximate Requirement
>3
1112
>3
44
Casein diet: 12
Soy diet: 2015
Casein diet: >15
961
Practical diet: 20
1109
<20-
a Symptoms listed are in addition to growth failure and decreased food
consumption.
- Mg/kg feed, dry weight.
- Unpublished data, H. F. Travis.
263
-------
on adult animals. Studies with rats indicate that a low zinc diet inter-
1154,1408
feres with wound healing in the adult male and with estrous
45,544 1640
cycling and reproduction in the adult female. Skin lesions
have been reported in adult humans whose zinc intake has been very low for
199
2 mo.
REQUIREMENT FOR ZINC IN THE DIET
Table 8-1 also estimates the amount of zinc that various animals
require in order to grow normally. Adult animals presumably require some-
what less than the amounts listed, except during periods of stress such as
pregnancy and lactation. The amount of zinc required varies somewhat with
the type of diet, because diets which contain seed protein (such as soy
protein) may increase the requirement for zinc. There may be other
conditions under which the requirement for zinc is increased, since
deficiencies have been reported in ruminants in the field on zinc intakes
which, based on laboratory studies, should have been adequate.
A lower than normal requirement for zinc has been reported in
1513
germ-free rats and a higher than normal requirement in a breed of
179
Danish calves with hereditary thymus hypoplasia. A genetic difference
has been suggested to affect the requirement of pigs. "°
Normally, little zinc is available to the body except that ingested
in the diet, and therefore zinc must be supplied in the diet more or less
continuously. Under some conditions, however, zinc stored in the tissues
may become available to the body. Such conditions include muscle and/or
462 1532
skin catabolism after severe injury, starvation, low protein
760 762,1599
intakes, and bone resorption during low calcium intake. The
animal may also be able to make some adjustment to a low zinc intake
264
-------
through decreased excretion of zinc and perhaps by increased absorption
199,447,1048,1456
efficiency.
Ruminants
Cattle. The suggested zinc requirement for cattle is 40 ppm for
1108 Hl4a
dairy cattle and 10-30 ppm for beef cattle. This rather wide
range exists because zinc deficiencies occur in the field on intakes of
zinc substantially higher than those which appear to be adequate under
experimental conditions.
Under experimental conditions, 8 ppm was sufficient for calves to
1064
grow,although plasma zinc levels were still low. This concentration
may be very close to the minimum, because in another study calves receiving
1052
7-8 ppm became severely deficient. On an intake of 17 ppm, lactating
cows did not show signs of deficiency nor was milk production affected,
1123
although the zinc concentration of the milk was decreased by about 25%.
1124,1126,1212,1213
Cows receiving low levels (5-17 ppm) of zinc absorbed
1125
and retained more zinc-65 than did cows fed 40 ppm. Addition of zinc
to a. barley-hay ration containing 25-32 ppm zinc did not improve weight
1302
gain of beef heifers. The experiments indicate that 30 ppm should be
an adequate zinc intake for beef cattle; but zinc supplements given to
beef calves in Greece on a ration already containing 32 ppm zinc resulted
in a 7% greater weight gain, and deficiences were reported in the field
1528
on Intakes of 20-40 ppm. Young Fresian bulls on a barley-swede
turnip ration containing 30-50 ppm zinc developed a pododermatitis that
375
was healed by oral zinc administration although growth was never
affected. Facial eczema in dairy cattle in New Zealand was treated
1345a
successfully with zinc supplements, although forage in the area had
552
not been considered to be low in the metal.
265
-------
The above cases suggest that some unknown factor influences the
zinc requirement of cattle. One possibility is phytate, although phytate
1050
content of the diet has not been considered to be a problem in ruminants.
Another factor that has not been carefully considered is soil ingestion.
657
Dairy cows may consume 1 Ib (0.45 kg) or more of soil a day. In
658,1510
some cases, animals might be able to utilize the zinc in the soil,
but ingestion of other soil types might reduce the availability of zinc.
Stress also may increase the requirement for zinc.
Sheep and goats. Although severe zinc deficiency has been developed
1064,1204
in sheep, little evidence exists of zinc deficiency occurring in
sheep in the field. A field survey of plasma levels of zinc in sheep in
552
various parts of New Zealand indicated that sheep in some areas had
plasma zinc levels as low as animals reduced in growth by experimental
1064,1204
zinc deficiency. However, no cases of zinc deficiency were
observed. A survey of 600 sheep in Greece showed that 1% of them had
severe symptoms of zinc deficiency, and 60% had milder symptoms probably
1204
associated with other factors (other deficiencies, poor management, etc.).
Zinc or zinc plus manganese supplements were reported to have improved repro-
ductive performance in ewes in Australia although management was apparently
418
also involved.
It is not known whether sheep are more resistant to zinc deficiency
than cattle, or if zinc deficiency in the field merely has not been recog-
nized. Sheep may need less zinc than cattle because deficiencies developed
1204
in sheep on a dietary level of 4 ppm zinc in the same amount of time
1052
in which deficiency symptoms developed in calves on 7-8 ppm dietary zinc.
656
The large amounts of soil ingested by sheep and the ability of sheep
658
ruminal fluid to extract zinc from it may also offer protection.
266
-------
Zinc deficiency in goats has not been studied much. Nine-month-old
male goats on a diet of 4 ppm zinc developed signs of deficiency in about
1127
5 wk. Adult female goats on a diet of 6-7 ppm did not develop signs of
zinc deficiency until they were lactating. Milk production was not affected,
but the zinc content of the milk dropped to approximately 50% of that pro-
569
duced by the controls after a month's lactation. Male kids raised on
the 6-7 ppm diet were severely stunted, but females apparently were not
375,945
affected. Other observations have indicated that the growing intact
males have an increased zinc requirement, but the differences have not been
as large as those reported in this experiment. It would seem that other
1204
factors must have been involved. The survey of ruminants in Greece
mentioned previously found that 2% of the 150 goats examined exhibited
severe symptoms of zinc deficiency.
Swine
Although 14-20 ppm zinc appears to be adequate for pigs on a diet
1479
with casein as the protein source, the requirement on a soy-based diet
1114
(in which the zinc is less available) is 50 ppm. However, soy is not
the only feed that increases the zinc requirement; pigs on a cassava-rice
bran diet supplying 40 ppm zinc displayed symptoms of zinc deficiency,
992
whereas pigs on a corn-soy (20%) diet with 48 ppm zinc developed normally.
Swine receiving a 13% soy meal intake showed no signs of deficiency although
the zinc concentration was only 32-34 ppm. Not surprisingly, neither the
1188
additions of ethylenediaminetetraacetic acid (EDTA) uor pre-
1187
chelated trace minerals to this diet improved weight gain of the pigs.
Addition of low levels of the pre-chelated trace minerals was said to
produce poorer results than the basal diet without the added trace elements;
267
-------
but none of the results were strikingly different and
considerable discrepancy was found between the amount of trace minerals cal-
culated to be in the diet and the actual content determined by analysis.
For the pig in particular, high calcium levels increase the zinc
614
requirement in diets containing large amounts of seed protein.
Rat
99,
Zinc deficiency has been studied extensively in the growing rat.
846,847,1195,1196,1197,1236,1640,1782
A casein diet containing 8 ppm
zinc yielded maximum growth, but 15 ppm was required to maintain bone
1197
and whole body zinc at normal levels.
Humans
Zinc was included in the list of recommended dietary allowances
1119
(RDA) for humans for the first time in 1974, and recommended levels are
given in Table 8-2. These recommendations are based partly on the
average intake of apparently healthy people and partly on balance studies.
In a balance study the amount of zinc ingested in the diet is compared to
that excreted in the urine and feces. Because the studies are cumbersome,
they usually involve small numbers of people for short periods of time on
a limited range of intakes. Such data allow only a crude approximation
of the requirement, but they provide the best information currently
available. Levels similar to those of the RDA's also were arrived at by
1806
a committee of the World Health Organization. Requirements in this
case were based partly on considerations of changes in lean body mass at
1404
various ages.
The adequacy of 15 mg zinc daily for adults is supported by recent
1532 1354
studies in which both adult American men and young New Zealand women
268
-------
TABLE 8-2
Recommended Dietary Allowances For Zinc
Infants
Children
Men
Women
Pregnant
Lactating
Age
(years)
0.0-0.5
0.5-1.0
1-3
4-6
7-10
11-14
15-18
19-22
23-50
51+
11-14
15-18
19-22
23-50
51+
Weight
(kg)
6
9
13
20
30
44
61
67
70
70
44
54
58
58
58
(Ibs)
14
20
28
44
66
97
134
147
154
154
97
119
128
128
128
Height
(cm)
60
71
86
110
135
158
172
172
172
172
155
162
162
162
162
(in)
24
28
34
44
54
63
69
69
69
69
62
65
65
65
65
Zinc
(mg)
3
5
10
10
10
15
15
15
15
15
15
15
15
15
15
20
25
?rt>m Recommended Dietary Allowances.
1119
269
-------
were in balance on a daily intake of approximately 17 mg. Adults have also
504,614,643,1119
been reported to be in balance on lower intakes. A zinc
intake of 4.5 mg in a diet typical of low income groups appeared to be
marginally adequate for 7-9-yr-old girls when additional nitrogen was added
1298
to the diet. Levels of zinc in diets of children from different insti-
1102
tutions varied from 2.7-6.4 mg/day. The allowance for the infant is
based primarily on the estimated zinc intake of an infant receiving breast milk.
This amount may not be enough to maintain bottle-fed infants in positive zinc
balance, because an intake of 1 mg/kg body weight was necessary to keep
infants less than a month old in positive zinc balance. In 3-4-month-old
471
infants, positive balances could be maintained with 0.75 mg. The rapid
decline in the zinc concentration in infants' hair in the U.S. shortly
617
after birth may also be an indicator that zinc intake is marginal.
615,621a
Hair zinc in infants in Thailand was not low after birth, and
hair zinc in infants in England did not decline as much as it did in the
614a
U.S. In addition, supplementing an infant milk formula with zinc
1722
increased the growth of male infants by age 6 mo. However, evidence
from animal studies shows that some change in zinc metabolism takes place
after birth, as plasma zinc levels decreased shortly after birth in both
757 879
rats and pigs. More information is needed on the zinc requirements
of infants and children.
562
Total parenteral nutrition also substantially lowered the plasma
zinc levels in two infants. Trace element formulas for use in intravenous
. ,„ 749,1483a
hyperalimentation solutions for children and adults have been described.
270
-------
Reproduction
Female. Zinc requirements for pregnancy and lactation have not been
extensively studied. In the rat and pig, the zinc level that allowed maxi-
1780,1782
mum growth was also adequate for gestation: 12 ppm in the rat and
65,1607b
20 ppm for swine on a casein diet and 34 ppm for a corn-soy diet.
The requirement for lactation is assumed to be higher than that for gesta-
tion. An adequate zinc intake for gestation in the goat resulted in severe
569
deficiency during lactation.
Little is known about the effect of zinc on reproduction in the
human female except that plasma and hair zinc levels decrease during
616 614
pregnancy and with intake of oral contraceptives. To some extent,
1000
this may simply reflect a redistribution of zinc in the body. Pregnant
teenagers who were given 30 mg zinc daily still maintained lower levels of
plasma zinc during the ninth month of gestation than they had 6 wk after
1439
parturition. Nonetheless, the values were higher than those reported
616
for normally noumished women in the ninth month of pregnancy. Zinc
concentrations in plasma and hair two days after parturition were lower in
women in Iran that had been living in villages or under low socioeconomic
conditions in the city than in those that had been living in higher socio-
1414
economic levels in the city. (But an attempt to correlate decreased
plasma zinc with abnormal outcome of pregnancy in East Harlem was
1515
unsuccessful. ) Several unsuccessful pregnancies have been reported
618
in patients with acrodermatitis enteropathica, a condition in which
very low plasma zinc levels have been reported. It recently has been
1085,1129a,1520a
discovered that the condition responds to zinc therapy.
The possibility of zinc deficiency occurring in human pregnancy
721,1474a,1475
has been of particular concern because zinc deficiency in
757,759,
rats has been shown to cause congenital malformations.
761,761a,1640,1732
To what extent these results are applicable to humans is
271
-------
unknown. When evaluating these results one must remember that extremely
low zinc diets are required to produce the malformations, and that so far
the only reported experiments have been conducted with rats. The results
of the rat studies may not be applicable to other species since vitamin
deficiencies, for example, have been known for a long time to be terato-
1730
genie in the rat; but comparable effects have not been seen in humans.
Similar experiments are difficult to simulate in other animals, and apparently
no experiments have been done on animals with long gestation times and only
one or two offspring. For animals that do not bear large litters, zinc may be
more important to normal mating and maintenance of pregnancy.
Male. A zinc intake somewhat higher than that required for normal
growth may be necessary to support normal reproductive function in the male.
Male sheep receiving a diet containing 17 ppm zinc exhibited reduced testic-
ular weight, low volume of seminal fluid, and an increased percentage of
abnormal sperm, compared to sheep receiving 32 ppm zinc, although food
1642
intake and weight gains for the two groups were the same. Young bulls
developed a zinc-responsive pododermatitis.although steers and heifers were
375
unaffected.
In the rat, the developing testes seem to be particularly susceptible
383,1174,1640,1642
to zinc deficiency. Because the testes of rats with
restricted food intake are not affected, the effect appears to be primarily
383,1642
a result of lack of zinc and not secondary to low food intake.
Changes in the adult male goat after relatively longer periods of zinc defi-
1127
ciency were apparently related more to lowered food consumption.
272
-------
SOURCES OF ZINC IN THE DIET
».
In an average diet, most of the zinc will be supplied with the protein
ingested. Table 8-3 sets forth the approximate zinc content of foods
relatively high in zinc.
TABLE 8-3
Zinc Content of Selected Human Food
mg/100 g (3.5 oz serving")
Oysters 100a
Roast beef 4.0-8.0^
Chicken, dark meat 3.CT7
white meat 1.0
Fish (perch) 2.5b
Liver 4.0
Egg 1.5 (0.7 mg/egg)a
Nuts 3.0a
Wheat germ 13.0
Whole wheat bread 3.0 (1 mg/slice)c
White bread 1.0C
Legumes 1.0—5.0
Milk 0.3/100 ml (1.4 mg/0.47 l)b
1425
a Data from Schlettwein-Gsell and Mommsen-Straub.
b Data from Osis et al.1179
T826
0 Data from Zook et al.
273
-------
One serving of oysters will more than meet the
adult zinc requirement for the day. Two servings of meat will probably
supply approximately half the daily requirement. Nuts, legumes, and whole
grains also contain relatively large amounts of zinc, but the zinc in these
foods may not be as available as the zinc in meats (see discussion of phytate).
The zinc content of foods as published in the literature has been
1425
comprehensively reported. Some additional zinc values of various foods
1179 1826
are listed by Osis; of wheat and wheat products by Zook; and of
1100,
market milk and evaporated milk, infant products, and human milk by Murphy.
1101 1115
Zinc contents of a few forages and animal foods have also been reported.
Representative zinc values for a number of common foods have been compiled by
the Consumer and Food Economics Institute, United States Department of Agrl-
109 3a
culture (USDA). These tables listing zinc content of foods make up
Appendix A.
1179
Since the values for cooked meat and fish are not very different
from what would be expected based on the values given for raw muscle, loss
of zinc in cooking meat appears small. Canned vegetables have been reported
1446
to have 40-80% less zinc than fresh vegetables. Since a negligible pro-
portion of zinc in the diet comes from fruits and vegetables other than
legumes, loss of zinc from this source would be of little consequence. In
addition, the canned and fresh vegetables that were compared in the study
were not from the same lot, so zinc contents may have been different even
before processing. The most serious losses in processing are those that
occur in refined foods such as wheat. For example, whole wheat bread has
1826
three times as much zinc as white bread.
274
-------
Although the concentration of zinc in milk is relatively low, an
adult who drinks a pint (0.47 1) of milk a day can get 107. of the zinc
requirement from this source. Reports of zinc concentration in cows'
1101,1593 1093a
milk varied from 3 to 8 yg/g, with 4 yg/g given as the average.
1771a 1100,1175,1244
Human milk was reported to have the same or less zinc
than cows' milk. It is possible that zinc in human milk is absorbed more
efficiently, however, since plasma zinc levels fell from 60 to 39 yg/100 ml
in a 2-yr-old boy with acrodermatitis enteropathica when he was no longer
619
given human milk. Improvement in patients with acrodermatitis who are
1085
given human milk may, of course, be from some factor other than increased
1107 1639
zinc absorption. Colostrum and partum mare's milk had twice as much
zinc as milk obtained at later times.
Availability
As was mentioned when discussing the requirement for zinc, the amount
of zinc needed in some diets is higher than in others. Presumably this is
caused by a difference in the availability of zinc in these diets. (Some
factors associated with reduced availability of zinc in certain diets are
discussed in the following section on interrelationships between zinc and
oth<=>.r dietary components.) Some work has been done to determine
the best way of measuring the availability of zinc.
Differences in zinc concentration that varied with protein source
were found in tibia of young Japanese quail although the birds' body
634a
weight was not affected. Rats on a moderately low zinc diet also
104
showed a difference in tibia zinc, although again body weight was similar.
103
Tibia zinc increased in rats from 3-16 weeks of age on normal diets.
Total zinc in the femurs of young rats repleted with 12 ppm dietary zinc
275
-------
1076
were reported to be linear. In a similar experiment, however, the zinc
1371
content of bone increased only slightly. Bone zinc concentration
254,1371
decreased rapidly in rats on a low zinc diet. In rats, at least,
bone zinc does not seem to be any better a measure of zinc availability than
is an increase in body weight in rats fed diets in which the amount of zinc
is less than that required for normal growth.
Fortification
To meet the RDA of 15 mg zinc/day, one must eat a fairly high protein
diet. From the data in Table 8-3, one can estimate that an adult who has
an egg and two pieces of whole wheat toast for breakfast, chicken for lunch
and roast beef for supper would consume approximately 12 mg zinc. If meat
is eaten only once a day, with cereals or legumes as the protein source for
other meals, the zinc intake would be significantly less. To raise the zinc
content of low protein diets, fortification of cereal foods with 22 ppm zinc
has been recommended by the Food and Nutrition Board of the National Research
1117
Council. Even with fortification of cereals it would be very difficult
for a woman on a low protein diet to meet the recommended allowance of zinc
for pregnancy and lactation. Even a woman on a high protein diet would have
difficulty meeting the 25 mg requirement for lactation without a zinc
supplement.
zinc.
Institutional diets generally fall below the 15 mg/day standard for
1033,1451
INTERRELATIONS BETWEEN ZINC AND OTHER DIETARY COMPONENTS
Phycace
Since most instances of zinc deficiency in the field have occurred
in diets in which most of the protein was obtained from plant sources
276
-------
(and seeds in particular), zinc from plants is considered to be less
available to monogastric animals and chickens than is zinc derived from
animal protein. This distinction has been attributed to the presence
of large amounts of phytic acid (inositolhexaphosphoric acid)
or phytate in seeds. It has been recognized that soy protein increases
the need for zinc in the diet, presumably because of the protein's phytate
904
content. Other seed meals have the same properties as soy so that their
use also might increase the zinc requirement. Protein concentrate from
mustard or rapeseed which also contained high levels of phytate interfered
with reproduction in female rats when it was fed from the time of breeding
1015a
to parturition. Since serum zinc levels were low, the effects were
attributed to zinc deficiency; but the performance of females receiving
these diets along with increased levels of zinc was not tested. A cassava-
rice bran diet caused zinc deficiency in pigs, although the zinc level in
992
the diet was 45 ppm. The high level of phytate in rice bran—approximately
5%—may have been responsible for the high zinc requirement. (Phytate con-
545
tents of some feed ingredients are supplied by Gontzea and Sutzescu and
1132
Nelson et al. )
The incidence of zinc deficiency in Iranian villagers was attributed
1325
to high levels of phytate in the bread. To demonstrate the effect of
phytate, high levels of phytate in the chemical form or tanok, an unleavened
bread high in naturally occurring phytate, were fed to three men. Zinc
excretion increased in two of the three men, but serum zinc levels were
1328
not consistently lowered. Prasad et al. reported that Iranian men whose
diet consisted almost exclusively of bread and beans had low levels of zinc
in their plasma, red blood cells, and hair, suggestive of zinc deficiency.1276
Incubating zinc-65-labeled high extraction wheat with yeast increased the
1329
solubility of zinc-65 in the wheat.
277
-------
1326
Since phytate phosphorus decreased in yeast-wheat mixtures, part of
the increased availability was probably from hydrolysis of phytate. How-
ever, the difference in solubility of zinc-65 was much greater than could
be accounted for by phytate destruction alone. There are also enzymes in
122,361a,1778
the animal intestine that degrade phytate, but they have not
been studied extensively.
Adding phytate to animal proteins decreased the availability of
their zinc to chicks; the effect was aggravated in the presence of high
1155
levels of dietary calcium. Assay of zinc availability in plant food
1156
ranged from 40-60%, quite a high availability compared to that of other
minerals. Zinc in zinc-65-labeled pea seeds was 75% available even in
1755
mature seeds with a 1.2% phytate content. Hencex factors other than
phytate contribute to the appearance of zinc deficiency in animals on diets
high in plant protein. The high fiber content of these diets may also con-
tribute to reduced zinc availability since fecal zinc loss has been reported
1439
to be significantly correlated with fecal dry solids.
The presence of phytate in the diet has not been considered a
problem in ruminants because the microorganisms in the rumen can degrade
545,1050
phytate. The development of a zinc-responsive pododermatitis in
375
young bulls on a diet containing swede turnip and barley suggests that
this conclusion should be reexamined, however, since the phytate content of
545
this ration could be relatively high. Weight gain of cattle on a soybean
meal ration was improved by adding a combination of trace elements, including
288
zinc, to the ration.
Calcium
Calcium generally does not depress zinc absorption except in the
1155
presence of phytate. Addition of 7.7 g calcium lactate
278
-------
to the diet of 7-9-yr-old girls, for example, did not change the excretion
1298
of zinc, nor did a sixfold increase in the calcium intake (from 200
1532
to 1,300 rag) affect the zinc balance in adult men. Phytate reduced
the rise in serum zinc in normal adults after zinc sulfate dosage,
1216a
but calcium had no additional effect. However, some indication
exists that high levels of calcium may affect zinc absorption when zinc
levels in the diet are marginal. Rats given 1.3% calcium excreted more
zinc-65 in the feces than did controls given 0.6% calcium. The effect
appeared to be at the intestinal level, since the calcium level in the
diet had no effect on retention or distribution of zinc-65 injected intra-
741
peritoneally. The high level of calcium (18%) in the clay ingested by
1510
some Iranians may also have contributed to zinc deficiency in this
population. Because the source of much dietary calcium in this country
is milk, it is interesting that oral administration of lactose and
500
zinc-65 increased the absorption and retention of zinc-65 in rats.
A low calcium diet has been reported to reduce the severity of
zinc deficiency in the pregnant rat, presumably by causing resorption of
762
bone and thereby releasing zinc from it. To support this theory, a
low calcium diet was shown not to alleviate zinc deficiency in pregnant
1599
rats parathyroidectomized to prevent bone resorption. Pigs on a low
zinc and low calcium diet showed no signs of zinc deficiency^ although those
on a high calcium-low zinc diet were severely affected. Bone zinc was
lower in pigs on the low calcium-low zinc diet, suggesting that either mobiliza-
tion of zinc from the bone or failure to deposit zinc in the bone had occurred.
In growing rats on a low calcium diet with a normal amount of zinc, bone zinc
464
increased.
279
-------
Protein and Amino Acids
Net protein utilization of a soy diet by chicks was increased
57
slightly when zinc was increased from a marginal to an adequate level.
Low levels of dietary protein in rats decreased both zinc absorption and
1671
levels of zinc in the liver and small intestine, implying that a
secondary zinc deficiency might be induced in cases of protein malnutrition.
Zinc absorption and retention by ruminants similarly was decreased on a low
1543
protein diet. In the ruminant, the effect was attributed to a lessened
need for zinc because of reduced growth, but the lowered tissue zinc concen-
trations found in the rats on the low protein diet suggests interference in
zinc metabolism.
Addition of 1% histidine or 0.2% histamine
to low zinc diets alleviated leg abnormalities in chicks
and skin lesions in pigs ° without affecting the zinc concentration of the
tissues. Histidine did increase alkaline phosphatase in tibia of chicks
QflS
fed a low zinc-soy protein diet. UJ Feeding a histidine-rich protein to
pigs also alleviated zinc deficiency although here absorption of zinc was
increased. ° Adding arginine to chick diets low in zinc
increased the severity of the leg abnormalities. Even with adequate zinc,
added arginine tended to depress tibia zinc concentration, though not as
markedly as with low levels of zinc. Since many seed proteins are high
in arginine, arginine, as well as phytate, may cause problems when seed
proteins are used in chick diets. 90^ The negative effects may be peculiar
to chicks since their metabolism of arginine differs from that of mammals.
Unlike the experimental animals, humans fed a diet rich in histidine became
zinc-depleted . 673
280
-------
Vitamins
Biotin has also been reported to alleviate somewhat
282
the symptoms of zinc deficiency in the rat. However, the observation
was based on a small sample of rats. In a more extensive study, biotin
levels from 2-50 rag/kg diet had no effect on weight gains or zinc levels in
1201
rats on a low zinc diet. Since folic acid concentration
1781
has been reported to be low in livers of zinc-deficient animals, these
workers also studied the effect of adding 1-5 mg folic acid/kg diet. The
additional folic acid produced no detectable effect. Similarly, additions
of 2, 5, and 10 times the requirement for thiamine, niacin, riboflavin,
pyridoxine, pantothenic acid, or vitamin B,^ failed to alleviate any of the
symptoms of zinc deficiency.
Low levels of vitamin A have been reported in
52,1413
zinc-deficient lambs and rats. -50ja»J--'-1-' Retinol-binding protein
1514
was also lower in serum of zinc-deficient animals. Since vitamin A
levels in the livers of zinc-deficient rats were higher than in ad libitum
1512
controls, it was concluded that zinc is required for normal mobiliza-
tion of vitamin A from the liver. The accumulation of vitamin A in the
liver of the deficient rat was related to the animal's lack of growth, however,
because zinc-supplemented animals with growth restricted by limited food
intake to the same rate of growth as the zinc-deficient animals also had high levels
of vitamin A in the liver and low levels in serum. The low serum level
in lambs may also have resulted from slow growth. Low levels of vitamin A
1356
in serum have been observed in protein-calorie malnutrition.
281
-------
Other Minerals
Since cobalt can replace zinc in vitro in some zinc-dependent
929
enzymes, the possibility of substituting cobalt for some of the in vivo
zinc requirement has been investigated. A preliminary report indicated
716
that this substitution was possible in pigs. In a more recent study with
rats, cobalt reduced the zinc content of liver and serum but was ineffective
in relieving any zinc deficiency symptoms. In the rat, cobalt cannot be
substituted for the zinc required in the diet. Iron and nickel were simi-
larly ineffective in relieving the zinc deficiency symptoms in rats but
848
did alter the concentrations of zinc in some tissues. Nor did Increasing
1194
the copper or manganese levels affect symptoms of zinc deficiency.
1102
Although cadmium normally is neglible in foods, a few studies
1236
have been done of zinc-cadmium interactions in the diet. Zinc or zinc
1047
plus copper in the diet protected against toxicosis from dietary cadmium.
Distribution of injected cadmium was not different in weanling hamsters fed
1045a
a low-zinc diet compared to those fed a stock ration. But the hamsters
on the low-zinc diet were apparently not very zinc-deficient. Injecting
cadmium on day 12 into pregnant rats caused more resorptions in females fed
a low zinc (7 ppm) diet on days 4-12 than in those fed a high zinc (135 ppm)
1211a 448
diet. Cadmium decreased the intestinal uptake of zinc-65 in rats
1352
and calves, but in the deficient rats the transport of zinc to the
carcass was not inhibited. Nor did copper inhibit zinc transport in zinc-
deficient rats although it had some effect on rats with adequate zinc
448 /
levels. Since more orally administered zinc-65 was excreted by copper-
deficient rats than by controls, it has been hypothesized that copper
282
-------
19
facilitates zinc absorption. The difference was small, however, and
could have been caused by differences in growth or food consumption.
There were no differences in zinc uptake and transfer by everted intesti-
1456
nal sacks of copper-deficient rats or the controls. However, binding
155a,156a,156b
of copper and zinc by liver proteins has been shown to be related.
Form of Zinc
Absorption or retention of zinc was about the same in rats, cattle,
and quail whether the intake was in the form of an inorganic salt or incor-
911,1122a,1128,1478
porated in a natural food. The anion associated with the
32
zinc does not affect utilization of zinc, although its solubility in the
155
alimentary tract of sheep varies with pH. Zinc sulfate has been reported
to result in greater weight gains in pigs than did zinc oxide , but
the data are not very convincing since the pigs supplemented with zinc sulfate
did not grow any better than those supplemented with magnesium sulfate
and no zinc.
OCCURRENCE OF ZINC DEFICIENCY
The extent to which zinc deficiency, particularly marginal zinc
deficiency, occurs in man and animals outside the laboratory is largely
unknown. The chief symptoms of zinc deficiency—decreased food intake and
cessation of growth—are not, of course, unique to zinc deficiency. A
test which will distinguish zinc deficiency from other deficiencies and is
suitable for screening large populations of animals has yet to be developed.
Because of the widespread presence of zinc in foodstuffs, zinc deficiency
was long considered not to be a problem of any practical importance.
283
-------
Increased use of supplements such as soybean meal in animal diets, particu-
larly for pigs and chickens, produced demonstrable zinc deficiencies despite
supposedly adequate amounts of zinc in soybeans. Diets for these animals are
1049
now routinely supplemented with zinc. Only a few severe cases of
1050
apparent zinc deficiency in cattle have been reported, but unrecognized
cases of mild deficiency may be extant, particularly in areas where the zinc
content of forages is low. Crested wheat grass (Agropyron cristatum) in
125
northern Nevada was reported to have a content of 13 ppm zinc, less than
that required by calves on a purified diet to maintain normal serum zinc
levels. Zinc levels as low as 6 ppm have been found in some grasses (Kubota,
unpublished data). Animals grazing on such grasses might well suffer from
zinc deficiency.
It is now known that zinc deficiency may occur in humans, particularly
in people who derive much of their protein from plant sources. A recent
617
report of apparent zinc deficiency in middle-class American children
indicates that the problem is more widespread than had been assumed. Forty-
nine percent of children selected for short stature in a group of Project
1723
Head Start children had hair low in zinc. Similarly, about half the
girls from low income families in another study had somewhat depressed zinc
1217
levels in their hair, although no correlation with growth seemed to
exist.* Hair zinc was low in Iranian and Egyptian 2^8»1 hypogonadal
patients with retarded growth.
ASSESSING ZINC STATUS IN ANIMALS
Because signs of a marginal zinc deficiency generally are nonspecific,
some other test of zinc status is needed. Attempts to correlate zinc
*See Burch et al.201a
284
-------
concentration in blood, hair, or urine with zinc status have met with limited
success, as have endeavors to correlate zinc status with enzymes or metabolites
in blood or urine. There have been recent attempts to correlate zinc status
with taste acuity and salivary zinc levels. The problems involved in using
these measures are discussed below.
Plasma Zinc
Although red blood cells contain significantly more zinc than do plasma
or serum, zinc in the red blood cell appears to be firmly bound and shows
little change in deficiency. Plasma or serum zinc is more variable and does
reflect changes in the zinc status of the individual. Serum or plasma zinc
in rats, for example, fell significantly after only 1-2 days on a deficient
1199,1773
diet, and it remained low throughout the period of low zinc intake.
199
This pattern was also true in humans. Low plasma zinc levels have been
linked to poor growth and symptoms of zinc deficiency in cattle and sheep,
although it was pointed out that at least two low serum zinc levels obtained
on separate occasions would provide more conclusive evidence than a single
1064
determination. Unfortunately, many other conditions are responsible for
reduced plasma zinc levels, including pregnancy, various diseases, and
614
stress. Food intake also affects serum zinc level. The plasma zinc levels
ftfif) 82T
of zinc-adequate quail and pigs and zinc-deficient chicks dropped when
they were refed protein-containing diets after fasting. An oral protein
1235
supplement lowered serum zinc in healthy adults. Administration of zinc
sulfate to fasting subjects increased serum zinc levels more than did
3,1216a
administering zinc with a meal.
Measurement of plasma zinc levels after oral dosing with zinc has
been suggested as a better method for determining zinc deficiency than
the usual measurement of plasma zinc, as rats on a zinc-deficient diet
had higher levels of zinc in plasma after dosing with zinc than did
285
-------
1774
those that had been on the control diet. The situation is
consistent with the observation that everted jejuaal sacs (intesti-
nal segments) from zinc-deficient rats took up more zinc than did those of
850,1456
fully fed controls. Other suggestions have been made that zinc
1048
absorption is increased in zinc deficiency, but the evidence is not con-
clusive. Uptake by animals on restricted intake was also altered, however,
suggesting that any animal receiving less than a normal food intake might
also show a higher level of plasma zinc after dosing. Thus it is unlikely
that determining plasma zinc levels after oral dosing offers any significant
advantage.
In summary, fairly severe zinc deficiency would be reflected in a
low plasma zinc concentration, but marginal deficiencies are unlikely to
be diagnosed by this index. For example, in cases of total starvation,
plasma zinc levels were not lowered although body zinc stores were being
1532
depleted, indicated by excretion of zinc in the urine.
Hair Zinc
Because hair samples are easy to collect and store, their use for
504,614
estimates of zinc status has been looked into by several investigators.
Various washing procedures were used to remove surface contamination, sweat,
and body oil from the hair. One study of such procedures concluded that
none of them satisfactorily overcame the effects of the cosmetic treatments
701
commonly used on human hair. With care, however, reliable estimates
1525
of zinc content of human hair probably can be obtained, although it
may be wise to exclude samples subjected to the more severe treatments
286
-------
such as permanent waving, bleaching, and dyeing. Use of pubic hair has
been suggested in cases where values from scalp hair might be unreliable
because zinc concentrations in scalp and pubic hair of pregnant women have
89
been found to be the same.
It is necessary to discern how useful the zinc content of the
hair is in determining zinc status. The zinc content of hair is a
reflection of chronic zinc status over the time the hair has been growing,
because hair zinc does not exchange with the body zinc pool. Hence it is
no surprise that no correlation was found between the zinc concentrations
999
in plasma and hair of Iranian village children. The lack of correlation
between plasma and hair zinc was demonstrated more strikingly by an aero-
dermatitis enteropathica patient whose plasma zinc levels were very low
(less than 0.4 yg/ml), but whose hair zinc level was in the normal range.
The normal hair zinc level may be related to the decreased hair growth in
1037a
these patients. An extensive study of hair zinc in young rats on
varied levels of zinc intake showed that a diet deficient enough to prevent
weight gain also prevented hair growth, and therefore zinc concentration
1198
did not change substantially. A level of dietary zinc that was still
suboptimal, yet which permitted some growth, resulted in increasing hair
1198
growth and reduced zinc concentration in the hair. Lack of correlation
855
between plasma and hair zinc was also reported in a Panamanian population,
atif.
although hair zinc in the females varied with geographic location.
617,1235b
Zinc concentration in human hair is also reported to vary with age.
Hair zinc content in female goats was suggested as a good indicator
of zinc status because concentrations were decreased in animals on the
deficient diet, but the concentration only was measured when the goats were
287
-------
569
severely deficient. Zinc concentration was not correlated with the
1150
color of bovine hair in samples collected from field locations, but
39 3a
it has been reported to vary with the season.
Fairly severe zinc deficiencies in animals, particularly over a
long period of time, would probably be reflected in a low hair zinc concen-
tration. Marginal deficiencies are unlikely to be diagnosed in this manner
617
although a study of hair zinc in middle class American children indicated
that the zinc level of hair might be a useful measure in detecting deficiency
in humans.
Urinary Zinc*
Although zinc is largely excreted in the feces—with only a small
amount excreted in urine—some indication exists that, at least in man,
urinary zinc may be decreased in states of deficiency. Urinary zinc has
not been measured extensively in animals, although urinary zinc excretion
was studied in rats after EDTA administration to determine whether this
896
technique might be a way of assessing zinc stores. Such a procedure
would be useful primarily for laboratory studies.
Zinc Enzymes
The existence in zinc-deficient animals of an inactive zinc-dependent
enzyme that could be activated by the in vitro addition of zinc would provide
an excellent assay for zinc deficiency. For example, a mutant of the bac-
terium Escherichia coli grown in low zinc media produces an inactive form
of the enzyme alkaline phosphatase, which can be activated if the enzyme is
602
incubated with zinc. The existence of this type of enzyme in blood
would be obviously advantageous to diagnosis of zinc deficiency. Unfortunately,
such an enzyme has not been found in animal tissues.
*See also Chapters 7 and 10.
288
-------
Zinc-dependent enzymes in blood are of the most interest for their
potential in assessing zinc status. Alkaline phosphatase is one of the
enzymes that has been measured extensively in young, zinc-deficient
animals. The enzyme appears to be consistently reduced in serum of zinc-
11,349,350,1146,1285
deficient pigs compared to controls, but in most cases
the controls have grown more than the deficient animals. However, an
instance of zinc deficiency in pigs was reported in which serum alkaline
201b
phosphatase was not decreased. The results with rat serum have been
742,849,1370
more variable and not always different from controls on a
restricted food intake, suggesting that decreased growth influenced the
decreased activity. Since alkaline phosphatase in serum comes primarily
1765
from bone and liver, slight differences in growth rate may significantly
affect the amount of the enzyme in serum. However, in a 22-yr-old aero-
dermatitis enteropathica patient presumably not in an active stage of
growth, the level of serum alkaline phosphatase rose parallel to the
increase in serum zinc when zinc treatment was applied. Addition of zinc
1129a
to serum in vitro did not increase the activity. Alkaline phosphatase
was also low in serum of a boy who appeared to be zinc-deficient by other
615 1363
criteria, but not in Iranian boys with low plasma zinc levels.
Calves with hereditary zinc deficiency were observed to have only slight
alterations in alkaline phosphatase activity, although serum zinc levels
decreased to 0.4 yg/ml. Symptoms of deficiency appeared 2-3 wk after the
881a
serum zinc fell to that level. Since the activity of alkaline phosphatase
can be altered by other conditions which interfere with normal metabolism
of bone or liver and because low levels are usually found along with low
289
-------
levels of serum zinc, serum alkaline phosphatase is of no greater diagnos-
tic value in determining possible zinc deficiency than the measurement of
serum or plasma zinc level.
1370
Other zinc enzymes, such as lactic and malic dehydrogenase, have
been measured in serum of zinc-deficient animals, but their enzymatic
activity was not affected. Carbonic anhydrase activity was lower in
deficient animals if the increase in numbers of erythrocytes from hemocon-
1374
centration in the deficient animals was taken into account. Since
restricted-feed intake can also cause hemoconcentration, decreased activity
was probably related to the reduced food consumption of the deficient
742
animals as was, in fact, found in another study.
Therefore^no presently known enzyme is a reliable measure of zinc
status.
Metabolites
Concentrations of various metabolites in blood and/or urine have
been reported to be changed in zinc deficiency, but none of the changes
appear to be observed consistently in deficient animals and are therefore
of little value as a diagnostic tool. Metabolites that have been reported
1783
to appear in increased concentrations are uric acid in blood
730 729 731 733
and urine; hydroxyproline, sulfate, taurine,
and alanine in urine; and taurine and free
fatty acids in plasma. Urinary ascorbic acid has been
93
reported as decreased. All these metabolites have been observed in
young animals, and it is quite possible that they are nonspecific changes
related to the slow rate of growth caused by the zinc deficiency. Reduced
290
-------
plasma protein in severely zinc-deficient rats was not related to decreased
1370,1598
growth or food intake; however, neither serum protein nor albumin
levels were lowered along with the very low serum zinc level of patients
1129a
with acrodermatitis enteropathica. Plasma protein would be affected
by many conditions other than zinc deficiency.
Taste Acuity and Salivary Zinc
Because some conditions of abnormal taste acuity have been zinc-
responsive, both taste acuity and salivary zinc have been measured in
connection with studies of possible zinc deficiency. Taste acuity was
617
impaired in children with poor growth and low levels of hair zinc, but
not in a young woman with acrodermatitis enteropathica, a zinc-responsive
1129a
disease in which plasma zinc levels are very low. Zinc secretion in
parotid saliva was lower in children who had low levels of hair zinc than
615
in normal children. The data are insufficient at present to indicate
whether either of these measures is a reliable and specific indicator of
zinc status. In any event, measurement of taste acuity would not be a
practical means of screening large populations. Salivary zinc could be
measured on animals as well as people. The large variation in zinc con-
centration of saliva collected from the same individual at different
1536
times suggests that there may be substantial problems in the use of
this criterion. Nonetheless, in view of the striking changes that have
973 79
been reported in the tongue of zinc-deficient sheep and monkeys,
1180 1146
in the mouth and esophagus of pigs, and in zinc-deficient
461,1102a 382
rats (including the fetal rat), measurement of salivary
zinc may be worth pursuing.
291
-------
METABOLIC CONSEQUENCES OF ZINC DEFICIENCY
Although many studies have examined in some detail the effects of
zinc deficiency on various systems in the animal, the critical roles of
zinc in these systems still are not known. Zinc is contained in a number
of enzymes, but the effect of zinc deficiency on the activity of these
enzymes is variable. Also, decreased activity of the known zinc-dependent
enzymes seems not to be a major factor in the disturbances observed in zinc
deficiency. Because some of the earliest effects of zinc deficiency are
reduced food intake and cessation of growth, it is very difficult to
distinguish between a primary effect of a lack of zinc on the animal and
secondary effects based on decreased food consumption and halted growth.
In theory, it is possible to control for the decreased food consump-
tion by feeding a zinc-adequate animal the same amount of food that the
zinc-deficient animal eats. The problem, however, is not so simple as it
might appear, since the deficient animal has a cyclical pattern of
270,271,1782
eating, will gain weight less quickly than a zinc-adequate
animal on the same amount of food, and will consume its food over a 24-h
period, whereas the pair-fed animal consumes its food in a much shorter
time.. To control for all these effects is difficult and often is not
done. Since food intake can influence metabolism in ways and over time
periods that are not obvious, differences between deficient animals and
their "controls" must be interpreted very cautiously.
DNA. RNA, AND PROTEIN SYNTHESIS
Because of the rapid effect of a lack of zinc on growth, a number
of studies have been conducted on DNA, RNA, and protein synthesis during
292
-------
zinc deficiency. Concentrations of DNA, UNA, and protein were not remarkably
467,493,494,775,878,1172,1280,1284,1285
different in the deficient animal, but
the concentration of these compounds may be less important than their meta-
bolic activity. The activity of these compounds has been estimated by
measuring the incorporation of radioactive precursors into DNA, KNA, or
protein. In order for such measurements to be reliable, however, the radio-
active precursor must have the same relative concentration in both deficient
and adequate animals. Since concentrations of free amlno acids were higher
732 567
in both skin and plasma of deficient animals than in controls, and
729a
since compounds such as thymldine and several amlno
733a,1607a
acids were metabolized to carbon dioxide to a greater
extent, the radioactive precursor may constitute a smaller proportion of the
precursor pool in the deficient animal than it does in the control.
Different concentrations of the radioactive precursor may be brought
about by differences in enzymes needed to enable the radioactive compound
to enter the precursor pool, as well as by differences in the concentration
of metabolites in the cell. For example, incorporation of radioactive
thymidine into the pool of DNA precursors in the cell depends on the
activity of thymidine kinase, an enzyme whose activity has been shown to
722 351a
vary diurnally, which is related somewhat to food consumption. The
different eating pattern of the zinc-deficient animal therefore could
make a difference in the time at which thymidine kinase activity occurs
and consequently in the time at which peak Incorporation of thymidine would
be observed. In rats on a low zinc diet, the peak of thymidine incorpora-
tion which follows partial hepatectomy was delayed compared to the controls,
but there was little difference in the amount of incorporation at the
293
-------
397,408
respective peaks. Others have reported differences in thytnidine incor-
poration between zinc-deficient animals and controls in which the magnitude
732,1002,1550
of the differences changed with time. Thymidine kinase activity,
as measured by thymidine incorporation into DNA in vitro, was reduced in
connective tissue from animals on a low zinc diet; the concentration of zinc
used in the assay systems differed, however, and the animals on the low zinc
1280
diet also were receiving 2.5 times as much phytate as the controls.
DNA
Thymidine incorporation was reduced in liver, kidney, and spleen in
young rats before growth and food consumption were affected by the low zinc
1779
diet. The sensitivity of these tissues to the early effects of the
deficiency was no doubt related to the rapid turnover of zinc in these
920
tissues. Thymidine incorporation was also reduced in 12-day-old embryos
of female rats given a deficient diet. However, mitoses in neuroepithelium
1584
were increased. In rapidly differentiating tissue such as embryonic
tissue, tissues from animals fed a deficient diet may not be in the same
stage of development as the controls although the time of gestation is
the same. Different stages of development at the same time of gestation
were observed between fetuses from folic acid-deficient females and fetuses
788a
from controls.
Thymidine incorporation was decreased in zinc-deficient rats more
736
than in controls after wounding the skin or implanting sponges under
1280 966
it, but not after an esophageal wound. Interpretation of studies
of incorporation after surgery is complicated by the hormonal changes
which accompany stress, since adrenal hormones, for example, affect thy-
542a
midine incorporation.
294
-------
In contrast to reduced mltotic activity observed in most tissues
383,461,1180
of the zinc-deficient animal, mltotic activity in the esophagus
and buccal mucosa was stimulated. The stimulation may be peculiar to
383,461
zinc deficiency because it did not occur in pair-fed controls.
Furthermore, a single dose of zinc in zinc-deficient rats reduced cell
division in the esophagus but increased the mitotic indices of epidermis
461 23
and liver. In addition to increased mitotic activity, thymidine incor-
24 1037
poration, and dry weight of cells in buccal mucosa of deficient rats,
epithelial thickness was increased.1180 The increase in epithelial thickness
occurred before the increase in mitotic activity and was thought to come from
interference with the normal shedding mechanism, perhaps from an increased
glycoprotein coating of keratinizing cells. Apparent interference with the
shedding mechanism was also observed in esophagi from 18-day-old rats nursed
by zinc-deficient dams and in pigs on a low zinc-high calcium diet.
Alterations in salivary and intestinal mucus have been reported in zinc-
. -. . t fc 1309
deficient rats.
Cell cultures also are affected by a lack of zinc. Results in these
systems are clearer, because differences from food intake and hormonal
changes are eliminated. Addition of a zinc chelating agent inhibited
931a,931b
incorporation of thymidine into DNA in primary rabbit kidney cells,
1785 • 269 1377,1378,1379
in human and porcine lymphocytes, and in chick embryo cells.
However, the zinc.requirement was lost rapidly by rabbit cells, and continu-
ously cultivated cells such as HeLa cells and L cells were also resistant
931a
to EDTA. Chick embryo cells retained the requirement for zinc but
1377
became quite resistant to EDTA after infection with Rous sarcoma virus.
295
-------
The effect of EDTA did not appear to be caused by damage to the cell membrane
1379
since cell movement and uptake of glucose were normal.
1494,1542 59,59a,
Although good evidence exists that both DNA- and RNA-
59b,1255
dependent DNA polymerases from some organisms are zinc-dependent
enzymes, EDTA probably does not affect enzymes directly involved in DNA syn-
thesis. A delay in the addition of EDTA decreased its effect in several
268,269,515,931a,931b
systems, and in continuously replicating cells EDTA
1377
was able to reduce thymidine incorporation only after a time lag. Both
these results are interpreted to mean that zinc is required for a critical
step preceding the onset of DNA synthesis in the cell, and that cells past
268,931b,1377
this step when EDTA is added will replicate normally.
Studies of microorganisms grown in low-zinc media are also consistent
with this hypothesis. Although growth of Mycobacterium smegmatis was
inhibited by a low-zinc medium, the activity of DNA polymerase in vitro was
201a
not affected. As has been pointed out, an enzyme which is zinc-dependent
in one species need not be so in another; however, the activity of the poly-
merase from Mycobacterium smegmatis was inhibited by a zinc chelating agent
1790
and was presumably a zinc-requiring enzyme. Euglena gracilis also
452
appeared to synthesize DNA normally, but it failed to divide. The DNA
synthesized could, of course, have been abnormal. In the zinc-deficient
Mycobacterium smegmatis, increased DNAase activity was suggested to be
related to increased need for DNA-repairing enzymes to compensate for
synthesis of abnormal DNA. DNA synthesis was not affected in Rhodotorula
292
gracilis.
In summary, the increased mitotic activity in esophagus, buccal
mucosa, wounded skin, and liver after partial hepatectomy, as well as the
296
-------
synthesis of connective tissue in response to an irritant, provide ample
evidence that zinc-deficient animals can synthesize DNA. The lack of growth
in zinc-deficient animals seems to be due to an effect on factors initiating
DNA synthesis rather than to any defect in the process once it has begun.
RNA
In general, incorporation of precursors into RNA is less affected
by a lack of zinc than is incorporation of precursors into DNA. Levels of
EDTA that almost completely inhibited thymidine incorporation into DNA had
little effect on incorporation of precursors into RNA in rabbit kidney
931a,931b 1785 268,269
cells, human, or porcine lymphocytes, and perfused
515
liver. EDTA reduced the incorporation of precursors into RNA in chick
embryo cells to 60% of normal, whereas incorporation of thymidine into DNA
1377
was almost completely inhibited. Incorporation of uridine
11 79
into brains of both zinc-deficient rats and controls was identical. '
Synthesis of RNA was not affected in Mycobacterium smegmatis grown in a medium
635a 292
low in zinc, but it was decreased in Rhodotorula gracilis and
452
Euglena gracilis.
1462
RNA polymerase has been reported to contain zinc, but its activity
was less inhibited by EDTA the later the EDTA was added to the incubation
mixture. This phenomenan has been interpreted to mean that zinc is required
for the initiation of RNA chains but not for their elongation. Because the
enzyme does contain zinc, its activity has been measured in zinc-deficient
1603 493
animals. Its activity was reduced in liver and brain from rat pups
nursed by zinc-deficient dams. However, the pups continued to grow despite
1603
the reduced polymerase in the liver; and the activity in brain, although
493
reduced at 6 days, was no longer reduced at 16 days. RNA polymerase
122a,1345b
activity in rat liver nuclei was also affected by starvation,
297
-------
but the relation between RNA synthesis in the animal and the measurement of
RNA polymerase in the test tube is not clear. Zinc stimulated orotate
incorporation in vivo without any detectable effect on RNA polymerase activity
1757 1757,1757a
measured in vitro. Zinc inhibited polymerase activity in vitro.
The increase in material in the monosome region of the sedimentation profiles
493 122b 1144a
of zinc-deficient rat pups, starved pigeons, and mice could repre-
sent defective RNA synthesis, but it may simply reflect the decreased protein
synthesis that is apt to follow decreased growth. Neither additional poly-
1345b 122b
merase nor initiation factors increased RNA synthesis in preparations
from starved animals, further suggesting that reduced polymerase activity
was the result rather than the cause of decreased growth.
RNAase activity was increased in testes of rats that had been on a
1284 1522a
deficient diet for 5 or 11 wk. Since the testes of growing rats
are particularly susceptible to zinc deficiency, increased RNAase activity
at these relatively late stages of the deficiency may also have been a result
rather than a cause of the problem. RNAase activity was not different in
brains of rats nursed by females on a low zinc diet, although, brain growth
494
itself was reduced.
Protein Synthesis
Nor did EDTA have much effect on incorporation of labeled amino acids
931a 1377
into protein in rabbit kidney cells, chick embryo cells, and perfused
515
rat liver. Since EDTA only affected amino acid incorporation into protein
269,1785
in lymphocytes at later stages of the cell cycle (but not initially),
EDTA presumably did not affect protein synthesis per se. A zinc-deficient
diet also had little effect on incorporation of labeled amino acids into
408,1172 533,731,735
protein in young rats. Incorporation of several amino acids,
298
-------
1003
including selenium analogs, was decreased in skin from deficient rats but
not in other tissues. Incorporation of radioactive amino acids into liver,
heart, and kidney was reduced in rat pups nursed by females on a low zinc
496
diet. Aside from the differences in incorporation in skin and in nursing
533
pups (perhaps partly caused by differences in amino acid metabolism by
the deficient animals), little indication has been found that a zinc-deficient
diet interferes with protein synthesis. The deficient rat was able to syn-
thesize new tissue in response to an irritant; and although the total amount
synthesized was less than that of controls, the amount synthesized relative
to weight gain was as high in the deficient animal as in the ad libitum
467,1284
control. Deficient rats also synthesized protein in response to
1218 267a
infection and after partial hepatectomy.
INTERMEDIARY METABOLISM
Glucose
Because the concentration of zinc in the pancreas decreases markedly
when zinc is deficient, and because the addition of zinc to insulin prepara-
tions prolongs their action, several studies have been made on the effect of
zinc deficiency on glucose tolerance. Results have conflicted,
somewhat accounted for in that glucose tolerance in rats appears to be
closely related to the food consumption of the animals in the 24 h preceding
479
the pretrial fast. In studies where differences in glucose tolerance
139,740
existed, comparisons were apparently made to ad libitum-fed controls.
139,1309a
Differences in glucose levels in fasted animals, glucose toler-
663,1221,1308 139,740 139,1221,
ance, insulin levels in blood before and during
1308,1309a 740,1221
glucose stimulation, and insulin levels in the pancreas
299
-------
between deficient animals and pair-fed controls generally have been small.
Blood glucose was higher in deficient animals that were given a second dose
663,1309a
of glucose within 2 h after the first dose. Pancreatic tissue from
deficient animals released less insulin in vitro in response to glucose
stimulation than did tissue from ad libitum-fed controls but the effect was
740
obtained only at a high glucose level. Insulin secretion in response to
659a
glucose stimulation is also affected by starvation. Resistance to
1309a
insulin coma was greater in zinc-deficient rats, but resistance to
1594a
insulin coma is also affected by starvation.
Fatty Acid
High levels of free fatty acid were observed in fasted rats during
1308
the first 3-4 wk after they were placed on a zinc-deficient diet. How-
ever, values approached those of the control group with time. Zinc stimulated
130 7b
the glucose uptake by rat adipose tissue in vitro, although the magnitude
1307c
of the effect varied with the species of rat. Glucose uptake by adipose
tissue from zinc-deficient rats was less than that from rats given a zinc
130 7b
supplement.
Amino Acid
Amino acid metabolism, particularly that of the sulfur-containing
amino acids, has been reported to be disturbed in zinc deficiency. Urinary
excretion of both total sulfur and sulfate from sulfur-labeled methionine,
733 42 733
cystine, and taurine, as well as sodium sulfate was increased in
deficient rats.
The urinary excretion of taurine increased in one investigation*2 but not in
another.567 Hydroxyproline excretion also increased in young zinc-deficient
300
-------
729
rats. Levels of free amino acids in rat skin were increased in deficient
732
animals. Activities of the amino acid-catabolizing enzymes arginase and
tryptophan pyrrolase were also increased, although the activities of serine
730
and threonine dehydratases were not affected. In these experiments, the
weight of the pair-fed animals exceeded that of the zinc-deficient ones.
Therefore, the differences observed may have been due to differences in
growth rate of the two groups of animals. Both food restriction and zinc
156,364,1342
deficiency affect zinc-binding proteins in rat liver.
ZINC DEFICIENCY AND BONE FORMATION
Bone abnormalities in zinc-deficient chicks and rat embryos have led
to studies on the effects of zinc deficiency on calcium metabolism and bone
99,100,102,221,755,949,951,1139,1146,1760
growth. Kinetic studies of calcium-
45 in zinc-deficient rats showed that turnover of calcium was slower in
deficient than in pair-fed animals and that pool size was smaller. Absorption
755
and excretion were not different, however. Sulfate uptake and turnover in
1139
growing regions of bone were impaired in the zinc-deficient chick. In
vitro sulfate uptake by skin from pigs given a moderately low zinc diet was
1610
not impaired. There was no difference in collagen or mucopolysaccharides
of zinc-deficient chicks, although the hexosamine content of primary spongiosa
1139
was somewhat higher. Hexosamine content in bone from deficient rats was
949
not different from the controls, although hexosamines in saliva were
1309
affected. No change occurred in the hexosamine content of skin from
1610
pigs given a moderately low zinc diet. Many changes that were observed
99,102,949,1146
in bone from deficient animals were similar to those observed
301
-------
with decreased feed intake. Zinc and manganese appeared to interact in
1306a
otolith formation in mice.
Zinc deficiency in chicks causes a swollen hock condition which
resembles arthritis. Certain anti-arthritic compounds relieved the con-
715 1323
dition while others did not. How the effective antidotes acted was
unclear, although in some cases sulfate uptake in areas of bone growth
increased. However, sulfate uptake increased in adequate as well as
395
deficient chicks.
Increased dietary histidine also relieved the swollen hock condition and
was postulated to function by increasing histamine concentration, but his-
tamine in tissues of deficient chicks was not different from controls, nor
1324
was it increased by the feeding of histidine. Histidine did increase
904,905
alkaline phosphatase activity measured in the entire tibia, although
histolbgic studies did not show any difference in the distribution of alka-
1760
line phosphatase in the epiphyseal plate. Some changes in bone histology
1760
were apparent before alkaline phosphatase would be produced, so that
deranged alkaline phosphatase may be a symptom rather than a cause of
problems in bone formation.
ZINC DEFICIENCY AND ZINC-RELATED ENZYMES
As with the DNA polymerases, the demonstration of a zinc-requirement
by an enzyme in an organism has led to its measurement in zinc-deficient
animals, although the enzyme tn the animal may not have been shown to be
zinc-dependent. Again, it must be emphasized that the metal requirement for
the same enzyme in different species need not be the same. A comparison of
302
-------
activities of various enzymes in zinc-deficient animals in relation to the
controls is given in Tables 8-4 and 8-5. The tables only are intended to
indicate whether a difference was observed between the activity of the
enzymes in the tissues of zinc-deficient animals and those of controls;
and, if values were given for both pair-fed and ad libitum controls,
whether the enzymatic activity in the tissues from the pair-fed animal
varied in the same direction as that of the deficient one. If a change
is shown in the deficient animal without a notation of change in the
pair-fed control, it may mean that information on pair-fed controls was
not given. Also, increased enzymatic activity in the pair-fed control
relative to the activity in the ad libitum will not be reflected in the
tables unless the enzyme activity was increased in the deficient animal.
It should also be pointed out that different authors expressed
their results differently. Ratings in the table were based on activity/
unit weight rather than on total activity, since the total would obviously
be influenced by the smaller size of the tissues in the deficient animals.
Nonetheless, activity/unit DNA was significantly different in some tissues
1285
in one study, whereas activity/unit protein was not. Also, in one of
the studies, pair-fed controls were not used because in a previous study
775
they had become "zinc-deficient." It seems more likely that the
animals were biotin-deficient, as diets containing egg white increase the
requirement for biotin. Since egg white protein is widely used in studies
of zinc deficiency because of its low zinc content, increased biotin require-
ment should be taken into consideration although most investigators do add
an excess of biotin to such diets.
303
-------
As shown in Table 8-4, the activities of presumably zinc-related
enzymes in the deficient animal have varied from tissue to tissue and from
experiment to experiment. In Table 8-5 it can be seen that other enzymes
not known as zinc-dependent enzymes were also altered in the zinc-deficient
animal.
Alkaline phosphatase in bone was consistently lowered in
the zinc-deficient animal, as was alcohol dehydrogenase in liver and
carboxypeptidase in the pancreas. The volume of pancreatic secretion was
1565
also smaller in zinc-deficient pigs. The decrease in a-mannosidase in
useful as
serum may be/ an indicator of zinc status because the activity of the enzyme
in liver was reported to decrease in zinc-deficient rats and be activated
by zinc in vitro, whereas the activity of the liver enzyme from zinc-adequate
1213a
rats was only slightly stimulated. As mentioned, this type of effect
would make a useful assay of zinc deficiency.
Little correlation has been found between zinc concentration in
tissues and the activity of the presumed zinc-dependent enzymes. Concen-
tration of zinc in the liver was not lowered in the deficient animal in two
1279,1372
of the studies, whereas liver alcohol dehydrogenase was
lowered. The activity of alcohol dehydrogenase correlated well with the
concentration of zinc if food were restricted but not if animals were fed
1330
normally.
Because enzymatic activity is not correlated with zinc in the tissues and
most enzymes are not activated by zinc in in vitro assays, it is possible
that enzymatic activity is low in the zinc-deficient animal because the
animal has stopped growing and consequently needs less enzyme. In any
304
-------
event, of the few enzyme activities that have been consistently lowered, none
seem likely to account for the symptoms (see Table 8-1) observed in the
deficient animal.
Alkaline phosphatase has been one of the most extensively studied
enzymes in zinc-deficient animals. Determination of alkaline phosphatase
activity in animals is complicated by the fact that enzymes from different
783a
tissues varied in pH optima, activity with different substrates, and
361a,361b,783a,948a
activity in presence of magnesium. Within a single
tissue several isoenzymes may exist,; as many as five have been reported in
36 la
intestinal mucosa. Only one isoenzyme—with a pH optimum at 10.5—'was
783a,1326a
activated by adding zinc in vitro, although intestinal homogenates
1778
have not been activated by in vitro addition of zinc. In cases where
alkaline phosphatase was activated, it happened to the control as well as
the deficient enzyme. Chromatography of homogenates from deficient and
control animals indicated that the same isoenzymes were present in both
animals. although some properties of the enzyme from the deficient animal
1326a
differed from those found in the control. The apparent differences,
however, may have been a consequence of differences in degree of purifica-
tion of the preparations.
ZINC AND REPRODUCTION
Another major effect of zinc deficiency is its interference with the
growth or function of the reproductive organs. This is another area in
which it is extremely difficult to distinguish between the effect of a lack
of zinc per se and the effect of reduced food intake. Underfeeding is well
£ 907a
known to interfere with reproductive function.
305
-------
Female
220,327,
In the rat, inadequate zinc severely affects reproduction.
759,761,1582,1640,1732
Effects include difficult parturition, congenitally
malformed young, failure to maintain pregnancy, and cessation of estrous cycles.
Congenital malformation or difficult parturition occurred after consuming a
46,756,1731 1582
low zinc diet or large amounts of a zinc chelate for just a
few days during a critical period of gestation. Development of preimplanta-
tion eggs was abnormal in females fed a deficient diet for only the first
758
few days of pregnancy; however, offspring born after dams received a deficient
756
diet for days zero to five had only a slight incidence of malformations.
Work with the teratogenic effects of trypan blue also
suggest that there is some mechanism for repairing early damage to the
621b
eggs.
Administration of zinc late in pregnancy prevented stress at parturi-
47
tion, but survival of offspring from dams fed a zinc-deficient diet during
the second week of gestation was poor although zinc nutriture after that was
757
adequate. Survival of pigs born to gilts on a low zinc diet for approxi-
1744a
mately the last third of pregnancy was also poor, and abnormalities in
1202
bone development were observed in the fetuses. Feeding the rat a zinc-
1102a
deficient diet during lactation caused severe zinc deficiency in the pups.
Insufficient zinc during pregnancy also interferes with reproduction
44
in the rabbit. Studies of zinc metabolism in zinc-adequate rabbits have
shown that zinc accumulation in the rabbit endometrium coincided with the
958a
blastocyst phase of embryonic development. About that time, luteal
tissue also showed a marked increase in specific activity of injected zinc-65,
306
-------
lOlla
although the zinc concentration in the tissue did not change. Turn-
over of zinc-65 in the endometrium decreased in the pregnant, pseudopregnant,
and superovulated rabbit. In placental tissues, zinc transport varied with
gestational age, and fetal placenta exchanged zinc with blood plasma four times
1011 757 1202
faster than maternal placenta. Fetal plasma in rats, pigs* and
929a
sheep and goats is higher in zinc concentration than maternal plasma.
The concentration of zinc in human amniotic fluid at the end of pregnancy
453a
has been correlated with the birth weight of the newborn.
Zinc in vitro has been shown to potentiate contractile responses of
354b
rat uteri to submaximal doses of acetylcholine.
Since uterine contractions have not been measured in the zinc-deficient preg-
nant animal, it is not known whether there is anything abnormal about the
contractions that could be related to the difficulty at parturition. Zinc
concentration in the deficient uterus was the same as in the controls. But
such a condition does not indicate of itself that contractility was normal
since the potentiation in vitro did not correlate with the zinc content of
354b
the uterus. Adding zinc to an endometrial homogenate has been reported
43 5a
to increase binding of g-estradiol to protein.
Most studies dealing with the relation of zinc to reproduction in
the female have been done with animals that bear large litters, such as the
rat, rabbit, and pig. Whether the work with these animals applies to other
species, particularly those that bear single offspring during relatively
long gestation times, is unknown. For animals that do not have large litters,
zinc may be more important for normal mating and maintenance of pregnancy
than for normal delivery.
307
-------
After consuming a low zinc diet for 2-3 wk before mating, rats failed
45
to maintain pregnancy. Consumption of a low zinc diet for longer periods
45,544
brought about cessation of estrous cycling. The effects on the estrous
cycle in particular are probably caused by reduced food consumption although
measurements of reproductive hormones indicated that effects of zinc deficiency
544
and restricted food intake were not identical.
614 1000
Oral contraceptives lower plasma zinc levels in humans and rats.
The percentage of zinc-65 in other tissues of rats given high levels of estro-
gen was increased somewhat; however, the dose administered signficantly
depressed weight gains in the growing females.
Male
The relation of zinc to the male reproductive function has received
considerable attention because of the high concentration of zinc present in
1640
the prostate gland and semen. The major portion of the zinc in semen
comes from the prostate gland, but its function is not known. Zinc content
1376b
of the epididymis is also high and may function in retarding oxidation
224
of sulfhydryl groups in the sperm. But sperm motility is not
789a
affected by large differences in zinc content, although sulfhydryl
224
groups in sperm flagellum have been thought to be involved in motility.
Zinc also has been postulated to protect the integrity of sperm since
42 7a
oxygen consumption by sperm in vitro was raised in the absence of zinc.
Increased oxygen consumption by sperm is associated with metabolic disorganiza-
tion. Zinc uptake by sperm was affected by factors other than the concentra-
tion of zinc in semen. a' ' c Interference with normal zinc metabolism
was postulated to be a mechanism of the antifertility agent a-chlorohydrin
since it increased radioactive zinc in regions of the
308
-------
575a
testes in which zinc concentration in sperm is normally reduced. Repro-
ductive hormones have also been shown to alter the distribution of zinc in
1640
male reproductive organs.
Insufficient zinc in growing male rats severely affected testicular
Development. The effect was apparently independent of the low food intake
since pair-weight rats had normal size testes whereas those of deficient rats
383
were considerably smaller than normal. Histologic studies of a few zinc-
deficient rats suggested that the earliest effect of zinc deficiency on the
testes was the inhibition of the transformation of round spermatids into
1174
elongated ones. Refceding the animals with zinc for 15 days subsequent
383
to the 28-day depletion period restored the histologic appearance to normal.
However, low food intake does affect the testes in some ways. Alpha"
mannosidase activity, for example, has been reported to decrease in epididy-
mal tissue of zinc-deficient rats. But enzymatic activity was restored in
castrated animals by injecting them with testosterone, yet zinc
1517a
concentration was not affected. The enzyme may have been affected by
the decreased testosterone levels associated with low feed intake.
Libido of adult male goats on a zinc-deficient diet has been reported
1127
to be reduced, but the goats may not have had a simple zinc deficiency.
EFFECT OF ZINC ON FOOD INTAKE
For both animals and man, one of the first effects of a low zinc
diet is decreased food consumption. In rats, food consumption also
became increasingly variable and cyclic.270'1782 Intake varied less
if the rats were given a 5% rather than a 20% protein diet. Moreover,
zinc-depleted rats ate more of a zinc-supplemented diet only when it
contained protein.270 Examination of the amino acid pattern
309
-------
in plasma and urine of animals fed low zinc diets containing either 24% or
6% protein indicated that plasma amino acids—particularly nonessential
ones—were elevated on the low zinc diets. Tyrosine
was the only amino acid that fluctuated significantly in relation to daily
food intake. Animals on a low protein diet may not be functionally zinc-
deficient, because growth is restricted by the lack of protein and the
increased catabolism of body protein may release enough zinc to satisfy
temporary maintenance requirements. Mean plasma zinc concentrations of rats
on a low protein diet were appreciably higher than in those fed 20%
protein diets.1376a
Zinc-deficient rats drank more of an acetic acid solution if they pre-
276
viously had received zinc in the solution. Zinc-deficient rats also drank
more sodium chloride, hydrochloric acid, and quinine sulfate solutions than did
pair-fed controls, and took in a greater total volume of fluid, including tastant
1004
solutions and water. Zinc-depleted chicks also chose a zinc-supplemented diet
in preference to a low zinc diet, after 5 days, however, the controls
also selected the supplemented diet. Turkey poults have been reported to
select a zinc-supplemented diet, "a but apparently because of a position
preference. Food consumption rose within a few hours after zinc repletion
11 QS
in young zinc-deficient rats, but it increased more slowly in zinc-
deficient pregnant rats who were transferred to a zinc-adequate diet.
EFFECTS OF ZINC DEFICIENCY ON WOUND HEALING AND TUMOR GROWTH IN ANIMALS
Reports of beneficial effects of zinc on wound healing in man have
led to studies of wound healing in both zinc-deficient and zinc-adequate
animals. Wound healing was impaired in zinc-deficient animals; *'
differences were more apt to occur during later stages of healing.1264 -jhe
level of zinc in wound fluid and wound plasma has been reported to increase
310
-------
930 931
in rats and dogs in early stages after wounding. However, the increases
observed were highly variable, as were the effects of different stresses on
1745 427,571,
serum zinc in dairy cows. Additional zinc generally has no effect
1307,1408 902,1034,1082
or only slight effect on wound healing in zinc-adequate
animals. Zinc sulfate did not improve collagen biosynthesis or fibroblast
1737b
proliferation in human tissue culture. Zinc seemed to have some protective
effect on the livers of rats injected with carbon tetrachloride427 and
on the thyroid glands of pigs given methylthiouracil, a thyroid
685
antagonist. The effectiveness of zinc in stimulating lymphocyte transfor-
98,1243 1307a
mation and increasing thymus weight suggests that excess zinc
might stimulate the immune system; but except for the effects mentioned,
beneficial effects of zinc treatments on zinc-adequate animals have not been
demonstrated.
Zinc deficiency has been reported to decrease the growth rate of several
tumors and increase the survival times of animals inoculated with them. If
Walker 256 carcinosarcoma (a rapidly dividing solid tumor) were injected
intramuscularly in young male rats that had been on a zinc-deficient diet for
8 days, the number of takes was reduced; survival time was increased and
381
tumor size reduced in those animals in which tumors did grow compared to
1018 381
either pair-fed or paired-weight controls. Growth of a variant of
the Walker 256 carcinosarcoma was reduced in young rats that had been on a
1235a
low zinc diet since weaning. Similar results occurred in mice with the
380
slower growing Lewis lung carcinoma. Survival times after injection with
various leukemia tumors were not as markedly affected by zinc deficiency as
81,380
sarcoma tumors and attempts to increase the effectiveness of the zinc
81
deficiency in treating leukemia tumors were unsuccessful.
311
-------
It is important to realize that in the cases in which tumor growth
was reduced in zinc deficiency, the animals were already deficient when
the tumor tissue was injected. Transferring animals to a low zinc diet
after a tumor is established has not been shown to reduce tumor growth.
Because of the hyperkeratosis and parakeratosis of the esophagus
that occur in zinc-deficient animals, a relationship between esophageal
cancer and zinc deficiency had been postulated. A marginal intake (7 ppm)
of zinc did not, however, affect the incidence of nitrosamine-induced
1683
esophageal carcinogenesis in rats.
ZINC DEFICIENCY AND BEHAVIOR
Zinc-deficient rats have performed poorly on different behavioral
220,601,950,1406
tests. Exploratory activity was decreased and performance
219
impaired in rats given a deficient diet for 7 wk after weaning. However,
the rats were apparently ill at the time of testing. Behavior was also
affected in offspring of rats given a deficient diet during gestation or
lactation although the offspring themselves were given a zinc-adequate
diet. Offspring from two rats given a deficient diet during lactation made
950
more errors on an elevated maze than did offspring from two pair-fed rats.
Offspring from successive litters of females fed a deficient diet through-
out gestation and lactation were less active in an open field test than
were controls. The number of deficient rats tested, however, was very
small and represented few litters, since not many offspring from the deficient
220
females survived to weaning.
Male offspring of 5 dams on a deficient diet only during days 15-20 of
gestation avoided shock less successfully than did offspring from 5 pair-fed
312
-------
rats. Offspring from both deficient and pair-fed animals, however, were less
active and extinguished the response faster than did offspring from ad libitum
601,1406
females. Since female offspring of zinc-deficient dams tolerated
600
shock less well than did those from pair-fed animals, poorer performance
of the males in the shock avoidance test also may have been due to a reduced
tolerance for shock. Females from pair-fed dams were less resistant to
shock than were those from ad libitum females.
These studies suggest that consumption of a zinc-deficient diet during
pregnancy or lactation has a residual effect on the offspring. Such an effect
737
has also been attributed to protein deficiency during gestation^ but the
number of pregnant females used in the zinc studies was small. In addition,
the effect of zinc may have resulted from differences in the care given the
46
pup by the dam, the suckling ability of the pup and hence its ability to
1816a
stimulate lactation, or the in utero nutrition of the fetus,although
the dams received the same amount of food.
Whatever the source of the difference, one must be very cautious in
extrapolating these results to other species since the brain of the rat is
relatively immature at birth and may be more susceptible to nutritional
insults. Brain maturation, as measured by the activity of the myelln-
associated enzyme, 2', 3'-cyclic nucleotide 3'-phosphohydrolase (CNP), was
1303
not different for rats suckling females on a zinc-deficient diet.
Studies of DNA, RNA, and protein in brains of animals whose mothers were
subjected to gestational or lactational zinc deficiencies have revealed
493,494
relatively little difference in the concentration of these compounds.
313
-------
TABLE 8-U
Effects of Zinc Deficiency on Tinaue Levels of ZIne-Relnted Enzyiron
Eeryma Blood
Alcohol
dehydrogenase'1
Bone Muscle Lung Heart Thyous
Decreasec(pl8) Ho change4
Decrease*
Spleen
Aldolase
Alkaline
Carbonic
anhydxase
Bo change
Decrease**
°(pl8)
Decrease^185
Decrease""18'
Decrease0'1'18'
Bo changeMple)
Decreasen(pl8)
DecreaseP(pl8)
Bo change1
Decrease
Decrease*1
Decrease
Decrease*'*1'*' So change*
Decreasek(chl<*'
DecreaSe^(chiclt)
Decrease?(pie)
Decreasetf(pl8)
Decrease
Decrease*^
Decrease
Bo change Bo change
;•
Bo change Decrease*
Bo change Bo change
Bo change*
Decrease*
Bo
CerboxypeptidAse
Gl'jtamlc
dehydrase
Bo change
Bo change
Lactic
dehydrogenaae
Malic dehydro-
genase
o>nannosldase
Increase*
Bo change
Bo change
a
Bo change
Decrease
D8crea.e
Bo change
a(pl«)
Decrease*1 Ho change Decrease* Bo change*
Bo change
Decrease
Decrease Bo change"
Decrease
314
-------
TABLE 8-4 (Cont'd.)
. Tissues
Bi«y»e
dehydrogenise*1
Aldolase
Alkaline
phosphausa •
Carbonic
anhydrase
Cartojypeptidase
Clutanic
dehydrase
lactic
dehydrogenase
Halle dehydro-
genase
d-nannosidase
Urer Pancreas
Decrease0'*18'
Decrease*
Decrease'
Decrease0'1118'
Ho change*
increase^"1*' Decrease*
Decrea,e* Decrease**
a(n\a\
Decrease011118'
Ho change1
Ho change to change4
Decrease0'*18'
Decrease*
Decrease*
Ho change
Ho change6'*18'
Increase*'
Decrease* Ho change1
Ho change0'*18' Ho change?'*18'
Decrease*'
Ho changer
Decrease"
Stonach Intestine
Decrease** DecreaseJ'(chlck'
Decrease* Ho change*'"*11*'
Ko change*'*1*'
Decrease
Decrease*0
Decrease"
Decrease0
Decrease Ho change1
Decrease* Decrease™
Ho change
Ho change Decrease*
Decrease
Kidney Testes Brain
Decrease01*8' Decrease*
Decrease*
Decrease0'*18' Ho change*
Ho change*
Decrease0'*18' So change*'chlck)
Ho change*'*16'
Decrease*
Decrease1
Decrease"
Ko change Ho change1
Decrease"
Ko change
Decrease"
Decrease* Ho change9
lib change* Ho change*
Ho change9
Decrease" HO change"
315
-------
TABLE 8-4 (Cont'd.)
Conditions:
Decrease - Deficient lover than either ad libitum or restricted-fed control,.
,-, . *.««.<«* lover than ad libitum controls; difference not necessarily
Decrease* - Restricted-fed as well as deficient lover tnan au
significant. , ,
No change - Deficient not different from ad libitum controls.
Increase - Deficient greater than ad libitum controls.
increase- - Restricted-fed as veil as deficient greater than ad libitum controls; difference not necessari*
significant.
^Studies done in the rat unless othervise noted.
Derived from Burch et al.
T pOc
^Derived from Prasad et al.
1 *vrfi
^Derived from Roth and Kirchgessner.
1279
^Derived from Prasad and Oberleas.
1372
•fDerived from Both and Kirchgessner.
^Derived from Agergaard and Palludan.
t, 9Ql»
Derived from Lease.
7^2
^Derived from Huber and Gershoff.
^ X«j *_
^Derived from Davies and Motzok.
Derived from Lease.
^Derived from Borrdic et. al.
775
^Derived from Iqbal.
3^9
"Derived from Dahmer et al.
°Derived from Iftbal.783*
^Derived from Dahmer et_ al.
^Derived from Williams.1778
> VYT^
Derived from Roth and Kirchgessner. '
'Derived from Roth and Kirchgessner. '
• 137U
Derived from Roth and Kirchgessner.
"Derived from Prohaska et al.1303
WDerived from Swenerton et al.1
^Derived from Patel and Ryman.1 13a
^Derived from Roth and Kirchgessner.
-316
-------
TABLE 8-5
Enzyme Blood Heart
Alanine and Mpareate fc(plg)
aminotransf erases" Ho change v* "*'
6-amlnolevulinate
dehydratase Decrease
Arginase
Glutamate, oxalate, and
pyruvate transaminases
Isocitrate dehydrogenase No change^plg^ Ho changeZ'^pig^
Leucine aminopeptidase Ho change pis'
Omithine transcarbamylase
Serins and threonine
dehydratases
Sorbitol dehydrogenase
Succinlc dehydrogenase
Tryptophan pyrrolase
Liver
Ho change0
Increase*
Decrease*^18*
Increase*6
Decrease*2^18)
No change1'
Ho change
Ho changeMpi6)
DecreaseMpi8)
Ho change
Decrease*6
Ho change^
Increase*
Pancreas Kidney
Ho change^ Ho change*^18'
Ho change?(pi8)
Ho change^
DecreaseZ'(pi8)
No chang^(pie)
Ho change'
Testes
Increase^
Ho change^
Conditions:
Decrease - Deficient lower than either ad libitum or restricted-fed controls.
Decrease* - Restricted-fed as well as deficient lower than ad libitum controls; difference not necessarily significant.
Ho change - Deficient not different from ad libitum controls.
Increase - Deficient greater than ad libitum controls.
Increase* - Restricted-fed as well as deficient greater than ad libitum controls; difference not necessarily significant.
Studies done In the rat unless otherwise noted.
^Derived from Burch et al.201*
"Derived from 7inelli et aO.
"Wived from Hsu and Anthony.730
"Derived from Roth and Kirchgessner.1372
^Derived from Prasad and Oberleas.1279
^Derived from Prasad et al^.1285
317
-------
CONCLUSIONS
1. A fairly constant supply of zinc is required by all species.
Signs of zinc deficiency develop rather quickly when zinc intake is low
because very little zinc is stored in the body in a readily available form.
Although a relatively large amount of zinc is stored in skin, muscle, and bone,
this zinc is not available to the animal unless those tissues are being
catabolized.
2. The extent to which marginal zinc deficiency occurs in humans and animals
is unknown.
Without a specific test for zinc deficiency, it is difficult to determine
a marginal occurrence of it. The high protein diet typical of middle and upper
income Americans will probably supply close to the 15 mg zinc that is the RDA
for adults. Therefore, deficiencies would not be expected in these individuals.
However,.people eating low protein diets are apt to receive significantly less
zinc. To what extent marginal zinc deficiency may occur under these conditions
is at present unknown. It is important to realize that the RDA and nutrient
requirements of animals are simply the best estimates that can be made with the
limited amount of data available. They will undoubtedly be modified as more is
known about the-factors affecting zinc requirements and zinc availability.
3. The aspects of cellular metabolism that are primarily affected by zinc
deficiency are not known.
Although zinc is contained in a number of enzymes, the effect of zinc
deficiency on enzymatic activity is varied, and decreased activity of known
zinc-dependent enzymes does not seem to be a major factor in disturbances
observed in zinc deficiency. Many of the effects observed may be a function of
low food intake or lack of growth.
318
-------
4. The amount of zinc occurring naturally in food is unlikely to be a health
hazard.
Ingested zinc is relatively nontoxic if the diet contains adequate copper
and iron; most animals appear to tolerate levels of the order of a milligram
per gram. Because of the possible interference with copper and iron metabolism,
animals with high zinc intakes should be monitored for signs of anemia.
319
-------
CHAPTER 9
ZINC IN METALLOPRDOEENS
SPECIFIC ZINC ENZYMES
The study of the specific biologic role of zinc has centered on its
function in a number of zinc metalloenzymes. Several of their
structures are now known on a molecular level; moreover, the three-dimen-
sional structure as determined by X-ray diffraction has allowed sane precise
descriptions of the role of zinc in the mechanisms of enzymatic action. The
characteristics of the zinc metalloenzymes which have been well documented
and the reactions catalyzed are listed in Table 9-1. Table 9-2 lists zinc-
containing enzymes for which extensive studies on the function of zinc have
not been carried out. Some enzymes, such as DNA and KNA polymerase, have
been reported to contain zinc when isolated, but the metal's particular
functional role in these enzymes has not been delineated.
The carboxypeptidases (part of the exocrine secretion of the pancreas)
are important in the C-terminal hydrolyses of peptides and proteins in the
mammalian gut. Additional proteases from bacteria have been discovered to
be zinc metalloenzymes. The most thoroughly studied example is thermolysin,
found in an extremely heat-stable enzyme containing both calcium and zinc.990
The Ca(II) appears to participate in the heat stability rather than in the
activity, whereas Zn(II) is essential to activity and appears to function much
as the Zn(II) ion in carboxypeptidase A. The specificity of thermolysin is analo-
gous to that of the latter enzyme, except that it is an endopeptidase and
does not require the terminal free-carboxyl on the substrate. The neutral
protease from Bacillus subtilis has been less extensively studied, but it is
i
known that Zn(II) is essential for activity.
320
-------
TABLE 9-1
VfeH-Characterized Zinc Metalloenzymes—
NJ
Qizyme
Carboxypeptidase A
Carboxypeptidase B
Thernolysin
Neutral protease
Leucine aninopeptidase
Carbonic anhydrase
Molecular Zinc per b
Weight Molecule Reaction Catalyzed- Reference
W f\ H
T B 1 T
34,500 1 J2>-C-N-CH2-C -f- ^-C-COO- Hartsuck and
0 H CH2 (BGP£ Lipscaiib6**
Jl \j H[ -
JJ-C-S-CHj-C 4* O-C-000-(HPIA]F
P
34,500 1 0-C-^I-CH2-C 4* S-j-COO~ Polk484
H R?=Arg or Lys
^C-w-CHo-C 4" O-C-COO""
R= Arg or Lys
O H H O
rt TJ |-N tj O f\
^J rl ^J ^^ E| JJ1 ^J ^ £ 1H
44,700 1-2 ^-O-C-B-CHj-C •+* N-C-nC-NHj Tsuru et al .
Tyr
54,000 2 H^-leucine 4* NB^tor peptide) Hiimelhoch706
^ _ 942
30,000 1 CO + H2O«M~) ?* H CO (HCO ) Lindskog et al .
RC-OR1 + H20g — ^RCOOH + R'CH
RCHD + HjO^ss^Rtfl
-------
TftBIE 9-1 (Continued)
N)
N>
Enzyme
Alkaline phosphatase
Aldolase (yeast)
Alcohol dehydrogenase
Superoxide disntutase
Aspartate transcarbamylase
6 catalytic
6 regulatory
Molecular
Weight
80,000
80,000
80,000
32,000
300,000
subunits
33,000
subunits
17,000
Zinc per
Molecule Reaction Catalyzed-
2 ROPO 4- HO ^ ' '"* ROH 4- HPO "
3 2 * 4
RSP03 4- H20 ;ps± RSH 4- HPO4=
ROPO2S= 4- H20 x — -H3H 4- HPO3S=
RNP03 4- H20^=^RNH24- HPO4=
2 Fructose-l,6-diP5*dihydroxy-
acetone-P 4- glyceraldehyde-3P
4; 2 NAD CH J3LOH 4- NAD5B±CH CHO 4- NADH 4- H+
•J & J
2; 2 Cu 0~ 4- 0~ 4- 2K+SgHH 00 4- 0^
2 2 ^22 2
6 H2N-C-O'*'P 4- Asp^Carbanyl-Asp + HPO^
Reference
1322
Reid and Wilson
724
Horecker et al.
833
Keleti
508
Fridovich
1772
Wiley et al.
—See also Chlebowski and Coleman.
—Benzoylglycylphenylalanlne
= bond hydrolyzed
-Hippurylphenyllactic acid
-------
TABLE 9-2
Enzymes Reported to Contain Zinc
Enzyme
RNA polyroerase
RNA. polymerase
DMA polymerase
Nucleotide pyrophosphatase
5 ' -Nucleotidase
Cyclic phosphodiesterase
U3
nPhosphomannose iscmerase
Phosphoglucomutase
o-D-Mannosidase
&-Lactamase
Protease
5'-Menosine monophosphate
aminohydrolase
Collagenase
Neutral protease
Dipeptidase
Phospholipase C
Source
Escherichia coli
T_ phage
Escherichia coli
Rat liver
Escherichia coli
Escherichia coli
Yeast
Yeast
Jack bean
(Canavalia ensiformis)
Bacillus cereus
Snake venom
Rat muscle
Clostridium
histolyticum
Bacillus cereus
Porcine kidney
Bacillus cereus
Molecular
Weight
370,000
110,000
109,000
52,000
450,000
26,000
290,000
105,000
47,000
Zinc Atoms
per Molecule Oof actor
2
2-4
2
0
1
0
1
1
0
0
1? 2 Ca
2
0
0
1
Reference
1462
Scrutton et al.
Coleman (unpublished)
1494
Slater et al.
316
Carder and Lowry
410
Dvorak and Heppel
410
Dvorak and Heppel
553
Gracy and Noltmann
708
Hirose et al.
1518
Snaith and Lewy
1388
Sabath and Finland
1713
Wagner and Prescott
1819
Zielke and Suelter
1470
Seifter et al.
458
Peder and Garrett
225
Campbell et al.
118T
Ottolenghi
-------
TABLE 9-2 (Continued)
Enzyme
Source
Molecular
Weight
Zinc Atoms
per Molecule
Cofactor
Reference
Dipeptidase
a-Atnylase
D-glyoeraldehyde-3P-
dehydrogenase
Tactic dehydrogenase
Malic dehydrogenase
Glutamic dehydrogenase
o3ranscarboxylase
£
Pyruvate carboxylase
Mercaptopyruvate sulfur
transferase
5-Aminolevulinic acid
dehydratase
Mouse ascites tutor
Bacillus subtilis
Porcine muscle
Rabbit muscle
Bovine heart
Bovine liver
Proteus shermanii
Yeast
Escherichia coli
Rhodano.se (sulfur transferase) Bovine liver
87,000
50,000
Bovine liver
40,000
1,000,000
670,000
600,000
23,800
37,000
260,000
1
0.3 Ca
3
0
1
2-4
4; 2 Co
4
1
2
3 NAD
NAD
0 NAD
6 Biotin
4 Biotin
654
Hayman and Patterson
1665
Vallee and Wacker
832
Keleti
1665
Vallee and Wacker
639
Harrison
9
Adelstein and Vallee
1148
Northrup and Wood
463
Scrutton et al.
1649
Vachek and Wood
1702
Volini et al.
1787
Wilson et al.
%towever, definitive data relating zinc to the structure or function
of tirese enzymes are not available.
-------
Carbonic anhydrase plays an iitportant role in catalyzing the rate of
attainment of equilibrium between carbon dioxide and the bicarbonate anion,
a process involved in many important cell functions in the mammal;
carbon dioxide diffusion at the alveoli, hydrochloric acid secretion
in the stomach (bicarbonate anion exchange for chlorine anion) and acid
secretion in the renal tubule. Carbonic anhydrase may also influence the
eg|li1^hrium in the leaves of green plants. Aside from carbon
dioxide transport itself, it is possible that catalysis of the CO^sft HCO~
equilibrium by carbonic anhydrase is involved in the general scheme of
anion transport and the maintenance of electroneutrality.
Alkaline phosphatase is widely distributed in nature. It is present
in bacterial cells like Escherichia coli (where it is located in the peri-
plastic space between cell wall and cell membrane), as well as in mammalian
cells such as the cells of the intestinal endothelium, and cells along the
line of newly calcifying bone. Both the bacterial and mammalian enzymes
are zinc metal loenzymes. The known function of alkaline phosphatase is the
nonspecific hydrolysis of phosphate monoesters, but many of the enzyme's
functions in the complex environment of the cell are not fully understood
and it also may participate in phosphate transport and phosphate transfer
reactions. Bone phosphatase is essential for bone calcification, as human
mutants lacking the enzyme activity do not calcify their bones. The enzyme
does participate in transferase activity in the test tube with suitable
acceptors such as Tris buffer and ethanolamine. Acceptors must
carry an amino function on a carbon adjacent to a carbon carrying an alcoholic
hydroxyl.
325
-------
The physiologic dinplications of alcohol dehydrogenase are obvious be-
cause of its central position in the metabolic pathway for the exogenous
(and possibly endogenous) metabolite, ethanol.
Superoxide dismutase, a zinc-copper protein also known as erytii-
rocuprein, has become the subject of recent interest because of the theory
organisms carrying out
that its presence is necessary for / aerobic metabolism by protecting the
organism from damage by the superoxide radical. This radical is the
product of a number of enzyme-catalyzed reactions involving oxygen: for
example, the oxidation of xanthine to urate by xanthine oxidase.
Radicals as products are likely to occur since reactions involving oxygen
usually proceed in one-electron steps involving free-radical intermediates
because of the spin restriction applying to the addition of two electrons
to molecular oxygen. The enzyme catalyzes the dismutation of the superoxide
radical rapidly, limited only by the difusion of substrate.
Aldolase is a central enzyme in the pathway of anaerobic glycolysis and
in organisms such as yeast, Zn(II) is an essential component although the
mammalian muscle enzyme is not a metalloenzyme. In this enzyme, a specific
lysyl -NIL,"1" group is the substitute for the zinc function.
Aspartate transcarbamylase is important in pyrimidine biosynthesis
and the bacterial enzyme is the only well-characterized zinc metalloenzyme
in which zinc directly and exclusively affects enzyme structure, since the
metal is contained in the regulatory subunit of this enzyme and the catalytic
subunit is active in the absence of metal. Zinc is involved indirectly in the
regulation of enzymatic activity, since its presence is required for the
proper association of regulatory and catalytic subunits to form the function-
ing hexamers of this allosteric enzyme.
326
-------
ROLE OF ZINC AT THE MOLECULAR LEVEL
The protonatable amino acid side chains of proteins have been the obvious
candidates as ligands to the zinc in metalloproteins. These chemical groups
include the carboxyl groups of aspartyl and glutamyl residues, the N-terminal
^-amino • groups of the
peptide chain, the I -amino groups of lysyl residues, the imidazole
nitrogens of histidyl residues -- the most likely candidates considering their
chemical properties of pKfl and formation constants — the phenolic hydroxyl
groups of tyrosyl residues, the sulfhydryl
groups of cysteinyl residues and the guanidino groups of
arginyl residues. The -NH-group of the peptide bond has been shown in certain
model systems, especially Zn(II)-peptide complexes, to form a coordinate bond
with loss of its proton. Thus this group is also a potential donor in the
formation of metal-protein complexes.
Of this group, only three have been identified as Zn(II) ligands in
Zn(II) metalloproteins: the imidasole nitrogen, the <-carboxyl group of
glutamic acid and the sulfhydryl group of cysteine. Carboxypeptidase A
coordinates the Zn(II) through the
imidazole nitrogens of two histidyl residues and the V1-carboxyl group of a
644
glutamyl residue. Thermolysin, a bacterial endopeptidase similar to
990
Carboxypeptidase A, has the same three ligands to the Zn(II) ion. In both
proteins, the fourth coordination position of a distorted tetrahedron is
occupied by a water molecule. The Zn(II) of carbonic anhydrase is coordinated
to the imidazole nitrogens of three histidyl residues. A fourth coordination
site is also open to solvent and occupied by a water molecule or an hydroxyl
ion.275a'303'9^2 Indirect evidence from nitrogen nuclear hyperfine splitting
of the electron spin resonance signal of the Cu(II) derivative of alkaline
327
-------
phosphatase suggests the presence of three nitrogen nuclei as ligands to
275a
the metal ion (probably imidazole nitrogens). Spectral data and amino
acid analysis have established that Zn(II) binds to sulfhydryl groups in the
protein metallothionein from mammalian kidney. ' Zn(II) also binds to
275a
sulfhydryl groups in alcohol dehydrogenase.
To draw conclusions about the nature of coordination sites to be found
at the active centers of metalloenzymes, the large body of information that
exists for small coordination complexes must be studied. In these systems,
however, the ligands are flexible and free to assume bond lengths and bond
angles dictated by the electron distribution in the d-orbitals of the metal
ion. Bond lengths and bond angles in enzyme coordination sites, however, may
be dictated as much by the stereochemistry of relatively inflexible protein
ligands as the electron distribution in the metal ion's orbitals. Such
inflexibility may be one of the advantages of a large protein molecule as a
catalyst in that a particularly unstable configuration of amino acid residues
(considered as an isolated structure) may be maintained at one site in the
molecule (e.g., the active site) by the stabilizing effect of structure in the
rest of the molecule. Such "strain" could radically affect the stability of
particular complexes and affect the reactivity of the metal ion toward a sub-
strate in a mixed complex formed during catalysis.
Highly purified zinc metalloenzymes for which the relative affinities of
several first transition and IIB metal ions for the metal -binding site have
been determined show an extraordinary preference for Zn(II) compared to Co(II),
Ni(II) i and Cu(II) , although the sites in most cases are made up of nitrogen
and oxygen ligands, which have greater affinity for the other three ions in
simple systems. Model systems would suggest that all Zn(II) binding sites
should contain sulfur ligands, since sulfur is known to have a particularly
high affinity for Zn(II) , a property that does not occur uniformly in nature.
328
-------
The Zn(II) ion does not appear to be necessary for the synthesis of the
apoprotein nor does it have to be present to assure the presence of the high
affinity Zn(II) binding site. The Zn(II) enzyme, alkaline phosphatase, has
been thoroughly examined in this regard. Escherichia coli grown in a zinc-
free medium derepressed for alkaline phosphatase will produce a zinc-free inactive
apoalkaline phosphatase with physicochemical characteristics identical to the
apoenzyme produced by removal of the metal with chelating agents.2753 The
apoenzyme synthesized by the Zn(II)-deficient organisms can be completely
reactivated by the addition of Zn(II). The organism can synthesize enzymes
containing Co(II), Cu(II), and Cd(II) if these metal ions are added to the zinc-
free medium, but Zn(II) will overpower the other three metals at relative con-
centrations much below those expected for competititve binding to model coor-
dination complexes. This dominance could be related to the fact that most of
the protein binding sites have highly distorted ligand geometry, a function of
the three-dimensional structure of proteins. The first transition metals with
unfilled d-shells generally derive significant stabilization of their complexes
from the ligand field stabilization energy, provided the geometric arrange-
ment of the ligands is such that low energy orbitals exist that can be occupied
and higher energy orbitals are left vacant or half-occupied. If the ligands
are prevented from assuming the ideal geometry that maximizes the splitting of
the orbital energy levels (as may occur because of restraints in the protein
structure), the metal complex will be destabilized in relation to the usual
models. However, zinc with a filled d-shell and no field stabilization energy
from the ligand may accommodate the distorted geometry without destabiliza-
tion resulting.
It has been speculated that zinc operates in its capacity as a Lewis acid
and withdraws electrons from certain groups of the substrate in mixed enzyme-
metal-substrate complexes. The open coordination sites in carboxypeptidase A,
329
-------
thermolysin, and carbonic anhydrase clearly suggest this possibility. Only in
the case of carboxypeptidase A has the structure of an enzyme-metal-substrate
complex been determined. The X-ray structure of the crystalline glycyl-L-
tyrosine complex with carboxypeptidase A shows the Zn(II) ion to be coordinated
to the carbonyl oxygen of the susceptible peptide bond which is placed in the
644
position of the coordinated water molecule in the free enzyme. Mechanisms
proposed for carbonic anhydrase suggest coordination between Zn(II) and the
oxygen of an active hydroxyl ion in hydration or the oxygen of bicarbonate in
the dehydration reaction, but no direct proof is available. a Whether a
simple Lewis acid function applies in all instances or even in all hydrolysis
or hydration reactions catalyzed by zinc metalloenzymes is not clear. Structural
roles for Zn(II) have been suggested, but evidence has been difficult to obtain.
In aspartate transcarbamylase the Zn(II) is not at the active site, but seems
1772
to participate primarily in the structure of the regulatory subunit.
EVOLUTION OF ZINC ENZYMES
If one of the major functions of zinc in the biosphere is its fundamental
catalytic role in metalloenzynes, then an important question is, how uni-
versal among the various phyla is it that the catalysts for these particular
reactions are zinc metal loproteins? Is the evolution of a given zinc
metalloenzyme a relatively restricted event, or have these zinc macrono-
lecular catalysts existed over long periods of evolution? A great amount of
phylogenetic information has been gathered on seme of these zinc enzymes fron
isolating and characterizing the enzymes (listed in Table 9-1) from plants, and
primitive and higher animals all the way up to primates, including man. The
sources from which the enzymes were obtained, the molecular weight, and the
zinc content are given in Table 9-3. Usually if an enzyme is a zinc enzyme
330
-------
in one species, it will contain zinc in another; for inany enzymes the zinc
is maintained in the enzyme from bacteria to man. Thus in an enzyme like
carbonic anhydrase, the active site of zinc is present in the enzymes of the
elasmbbranch, a species that split off from the mainstream of animal evolution
approximately 350 million years before the evolution of mammals. Therefore
some zinc enzymes are under little evolutionary pressure to change.
However, there are notable exceptions. The aldolase of yeast is a zinc
enzyme, whereas the aldolase of the mammalian muscle is definitely not. An
c-amino group of a lysyl residue of the protein provides
a complete reaction through the formation of a ketimine intermediate without
the assistance of a metal ion. Another notable substitution occurs within
the enzyme superoxide dismutase: the bacterial enzyme is a manganese metallo-
protein, but the mammalian enzyme is a zinc-copper protein.
OTHER POSSIBLE BIOLOGIC FUNCTIONS OF ZINC
Whereas the enzymatic functions of zinc outlined above form a specific
and fundamental basis for the biologic function of zinc, a number of complex
1652
syndromes have been described in animals as a result of zinc deficiency.
In many of these syndromes, however, it is very difficult to ascribe the patho-
logic findings (both chemical and morphologic) to the malfunction of all or some
of the zinc enzymes described above. This may relate to the difficulty of
predicting the chemical and morphologic changes responsible for disrupting
a single or limited number of enzymatic steps in a complex and interrelated
series of metabolic pathways. For example, a disorder similar to cirrhosis of
the liver appears in zinc-deficient pigs and a relationship to alcohol dchydro-
genase might be postulated. It has been found that serum zinc content of
alcoholic humans with cirrhosis has been low and urinary zinc excretion has
been high.
331
-------
However, some features of zinc deficiency syndromes reflect unknown
biologic functions of zinc. One example is the well-characterized zinc
1710
deficiency that occurs in Euglena gracilis. In zinc-deficient
media, these organisms fail to grow well, and undergo peculiar morphologic
changes. Chemical analyses have shown that the concentration of DNA per cell
doubles, whereas protein and RNA synthesis are radically depressed. In
addition, a derangement of the relative concentration of other metals and a
1710
striking increase in acid-insoluble polyphosphate occur. It has been
suggested that zinc participates either in the structure and function of nucleic
acids or in protein synthesis, through effects on ribosomal structure or on
specific enzymes involved in protein synthesis. Indeed, it has been shown
1712
that RNA isolated from diverse biologic sources does contain zinc and that
added zinc and other metal ions significantly affect RNA structure, as revealed
518
in melting curves.
Zinc may have important enzymatic functions of which we are not yet fully
aware. For example, isolated samples of DMA. and RNA polymerase contain
zinc (Table 9-2). If zinc were essential to proper transcription in Euglena,
the chemical findings reported would be consistent with the activity necessary
to that function. However, finding zinc in a given biologic specimen is not
in itself sufficient evidence for function, because zinc is a ubiquitous element
in biologic fluids, second only to iron in relative concentration among the
transition elements. More direct relationships between the contained metal
and enzymatic activity must be demonstrated. Zinc has been clearly proved to
participate in the structures and mechanisms of action of many enzymes. But
it is possible that zinc also functions in maintaining the ionic environment
necessary for certain biologic processes or in the maintenance of the structure
of certain nonenzymatic proteins. For example, the cadndum-containing protein
807
metanothionein has been isolated from kidney, and shown to contain sub-
stantial amounts of zinc. The biologic function of this protein is not known
at present.
332
-------
9-3
Comparative Biochemistry of Zinc Metal loenzymes
Enzyme
Source
Molecular Zinc Content,
Weight gram atom/moleg.
Reference
Carboxypeptidase A
Carboxypeptidase B
Ul
Carbonic anhydrase—
Bovine pancreas 34,500
Porcine pancreas 34,000
Spiny dogfish pancreas 35,000
Bovine pancreas 34,000
Porcine pancreas 34,300
Spiny dogfish pancreas 35,000-
37,000
Hunan red cell 30,000^
Rhesus monkey red cell 30,000
Bovine red cell 30,000
Equine red cell 30,000
Porcine red cell 30,000
Canine red cell 30,000
Guinea pig red cell 30,000
Elastnobranch red opll
Bull shark 40,000
Tiger shark 40,000
Cuttlefish gin
Sepia of f icingi *-**
l
1
contains zinc
1
1
contains zinc
1
1
1
1
1
1
1
1
1
contains zinc,
but inhibited by
chelating agents
1661
Vallee and Neurath
486
Folk and Schirmer
889
Lacko and Neurath
1792
Wintersberger et al.
~
Folk and Gladner
1270
Prahl and Neurath
33
Armstrong et al.
405
Duff and Coleroan
831
Keilin and Mann
517
Furth
1596
Tanis et al.
214
Byvoet and Qotti
243
Carter and Parsons
997
Maynard and Ooleman
Addink
-------
9-3 (Continued)
Enzyme
Source
Molecular zinc Content,
Weight gram atom/moleg.
Reference
Carbonic anhydrase-
Alcohol dehydrogenase
£> Alkaline phosphatase
Aldolase (type I)
Aldolase (type II)
Parsley leaves
Neisseria sicca
Hunan liver
Horse liver
Yeast
Human placenta
Human . leukocytes
Human bone
Calf intestine
Escherichia coli
Bacillus lichenif ormis
SUbtUis
Babbit muscle
Other mammalian muscle
Yeast
180,000
(29,000
submit)
28,000
87,000
84,000
150,000
125,000
100,000
80,000-
89,000
121,000
1 29,000 g
1
2-4
2-4
4
0.15%
0.15%
zinc enzyme by
histochemical
staining with
zinc chelate
0.20%
no data
100,000
160,000
160,000
80,000
2
no zinc
no zinc
2
1624
Tobin
180
Brundell et al.
1706
von Wartburg et al.
1659
Vallee and Hoch
1658
Vallee and Hoch
1658
Vallee and Hoch
1629
Trubowitz et al.
142
Bourne
985
Mathies
1254
Plocke et al.; 48
Applebury and Coleman
Hulett-Cqwling and
Campbell747
Yoshizumi and Coleman
724
Horecker et al.
1812
Richards and Rutter
1345
-------
TABLE 9-3 (Continued)
u>
Ul
Enzyme Source
•
Aldolase (type II) Asocmycetes niger
C. utilis
Superoxide disnutase
Human red cell
Bovine red cell
Bovine liver
Yeast
Neurospora crassa
Escherichia coli
Streptococcus mutans
Molecular
Weight
•
—
Both of
two sub-
units
32,000
32,000
32,500
31,200
31,000
40,000
40,250
Zinc Content,
gram atom/moleJL
contains zinc
contains zinc
2 plus
2 copper
2 plus
2 copper
2 plus
2 copper
2 plus
2 copper
2 plus
2 copper
2 manganese
no zinc or copper
2 manganese
no zinc or copper
Reference
789
Jagannathan et al.
F72
Kbwal et al.
974
Mann and Keilin
508
Eridovich
1756
Weser et al.
1756
Weser et al.
1070
Misra and Fridovich
828
Keele et al.
1675
Vance et al.
^Unless otherwise noted.
-Mammalian erythrocyte carbonic anhydrases all have molecular weights near
30,000. Actual experimental data vary from 28,000-31,000.
-------
CHAPTER 10
CLINICAL ASPECTS OF ZINC METABOLISM
ABNORMALITIES OF ZINC METABOLISM IN DISEASE
Liver Disease
Zinc loss is a common accompaniment of all acute and chronic liver
disease. Decreased concentration of zinc in serum and liver tissue and the
increased excretion of zinc in the urine have been reported in patients with
610,1568,1666,1667,1668 1273
alcoholic cirrhosis of the liver. Prasad et al.
found decreased zinc in red blood cells of cirrhotic subjects. Oral adminis-
1568
tration of zine sulfate was reported to improve some liver functions,
but these studies were uncontrolled and attempts to
confirm their results have not been successful. 1666,1667 in chronic alco-
holism, patients commonly exhibit serum zinc concentrations two standard
deviations below the normal mean, elevated urinary zinc excretion, and increased
1 568
clearance of renal zinc. Urinary zinc excretion returned to normal
in some patients without cirrhosis who abstained from alcohol for 1-2 wk.
1568
But in patients with postalcoholic cirrhosis the changes persist.
Administration of alcohol to normal subjects has been associated with a
slight increase in urinary zinc excretion, but it is difficult to predict
1564
whether such an increase may occur.
The zincuria of alcoholism may be somewhat similar to the renal defect
1568
for magnesium found in alcoholics. Prolonged and excessive intake
336
-------
of alcohol with insufficient dietary intake of protein and other nutrients may
produce primary renal dysfunction with excessive excretion of zinc and/or
magnesium. Use of intravenous zinc-65 suggested that the hyperzincuria in
cirrhotic patients was not caused by a specific renal defect;1567 a more
general abnormality of cellular zinc metabolism was postulated.
Rats with cirrhosis exhibited reduced serum and liver zinc levels.810
Laboratory models of hepatic cirrhosis induced by carbon tetrachloride
and other agents have also been used to study the role of zinc in liver ab-
normalities. Liver damaged by carbon tetrachloride has been used to study
other manifestations of liver pathology related to zinc. Voigt and Saldeen1701
showed that water-soluble zinc salts given parenterally inhibit liver damage
produced by manganese in golden hamsters and carbon tetrachloride in rats.
A histochemical study139^ demonstrated that mice with hepatic damage from
carbon tetrachloride had much less pronounced enzyme changes when treated with
zinc beforehand to protect them. In animals treated
simultaneously with carbon tetrachloride and zinc chloride, alkaline
phosphatase increased in newly formed tissue in the peripheral zone of the
damaged liver. However, no difference in activity between zinc metalloenzymes
or zinc-dependent enzymes was found.1396 The uptake of zinc-65 in rat liver
damaged by carbon tetrachloride increased; uptake was greater in animals with damaged
liveuthan in controls.1809 Zinc was taken up by the parenchymal cells but not by
the Kupffer cells.1396 However, even after acute carbon tetrachloride damage,
uptake of zinc-65 was still rapid,89a suggesting that any alteration in zinc-65
uptake in damaged rat liver requires time following the insult to be apparent.
337
-------
In all patients with infectious hepatitis, serum concentrations of
total zinc decreased early in the acute phase of the viral disease and then rose
683
to normal as the illness subsided; diffusible serum zinc concentrations
were elevated early in the acute phase of the disease and fell to normal as
683
the disease subsided. Serum changes were always accompanied by a signif-
icant zincuria during the early acute phase of the illness and by a return to
normal as the illness subsided.
Little about the zincuria or the levels of plasma zinc of hepatitis
patients has been agreed upon; some investigators discovered no change from
normal urinary zinc excretion but a lowering of plasma zinc (which eventually
819
returned to normal), while others noted variability in urinary excretion
361,608,809
and plasma concentrations of zinc. The disagreement may be related
either to the failure to specify the stage of the illness at which the patients
were studied or to differences in methodology. Studies carried out late in
the disease may have missed the elevated urinary zinc excretion and lowered
serum zinc levels observed in the acute phase.
Decreased concentrations of total plasma zinc and increased urinary zinc
610,
excretion have been observed in patients with several forms of liver disease.
683,819
Attempts have been made to correlate abnormal levels of plasma zinc
661,809
with the severity of the hepatic disease with moderate success. In
patients with acute viral hepatitis, the lowest serum total zinc concentrations
and the highest levels of diffusible serum zinc and urinary zinc excretion
were observed in patients hospitalized because of the severity of their
683
disease.
338
-------
Normally, there are two classes of zinc ligands in serum. Macromolec-
ular zinc ligands, including albumin and o.-macroglobulin, exemplify one
537,538,668 2 * '537f66g
class; micromolecular zinc ligands, including amino acids
and species such as porphyrins and peptides, exemplify the other,, The
equilibrium of these ligands in serum depends upon their respective concen-
trations of albumin and amino acids, particularly levels of histidine
and cysteine.537'538'668 In vitro,
the amount of ultrafilterable or diffusible zinc has been shown to increase as
the concentration of these amino acids increases. Exchangeable zinc is bound
mainly to albumin, and the affinity of zinc for albumin is altered sharply
in favor of the amino acids histidine and cysteine if their concentration
537,538,668
in serum is elevated or if albumin concentration is decreased <,
However, urinary zinc excretion is related primarily to the amount of
micromolecular-liganded zinc complexes because they readily pass the renal
glomerulus and are excreted in urine. Indeed, oral administration of histidine
(\T\ 679
to humans has produced significant hyperzincuria and hypozincemia. *
In general, significant decreases in plasma albumin or globulin do not
1481a
occur in acute viral hepatitis. No such decreases were observed in
683
the patients studied by Henkin and Smith. However, the concentration of
some amino acids in plasma have
339
-------
460a
been reported to increase by 20% and increased urinary excretion of
amino acids such as histidine, lysine, and glutamine
has been observed. a These amino acids also
may be released into the circulation during the destruction of liver cells
683
that accompanies severe liver disease. Thus, the increased concentra-
tion of plasma and urinary free amino acids observed in hepatitis might be
associated with shifts in the binding of zinc from macromolecular to micro-
molecular ligands and consequent increases in diffusible zinc. These
changes would account for the significant zincuria and liver losses of zinc
observed in this disease.
Another mechanism by which diffusible zinc might be increased during
hepatitis is by reducing the affinity of the macromolecular zinc ligands for
zinc. In those patients observed by Henkin and Smith, serum diffusible zinc
683
varied directly with bilirubin concentration. When calculated separately,
the correlation between plasma bilirubin and diffusible zinc in patients
with bilirubin concentrations greater than 6 mg/100 ml (correlation coefficient
0.949) was greater than that in patients with bilirubin concentrations less
than 6 mg/100 ml (correlation coeficient = 0.605). Whereas the correlations
in the entire group and in each of the two subgroups were all highly signif-
icant (probability < 0.001), the higher correlation coefficient suggests a
closer relationship between bilirubin and diffusible zinc at higher bilirubin
concentrations. Nevertheless, the normal concentration of albumin in plasma
(650 uM) is so much greater than that of zinc (15 yM) or bilirubin (1 y M),
that even
340
-------
in the event of severe pathologic increases in bilirubin,478a»762b'1652»1809a
the displacement of zinc by bilirubin from a common binding site cannot account
for the observed increase in the concentration of diffusible zinc.
One attempt to clarify this complex relationship was made by Lindeman
and Baxter, who suggested that alterations in serum binding of zinc-65 were
present in patients with cirrhosis.933* Schecter et al. * studied the
distribution of serum zinc between albumin and ou-macroglobulin in patients
with decompensated hepatic cirrhosis. Not only did those patients with
cirrhosis exhibit lower than normal zinc levels, but a greater
proportion of the zinc in their serum (about 50%) was associated with
cu-macroglobulin, an amount significantly higher than percentages found in
normal subjects (between 20-30%). Schecter et al.1420a also noted that
albumin-bound zinc in serum was lower than that of normals, as expected. ' a
Decreased levels of zinc have also been found in the serum, red and white
blood cells,5^ and hair*28^ of cirrhotic patients. Increased urinary
zinc excretion and a decreased total body zinc pool, was observed. All of
these symptoms indicate that patients with decompensated hepatic cirrhosis
exhibit a total body loss of zinc.
Other Gastrointestinal Disorders and Malabsorption
Various abnormalities of zinc in blood have been reported in patients
with gastrointestinal disorders. Lower than normal levels of serum zinc
have been reported in patients with kwashiorkor,630a>1412'1504 although hair
zinc levels are elevated above normal.867a Lower than normal levels of zinc
in blood have been reported in patients with regional enteritis;14043 »1522
several malabsorptive
341
-------
998a,1715a 140
states; acute dysentery; gastric ulcers complicated by pyloric
1563 1802
stenosis; and achylia gastrica. However, no abnormal hematic zinc
361
levels were found in patients with acute gastrointestinal hemorrhage or
972a
following partial distal gastrectomy. Indeed, experimental production
of gastric ulcers in dogs revealed decreased fecal zinc excretion and pre-
sumed zinc retention in tissues. Increases in whole blood zinc
in patients with chronic pancreatitis have been reported, the result of de-
1817a
creased plasma zinc and increased red blood cell zinc. Zinc deficiency
may affect exocrine activity of the pancreas. The metal has been shown to
1661
be an important component of pancreatic carboxypeptidase, and the activity
of this enzyme was reduced in zinc-deficient rats.
Hypogonadal Dwarfism Syndrome
Dietary zinc deficiency has been associated with clinical symptoms in humans.
Prasad et al. reported on a group of Iranian men whose diet consisted almost ex-
17 76
clusively of bread and beans. Their deficiency was related not only to the low
level of zinc and protein in their diet, but the presence of phytate in the bread.
Phytate tends to decrease the intestinal absorption of endogenous dietary zinc.
1276,1277,1278,1287,1412 _ f * *• • -u • t j A
Symptoms of deficiency in these patients were decreased
zinc in plasma, red blood cells and hair; decreased urinary zinc excretion; rapid
turnover rate of zinc-65 with decreased 24-h exchangeable pool; and decreased
excretion of zinc-65 in feces and urine.
Iron deficiency anemia, dwarfism, hypogonadism, hepatosplenomegaly, depression
of adrenocorticotropin- (ACTH) production, and increased sensitivity to insulin
1412
was also found in a group of Egyptian adult males on a low-protein diet similar
to that observed in the previously reported Iranian men. Although the Egyptian
boys studied were infested with liver schistosomes, they excreted less urinary
zinc than did normal subjects; this tendency contrasts with the zincuria seen in
342
-------
patients with cirrhosis of the liver on zinc-adequate diets. This observation
suggested that these subjects, taking a diet which was apparently low in zinc,
conserved zinc by reducing its excretion in urine.
Anemia, hepatosplenomegaly, growth retardation and lower than normal levels of
zinc in plasma were also observed in preadolescent Iranian children.4363 The basis
for this syndrome was also considered to be the large amounts of phytate and fiber
present in their diet. Dwarfism, hypogonadism and alterations in zinc metabolism
have also been reported to occur in the U.S. without association with the dietary
intake of large amounts of phytate or fiber. 214a
Zinc salts and a good animal protein diet were fed to some Egyptian boys.
All symptoms of the syndrome were reported to diminish or disappear, and the
boys gained weight. The level of zinc in body fluids and hair were reported to
return to normal levels. One patient only on an iron supplementation did not
show these changes. After their plasma zinc had become normal, five patients
returned to their native villages and resumed their original diets; their
plasma zinc levels gradually decreased, reinforcing the conclusion that the described
syndrome was caused by zinc deficiency. Ronaghy £t al^ reported that administration
of zinc supplements to malnourished Iranian school boys improved their skeletal
«. 1363
growth .
Several investigators have doubted that zinc deficiency alone produced
this syndrome. The reported symptoms of these patients coexisted not only with
zinc deficiency, but also with protein deficiency, and were manifested by de-
248
pressed levels of serum albumin and plasma protein. Thus, Coughey con-
sidered the role of zinc deficiency overemphasized in these patients. In the only
controlled clinical trial carried out in this patient group, a double*blind study
of the administration of zinc sulfate and placebo indicated that no differences
could be shown between the treatment groups. Plasma and erythrocyte zinc levels
QO e
are below normal in children suffering from protein calorie malnutrition; these
885
levels are lower in kwashiokor than they are in marasmus. Hair zinc in growth-
retarded Iranian248and Egyptian248'1558 hypogonadal patients was below normal,
343
248
-------
although hair zinc levels in patients with either kwashiorkor or marasmus have
been found to be similar & to or appreciably higher than in normal subjects. a
In these latter patients plasma or erythrocyte zinc is lower than in normals or
in patients with dietary zinc deficiency. These differences may be useful in
differentiating between patients with abnormalities in zinc metabolism caused by
dietary deficiency from those with protein calorie malnutrition.
Some Egyptian patients with hypogonadism and growth retardation have attained
sexual maturity and a normal stature without treatment or change in their low
plasma zinc level. Thus the importance of the plasma zinc level as a reflection
of zinc deficiency may be limited. Whether zinc is the sole factor limiting
growth and development in these Iranian and Egyptian adolescents has also been
249 250 1281
questioned. A debate about the limited " versus sole importance of zinc
in growth and development in these patients has continued over the past
few years. Although this problem has not yet been resolved,
it is clear that zinc deficiency itself can be a restrictive factor in growth and
612
development. That zinc deficiency can alter other biologic systems and offer
an environment in which other pathologic processes may occur is readily apparent.
609,1282,1331 _ . -,,.,. * *• •
In humans several pathologic processes may accompany zinc deficiency,
making the issue of zinc deficiency per j3£ difficult to define and the effects of
treatment difficult to evaluate.
Acrodermatitis Enteropathica
rare
Acrodermatitis enteropathica is a/ autosomal recessive disorder characterized
by pathologic changes in several organ systems. Skin changes include alopecia
totalis, paronychia, and bullous-pustular dermatitis of the extremities of oral,
anal, and genital areas. Ophthalmic manifestations include blepharitis,
conjunctivitis, photophobia and corneal opacities. Gastrointestinal manifes-
tations include severe, often chronic diarrhea, malabsorption, steatorrhea
344
-------
and lactose intolerance. Tremor has been observed, along with occasional
cerebellar ataxia, emotional lability, and irritability. Infections caused
by Candida, albicans are frequent, as are retarded growth and hypogonadism.
The disorder, described by Danbolt and Closs,354c and the clinical
changes observed .since, have been defined and reviewed 440b'504c'802a>1232a'
1355a,1572a,1755a
The disorder, although rare, may be observed more frequently
in infants of Italian, Armenian, or Iranian origin, and usually develops after
weaning from breast feeding.
Diiodohydroxyquin therapy has been used with some success in these
384a
patients since 1953, but neither the mechanism of the drug's action nor
the pathologic basis for this disease was amenable to study until recently.
In 1973, decreased levels of serum zinc were noted in an infant with
acrodermatitis enteropathica, and after oral treatment with zinc sulfate a
77a,1086
complete remission of symptoms occurred. After inadvertent omission
of zinc sulfate from the patient's treatment, the symptoms of the disease
recurred; but they remitted again following reinstitution of zinc sulfate.
The dramatic alleviation of this condition with oral zinc sulfate has been con-
1037b,1129a,1266a
firmed.
Reasons for the changes observed in these infants after therapy are not
clear. That changes developed after weaning suggests either that infants
620a
received too little dietary zinc following weaning or that absorption of
zinc was impaired. Because very small amounts of zinc added to the diet
appear to correct abnormalities, it is rather unlikely that a specific dietary
672b,1129a
lack of zinc per se produces symptoms of acrodermatitis enteropathica.
That treatment with diiodohydroxyquin is effective might be related to the
1085b
formation of an absorbable zinc chelate. Human milk therapy may be helpful
950b
for similar reasons.
345
-------
Although there may be a significant abnormality in the manner by which
950a,950b 515a,672b
orally administered zinc-65 is absorbed, diverse data
suggest that malabsorption cannot be the primary defect in acrodermatitis
enteropathica. Although the nature
of the defect is not clear, the lack of an appropriate factor in gastro-
intestinal transport corrected by administration of even small amounts of
oral zinc is an attractive hypothesis, because zinc binding and transport
672b
after absorption appear to be intact. Such a zinc-binding protein has
873
been isolated from rat jejunal mucosa, although its role in
active transport across the gut mucosa has not been firmly documented.
Blood Dyscrasias
Plasma zinc was found to be below normal in pernicious
1369,1802 1802 1802
anemia, chronic lymphocytic leukemia, multiple myeloma,
58 1802
Hodgkin's disease, and various other types of anemiaso In Hodgkin's
58
disease, whole blood zinc also, is diminished. After vitamin IL-
therapy, the zinc serum level returned to normal in patients with pernicious
1802
anemia.
Lower than normal levels of serum zinc have also been observed in some
patients with thalassemia1275 and sickle-cell anemia,158a>1287a It has been
suggested that zinc may counteract the deleterious effect of calcium on the red
blood cell membrane. In uncontrolled studies administration of exogenous zinc
has been claimed to improve the clinical status of patients with sickle cell
anemia, assist in the healing of their peripheral ulcers and improve their
clinical condition in general. Controlled clinical trials will be of importance
to verify these provocative early findings.
1802
Elevated plasma zinc concentration was observed in patients with eosinophilia,
346
-------
lymphocytic lymphoma, severe untreated megaloblastic anemia, and multiple myeloma.58
These latter results in patients with multiple myeloma are at variance with those
noted by other investigators,1586>1589and make it difficult to sort out the changes
that occur in blood zinc levels in this condition. In megaloblastic anemia,
treatment with folic acid returned zinc levels to normal after a few
58
days. No changes in the plasma zinc level were observed in six patients with
polycythemia vera or in nine patients with acute myelogenous leukemia.
1367
Erythrocyte and plasma zinc were found to be reduced in Hodgkin's disease.
1367
Valberg ejt al. found an increase in the number of erythrocytes containing zinc
in Hodgkin's disease, chronic myeloid and lymphatic leukemia, and in multiple
58
myeloma, an observation contrasting with Auerbach's results. The discrepancy
may have been caused by different diagnostic criteria and whether patients were
studied before or during therapy. The erythrocyte zinc of the leukemic subjects
was higher than in the controls, ' except in patients with acute granulocytic
leukemia. High erythrocyte zinc levels also were found in patients with un-
. 1592, 1802 . . . . . 1592
treated pernicious anemia, and pernicious anemia in relapse,
although the degree of elevation did not correlate with the severity of the
disease. Elevated erythocyte zinc values were also reported in patients with
1369
lymphoma and multiple myeloma, in contrast with lowered values reported by
1367
others. In megaloblastic anemia, high zinc levels in erythrocytes have been
found to decrease with therapy, but elevated zinc values have not always
1369
been found in megaloblastic or pernicious anemia.
In some cases, erythocyte zinc content correlated with the mean corpus-
cular volume of the erythrocytes, as it did in cases of polycythemia rubra vera
and myeloid metaplasia. However, this correlation was not observed in leukemic
patients.505 Talbot and Ross1592 attributed the greater amounts of zinc in erythro-
cytes to the increased carbonic anhydrase that they found in patients with these
disorders.
347
-------
Zinc in leukocytes of patients with lymphatic leukemia has been reported
to decrease as the number of leukocytes increased and the disease process
worsened. This inverse relationship is particularly common in patients with
377
a majority of premature forms of cells in their peripheral smear. In 56
t
patients with chronic lymphocytic and granulocytic leukemia, or with acute
lymphocytic, monocytic, and granulocytic leukemias or myeloid metaplasia,
values for zinc in leukocytes were less than normal in all groups. The
lowest values occurred in subjects with chronic and acute lymphocytic leukemia.
No correlation between zinc in leukocytes and total or differential leukocyte
505,1802
number could be established. In patients with chronic granulocytic
505
leukemia zinc in leukocytes rose in response to treatment. However,
510
Frischauf est al. did not find an essential difference in the zinc content
in leukocytes of leukemia patients.
High concentrations of zinc in leukocytes were found in patients with
predominantly lymphoblastic forms of leukemias, whereas in
lymphosarcoma, Hodgkin's disease, lymphoma, and nonlymphomatous undifferentiated
237
carcinoma, the level of zinc was close to the normal range.
The zinc content in granulocytes was studied in bone marrow and peripheral
blood by a histochemical technique and results were graded on a semiquantative
1589
scale. Decreased zinc content was demonstrated in acute and chronic
myelocytic leukemia, plasmacytoma, and Hodgkin's disease. During the remis-
sion of leukemia the zinc content increased, but it was still below normal.158
903,1587
Rising zinc values were observed in lymphocytic leukemia. In patients
1589
with myelosclerosis, the zinc content of bone marrow granulocytes was elevated.
In four
348
-------
1587
cases of chronic lymphocytic leukemia, increased 2inc levels were obsc-rved.
1588
Szmigielski suggested that the amount of zinc protein in granulo-
cytes decreased in leukemic patients. Free protoporphyrin
can be demonstrated in leukemic cells, but it cannot be found in normal
granulocytes; he therefore hypothesized that protein synthesis with a pro-
toporphyrin prosthetic group was inhibited in leukemic cells. He related
the decrease in zinc content to the increase of free protoporphyrin in
leukemic cells, but it did not appear that the protein which regulated the
maturity of granulocytes contained a zinc-protoporphyrin-prosthetic group.
Decreased zinc in granulocytes of patients with acute and chronic myeloid
leukemia was one of the most constant signs of the disturbances in leukemic
903
cells. As so often in zinc metabolism, the meaning of these differing
observations is not immediately apparent.
373a
Delves et al. reported that the ratio of plasma copper to zinc
was elevated in children with untreated leukemia. This ratio was useful
in evaluating patients' responses to therapy.
Other Cancers
Numerous studies of the role of or the change in serum zinc in malig-
nancies have been performed, but the meaning of these findings is obscure.
The behavior of zinc in malignancy may depend upon the site, nature and
state of the cancer as well as the nature of the treatment.
349
-------
361
Davies et^ al. suggested that decreased zinc levels in serum of
patients with carcinoma of the bronchus are one of the most constant char-
acteristics of the disease because they remain in normal ranges in all other
pulmonary illnesses. The constancy of these decreased levels has
361,1802
been contradicted. Lowered zinc plasma levels have been reported
361 1802
in patients with carcinoma of the colon. Wolff, who studied 45
patients with carcinoma of different etiologies and sites, also found de-
361
creased serum zinc levels. After studying 49 cases, Davies et_ £l. did
not find significantly lower serum zinc values in patients with .carcinomas
than in the control group, except for those with carcinoma of the bronchus
and colon. X-ray therapy also has been said to produce an immediate in-
1802
crease in serum zinc values.
More recently, patients with primary osteosarcoma had elevated concen-
474a 800a
trations of serum zinc, a finding previously demonstrated. However,
474a
Fisher et^ al. also noted that patients with osteosarcoma with metastases
had depressed serum zinc.
It has been suggested that the level of zinc in cancerous organs was
higher than in normal tissue, although the tumors themselves may exhibit
813,1241
decreased zinc content depending on the phase of the disease.
7
Serum zinc levels have been elevated in some forms of leukemia, and lower
than normal levels of zinc were found in leukocytes of patients with various
505
acute and chronic leukemias.
Urinary zinc was reported to be three times as high in patients with
various malignancies as in normal subjects, and their urinary excretion
of molybdenum was also decreased; a zinc:molybdenum ratio
1241
higher than 300:1 was considered a manifestation of cancer. An increase
of the zinc:molybdenum ratio in urine was observed during the progression
1650 *
of the malignancy.
350
-------
A small increase of zinc in 30% of patients with carcinoma
was
observed in erythrocytes,1650 whereas zinc in the leukocytes of patients
with different types of carcinoma showed a decrease even in
18/i,185
very early stages of the disease. This change has been suggested
as a useful early test of the disease. A decrease of plasma
zinc even as an early prognostic of cancer was suggested after analyzing
1588
the zinc content of blood granulocytes of 50 patients. The granulo-
cyte zinc content in cancer of the skin was reported to decrease in advanced
stages of the malignancy.
The content of zinc in carcinoma of the prostate is of special interest
because this organ normally contains the highest amount of zinc in any
soft tissue in the body. In prostate, the level of zinc in malignant
1586
cells was much lower than in normal pro static tissue. As in pros-
tatic carcinoma, zinc content was increased rather than decreased in hyper-
plastic prostate glands.
Cystic Flbrosis (CF) of the Pancreas
Patients with CF exhibit higher levels of various electrolytes—
particularly sodium and potassium in eccrine sweat—than do normal sub-
jects, and this phenomenon has been useful in diagnosing the condition.
Spvever. zinc levels in hair and nails were reported to be lower than in
86ft
comparable controls, although actual levels were not given. Mean zinc
concentration in cerumen was 16 times higher in controls than in patients with
151
CF. . Pancreatic zinc concentrations in patients with CF were significantly
lower than those of controls. The mean zinc concentration in the pancreases
351
-------
from 17 controls was 193+94 ug zinc, whereas mean concentrations for 35 patients
with CF was only 77 + 73 yg zinc/g dry pancreas. Lower levels were associated with
the more seveVe form of the disease. Duodenal fluid from a patient with CF contained
868
almost no zinc. Differences among cerumen, hair and nails, which have lower than
normal zinc levels, and sweat and saliva, which have been reported to contain higher
levels, do not yield to simple interpretation.
Of further interest is the relative sterility of males and the reduced fertility
of women with CF. Decreased gonadal function is a common accompaniment of zinc
deficiency. Yet zinc levels in semen from men with CF are reportedly higher than
1385
in corresponding controls, because the ejaculate of CF patients contains mainly
fluid from the prostate with little or no fluid from the testis or epididymus.
Infectious Processes
Zinc in serum and urine of subjects in whom experimental viral and
bacterial Infections have been induced has been carefully studied. Under controlled
conditions,
Subjects were inf ected with viral and bacterial agents. Serum zinc levels
decreased rapidly after the onset of the viremia or bacteremia, but before
1219
fever was measured. The zinc decreased throughout the course of the
infectious process and was accompanied by a significant zincuria.
i
These changes have been related to an alteration of body zinc pools
by a leukocytic endogenous mediator (LEM), a heat-labile protein released
from sensitized polymorphonuclear leukocytes. LEM appears to produce a
94a,94b,94c,
prompt hypozincemia and a concomitant increase in hepatic zinc.
414a,720a,814a,1217a,1217b,1219,1219a,1285a
352
-------
Lower than normal levels of zinc in blood have been observed in patients with
typhus and acute dysentery. 140
Changes in serum and urinary zinc following infection may reflect a
natural body defense mechanism.1748 Because zinc is necessary to the growth
of viral or bacterial agents, decreased hematic zinc could represent a signifi-
cant factor in the virostatic and bacteriostatic processes of the body.1748
Decreased serum zinc and increased serum copper and iron are thought to be changes
that occur in response to invasion by viruses and bacteria; these alterations may
1748
be part of the normally active homeostatlc mechanism of the body. It is curious,
however, that zinc was able to display antiviral activity at the nontoxic concentra-
870
tion of 0.1 yM in tissue culture of viruses. The metal inhibited formation of
infectious virions at any stage of the replication cycle.870 Zinc also
inhibited the cleavage of precursor molecules. Out of 11 other metals, in-
cluding cadmium, copper, cobalt, mercury, molybdenum and nickel, only zinc
871
had these antiviral effects at nontoxic levels of the metal.
In vitro data suggest that at higher concentrations zinc markedly inhibits
the cleavage of polio and encephalomyocarditis virus polypeptides in relation
871
to rhinoviruses.
%
Drug Metabolism
Various drugs alter zinc metabolism in man and animals. In the rat,
N-methyl-N-nitrosourea results in retinal atrophy, cataracts,
877
and significantly increased zinc content in the eyes. However, administration
of 6-azauridine,
an antimetabolite effective in inhibiting de novo pyrimidine biosynthesis
and treating psoriasis, uniformly is linked to reduced serum zinc concentration
and increased urinary zinc excretion.1495 Administration of 6-azauridine has
been associated with the appearances of histidinuria, homocystinuria, and ex-
cessive levels
353
-------
of histidine, cysteine, and homocysteine in blood.
Other drugs may be associated with total body zinc loss and the
appearance of the most common symptoms of it. Drugs that interfere with
protein synthesis most commonly produce zinc loss, and they are used in
1092
the treatment of malignant processes. Common clinical symptoms of
zinc deficiency observed have been anorexia and weight loss, hypogeusia,
671
hyposmia, dysgeusia, and dysosmia.
However, zinc deficiency in the rat has been associated with a
reduction in the metabolic rates of pentobarbital,
aminopyrine, and £-nitrobenzolc acid. Microsomal
cytochrome P-450 was decreased- in livers of these zinc-
deficient rats. All biochemical and pharmacologic abnormalities reverted
to normal after 14 days of zinc repletion.
Renal Disease and Chronic Dialysis
In patients with proteinuria related to any renal lesion, associated
zincuria and a subsequent loss of total body zinc are reflected in decreased
serum zinc concentrations. In renal diseases in which proteinuria is not a
significant factor, total body zinc levels are variable. In one study of
patients with uremia, plasma zinc levels were lower than normal, urinary zinc
excretion was within normal limits, and levels of zinc were normal in most
308
tissues except for those in the kidney, which were lower.
354
-------
or) o
The zinc within various body pools of these patients has been redistributed.
613,975,1440a,1802
This relationship has been well documented. In rats, plasma
308
zinc levels decreased after ureteral'ligation, but kidney zinc increased.
Other organs showed little or no change.
Several changes have occurred in patients undergoing chronic renal
dialysis. In early trials with extracorporeal dialysis, loss of total body
copper and a subsequent neuropathy occurred. Some attempts to counteract
the loss produced acute copper toxicosis. 7>971,9/5,976,989a changes in zinc
in blood and tissues also occurred during these episodes, perhaps reflecting zinc
127,1075b
content of the dialysis fluids and the tubing as well. When these
materials were purified, many of the acute changes in copper and zinc metabolism
were obviated. The values obtained appear to depend, in part, on the type of
membrane coils used, but the state of the patient before dialysis and the length
1816d
of the dialysis may also have their effect. Patients undergoing chronic extra-
corporeal dialysis (with current technology) have a slight but consistent decrease
in serum zinc concentration from the zinc found in the dialysate. Although zinc is
reduced, a small but consistent increase in copper is found in the serum and in some
tissues of dialysis patients (unpublished observations, R. I. Henkin and C. W. Mueller)
It also has been suggested that the way the kidney handles zinc differs from
1547a
the way in which it handles sodium, calcium, and magnesium.
355
-------
Schizophrenia and Emotional Disorders
Serum zinc levels of schizophrenics have been reported to be lower
840,1239
than in controls; brain autopsy specimens from schizophrenics
were found to contain approximately half as much zinc as brains of patients
with diseases such as progressive paralysis, congenital syphilis,
840
epilepsy, and erysipelas. The urinary zinc level was elevated in a
378a
patient with porphyria and symptoms of schizophrenia. In this patient,
the authors related the changes in zinc levels to the psychiatric symptoms
observed, but it is well known that patients with acute porphyria exhibit
hyperzincemia. However, in a study of 20 untreated patients with acute and chronic
schizophrenia, Gillin (unpublished observations) could ascertain no
difference between blood, urine, cerebrospinal fluid, or gastric
fluid zinc levels of patients with untreated schizophrenia and controls.
These findings are set forth in Table 10-1.
TABLE 10-1
Comparison of Zinc Content in Various Tissues
of Patients with Untreated Schizophrenia and Normal Controls^
Serum, Urine, CSF,— Gastric Fluid,
Subjects No. yg/dl yg/24 h pg/dl ug/dl
Schizophrenics 20 92 ± 3^ 286 ±35 3 ± 1 42 ± 4
Controls 82 92 ± 2 353 ± 23 7 ± 2 40 ± 5
-Data from J. C. Gillin
-Cerebrospinal fluid
- + 1 SEM
356
-------
During insulin shock, serum zinc concentrations were reported to
840 J
increase; it was suggested that the enzymes responsible for glycolysis
were zinc-containing enzymes and that insulin treatment increased the
activity of carbonic anhydrase. Zinc loss has been correlated
with abnormal mental behavior. Zinc-deficient rats were reported to exhibit
950
impaired maze learning compared to pair-fed and ad libitum controls.
Zinc-deficient children were reported to perform below normal on cognitive
1246a
and perceptual motor tests. Disturbances of mood have been observed
1085a
in patients with acrodermatitis enteropathica and kwashiorkor, and the
acute loss of zinc following L-histidine administration commonly is associated
with psychotic ideations, including paranoid delusions, depression, and other
profound changes in mental state, symptoms which have been reversed after zinc
679
sulfate therapy.
357
-------
Other Disorders
Reduced plasma zinc levels were found in patients with psoriasis,
559 874 1794
various dermatoses, and venous leg ulceration. ' Withers et al.
examined patients with chronic venous ulceration and observed lowered plasma
zinc.
1802
Wolff measured serum zinc in many patients and found it to be
lower than normal in many acute and chronic conditions, including hypothy-
roidism (higher than normal levels were found in hyperthyroidism and hyper-
tension) . Lower than normal levels of serum and plasma zinc have also been
1252
reported in patients with rheumatoid arthritis.
Parenteral Nutrition
Fluids rich in amino acids are being given parenterally in increasingly
common fashion to assist the alimentation of patients with various disorders
or appetitive
when their own absorptive/mechanisms prove faulty. Parenteral hyperalimen-
tation usually is accomplished with a mixture of essential amino acids,
glucose, and electrolytes. Administration of these fluids
Is commonly followed by losses of
358
-------
total body zinc, ' ' perhaps because of the interaction of those
with
amino acids / the zinc bound to albumin. Albumin-bound zinc is shifted to
the amino acids and this amino acid-bound zinc freely passes the renal glo-
merulus and is excreted in the urine. The loss of total body zinc may become
profound and can contribute to the patient's already severe systemic abnormalities.
One way to obviate this loss is through the appropriate and judicial use
of zinc added to the parenteral fluids administered.
Cardiovascular Disease
Various abnormalities of zinc metabolism have been observed in patients
613,632a,999a
with myocardial infractions. Changes occur in hematic zinc levels and
1711 999a
metalloenzymes, and abnormalities of acute myocardial injury in man
632a
and experimental animals. It is unclear whether the changes observed
in blood zinc levels reflect specific changes related to myocardial function
or general changes from stress associated with it or other similar condi-
462,479a,1466a
tions.
A relationship between decreased incidence of cardiovascular mortality
in areas supplied by hard water, has been proposed, although mean serum
cholesterol and triglyceride levels were higher in subjects drinking hard
llla,333b,873a,982a,1446a
water than in those drinking soft water. what
characteristics of hard water that appear to convey this protection is not
Ilia
clear, but the magnesium and calcium content of the water may be important,
982a
as might be the trace metal content.
359
-------
Klevay has suggested that an imbalance between zinc and copper is an
858
important factor in the production of hypercholesterolemia in rats and
perhaps in humans. He has suggested that animal fat ingestion and the zinc:
copper ratio in milk may be important in the etiology of cardiovascular
Q C QV
disorders in man. These hypotheses have yet to be fully tested and are not
generally accepted.
Vitamin Metabolism
In 1939, Patek and Haig observed that some patients with hepatic
cirrhosis exhibited a night blindness that did not improve with treatment
with vitamin A.1213b Later studies also suggested a clinical
612a,851a
correlation between some forms of liver disease and night blindness.
During the acute phase of viral hepatitis, patients not only had serum
683
zinc levels lower than normal, but serum vitamin A and retinol
682a
(vitamin A alcohol) binding protein levels were similarly low.
Although hepatic levels of vitamin A are normal in zinc-deficient
1512
rats, hematic vitamin A levels appear to be low, as are serum levels
1514
of retinol and retinol-blnding protein. Concentrations of serum
retinol were shown to be lower in zinc-deficient than in zinc-supplemented
, . 1413
lambs.
These studies suggest a relationship between zinc metabolism and
vitamin A metabolism. Some of the
animal studies were carried out with diets in which less than adequate
amounts of protein were fed to the experimental group and pair-
fed controls were not systematically employed. Nevertheless, zinc might
be necessary to convert retinol to retinene (vitamin A aldehyde)
360
-------
an energy-requiring process involving the zinc-containing enzyme
retinene reductase. This chemical conversion in the retina requires retinene
reductase, an enzyme similar to alkaline phosphatase in molecular weight,
amino composition, and dependence upon zinc.
420
Relationships between zinc and the metabolism of several B vitamins,
728a 528
and between zinc and thiamine, and with pyridoxine,
282 1781
biotin, and folic acid have been noted. The action of
zinc and vitamins on food intake in animals was noted in 1934.
Feeding either biotin or folic acid had little effect on the growth,
food intake, or depleted levels of zinc in liver, hair or serum of growing,
1201
zinc-deficient rats.
Congenital Malformations
There are no direct data which demonstrate a sure teratogenic effect
in humans caused by zinc deficiency. However, Sever and Emanuel have deduced
from preliminary epidemiologic data that a possible teratogenic effect of
1475
zinc deficiency in man may exist. Some of these speculations were based
upon the high rates of malformations in the central nervous systems of
infants born in Egypt and Iran, countries where zinc deficiency has been
, t , 353a 756,757,759,761a 1584,1731,1732
commonly observed. Hurley and others
also have speculated that maternal zinc deficiency may cause congenital
754a
malformation in humans.
However, the rat, the model with which Hurley has worked extensively,
is highly susceptible to the effects of various teratogenic agents such as
antibiotics and cancer chemotheropeutic agents and therefore may not be
a relevant model for man. Yet a dam may be fed a zinc-deficient ration
361
-------
with, little pr no change in total body zinc content, although the rat fetus may
be severely damaged.756'757 Indeed, the fetus may be damaged even if zinc in-
take decreased only for a short time during the pregnancy.
Some investigators have speculated that American women may ingest marginal to
deficient amounts of zinc. Thus, it is conceivable that during some periods
of pregnancy, particularly in the first trimester, too little zinc reaches the
fetus. How this affects fetal growth or development is not fully known. Studies
of serum zinc levels generate little useful data because estrogens and other gonadal
and placental hormones may depress serum zinc levels regardless of the amount of
zinc intake. Hambidge reported a significant decline in hair zinc in
pregnant women between the seventeenth and thirty-seventh weeks of gestation,
but the meaning of these reductions cannot be readily evaluated because hair
zinc does not always reflect the level of total body zinc accurately with respect
to time of sampling.
Porphyria
Abnormalities of divalent transition metals are associated with various types
f , . 378a,1171a,1235c,1349a,1360
of porphyria. Patients with active porphyria
of several types have been reported to excrete excessive amounts of zinc in
their urine, C) although their serum levels of zinc may not be altered?253'1360
In 1929, elevated urinary zinc excretion was observed in a patient during an acute
exacerbation of porphyria. a Zinc-uroporphyrin chelates in urine have been
362
-------
1349a,1738a
measured, and found to be elevated in porphyria patients during
acute attacks. Abnormalities of zinc and/or copper metabolism have occurred
1235c,1360
in patients with the cutanea tarda hereditaria variety. These
studies led to the use of chelating agents like ethylenediaminetetraacetic
acid (EDTA) and British antilewisite (BAL)
as antidotes for these conditions. Although these therapeutic regimens were claimed
to be successful, data relating porphyria and its underlying abnormalities to
trace metal metabolism were not clear enough to establish how trace metals
1171a
may affect this disease. Indeed, conflicting data were presented, and
whether abnormalities of metal metabolism are involved in the etiology of
porphyria at all is still being disputed.
925a
Levine et al. studied the zinc metabolism of patients with several
types of porphyria. They found that after histidine was administered, the
half-time of orally administered zinc-65 shortened significantly, which
demonstrated a total body loss of the metal. They also found that during
histidine administration, urinary excretion of various porphyrins decreased,
668,679
although urinary zinc excretion increased. This previously observed
decreased in excreted porphyrin c was related by Levine et al. to the
2 26*i
specific action of histidine on the zinc requirement ' of
6-ALA synthetase, the critical enzyme controlling the rate of porphyrin synthesis.
Histidine administration dramatically lowered the urinary porphyrin excre-
tion to or toward normal in each patient studied. The role of histidine
was found to be useful in reducing urinary porphyrin excretion in these patients. a
2,265
Because zinc is a component of 6-aminolevulinic acid dehydiatase in several species,
it is possible that abnormalities of zinc metabolism may influence the synthesis of
porphyrins.
363
-------
THERAPEUTIC USES OF ZINC
Wound Healing
515,1410a,1584
Zinc plays an important role in protein synthesis. It
is essential for the normal activity of DNA polymerase and is present in
1462
RNA polymerase. Zinc also appears to influence the synthesis of
729
collagen. Removing zinc from DNA polymerase inhibits polymerase activity,
whereas complete activity is restored after its incubation with zinc ions.
Physiologically, zinc is important to growth and development. Some
108
bacteria require zinc to synthesize several essential amino acids. Zinc
1523
is also an essential nutrient for higher plants. Animals on zinc-deficient
diets develop anorexia, hypogeusia, growth retardation, and various skin
270,271,1004
dystrophies. Each sign of zinc deficiency found in animals has
also been reported in man, although the relationship between zinc deficiency
and the abnormalities observed have often been difficult to specify.
364
-------
These biochemical and physiologic observations indicate the impor-
tance of zinc to cell division in several systems of different species.
Therefore, it is not surprising that zinc would affect any system under-
going rapid cellular division, such as a healing wound. Such an obser-
vation was made in apparently healthy men undergoing surgical excision of
1262,1263,1265
pilonidal cysts. In this study, conducted without the
benefit of systematic controls, orally administered zinc was claimed to
accelerate the healing rate of surgical wounds.
Zinc may accelerate healing because zinc pools shift radically after
an operation. In the postoperative or wounded period, more zinc is necessary
to promote protein synthesis, collagen formation, or the incorporation of
zinc into enzyme systems. Radioactive zinc has been found to concentrate
in healing tissue, and highest levels have been found immediately after
1418
injury. The greatest activity of zinc in wound healing has been sug-
gested to occur during epithelization when the large store of zinc in the
1265
skin may be a convenient source of the metal. Zinc has been shown to
preferentially concentrate in healing tissues, with peak levels reached
1418
the third day after injury.
Serum and urine zinc have been reported to be lower than normal in
1
patients with bedsores and interpreted to reflect the depleted body zinc
295
of these patients. Oral zinc sulfate has been said to promote healing.
Zinc given to animals to aid wound healing is not consistently effective
1054,1058,1096,1411
in increasing the healing rate. These studies in animals
75a,291b,559a,1102b
and in man cast doubt on claims that zinc is influ-
ential in wound healing.
365
-------
75a 295 504c
Other controlled and uncontrolled studies ' ' have been
performed to evaluate the claims that zinc accelerates the healing rate of
surgical wounds. Single- and double-blind studies involving oral admin-
istration of zinc and placebo have been carried out in man and animals and have
291b,559a,762a,1102b,1305b,1471a
produced conflicting results. Some verified
v 559a,762a,
the efficacy of zinc in accelerating the rate of wound healing.
1265,1305b,l471a 75a,291b,lK)2b
Others did not. But if zinc plays such an
important role in cell division and protein synthesis, how can the negative
results be explained?
These studies are problematic. How can the rate or end point of the
healing process be established to the equal satisfaction of different investi-
gators? The absorption of oral zinc by humans is variable-dependent upon several
factors, including the counterion used and the nutritional status of the patient?
the results of different studies cannot always be compared directly. Evalua-
tion of the results may also be difficult because human zinc
absorption is slow and variable during the first few weeks of oral intake. As
long as 6-8 wk may be required before some aspects of bodily changes in zinc
metabolism can be measured.
Although no definitive answers have been found, a double-blind study
was reported in which the effects of zinc and placebo were studied on the rate of
605
healing of venous leg ulcers. If all data from that study were pooled and the
total results analyzed, no significant difference would be found between zinc and
placebo in accelerating the rate of wound healing. However, when patients with
lower concentrations of serum zinc were evaluated as a group, a statistically
significant acceleration of the rate of
366
-------
healing was demonstrated, whereas no differences could be
shown in patients with what investigators considered normal concentrations of
serum zinc. Similar outcomes (that is, open to two interpretations) were
also demonstrated in zinc-deficient rats and rats on diets supplemented
1411
with zinc. These results suggest that zinc aids wound
healing in zinc-deficient states, but not in states where zinc is adequate.
If these latter studies are confirmed, the only problem would be to identify
367
-------
the patient or subject group at risk.
Unfortunately, this latter task is simpler to state than to solve. In
rats on a zinc-deficient diet, the serum zinc concentration may be an adequate
1522
index of --' zinc deficiency. However, in humans, serum zinc concentration
may not - . accurately reflect the status of total body zinc metabolism.
Acute or chronic infectious processes, anemias, various drugs, liver diseases,
malignancy and other pathologic processes may depress the concentration of serum zinc
and affect a redistribution of body zinc pools, yet not necessarily produce
a total body zinc loss or a state of zinc deficiency. How then is the patient
at risk to be readily Identified?
No simple answer exists although several approaches are possible. The
clinical ideal would be a simple, direct method of assessing total body zinc
status with one quick and easy test. Since assessing status of the serum
zinc pool may be misleading, measurement of urinary zinc excretion along with
it may be helpful. However, this index also must be used cautiously, for
during decreased food intake or starvation human serum zinc concentration
may be normal, whereas urinary zinc excretion may be quite elevated. Any
bodily process which involves the breakdown or rapid turnover of cells is
1532 1537
associated with increased urinary zinc excretion, ' changes which may
or may not be reflected in serum zinc concentration. Since mobilization of
the hypothalamic-pituitary-adrenal axis is associated with decreased serum zinc
concentration and increased urinary zinc excretion, stress from any significant
source, including surgery itself, could be manifested by a redistribution of
body zinc and a transient zinc loss, both of which may be evident for a few
days. Measurement of hair zinc is relatively simple but it does not reflect
368
-------
the dynamic changes inherent in body zinc metabolism. Measurement of salivary
678
zinc may offer a direct index of total body zinc, but too little is known
about this new technique.
Administering zinc to zinc-deficient humans or animals may well
correct the numerous defects produced by the deficiency. The mechanisms by
which these abnormalities are corrected are multiple, and they are related
to the specifics of the several organ systems affected by the deficiency state.
However,
/the incidence of zinc deficiency in man has not been established clearly, and
it
as in most disease states, /is represented by a spectrum of severity. Secondary
effects of zinc, unrelated to net total body zinc loss, also may be important,
particularly in organ systems which metabolize rapidly. If the role of zinc
in cell division and wound healing in man is to be comprehended, changes in
the intake, absorption, excretion and distribution of zinc, as well as the
many biochemical, physiologic and pathologic factors which influence the
dynamic nature of these processes will have to be evaluated carefully.
States of Zinc Deficiency
Clinical zinc deficiencies in humans have been related to zinc
malabsorption caused by gastrointestinal diseases of several etiologies,
150a,214a,998a, 1412,1522 decreased zinc intake> and loss of
, ^ f , 296,899a,899b,899c,1793a TT ,
rich fluids. Urinary losses following hepatic and
309a,720a,
renal damage, the effects of certain drugs, excessive perspiration,
1288
' or some combinations of these phenomena also may be responsible
for zinc deficiency. Whatever the etiology, clinical symptoms of anorexia
369
-------
taste and wnell abnornialities have been considered early signs of Insufficient
673,679
zinc.
Taste and Appetite Disorders
fi 70
The role of zinc in appetite has been studied in normal subjects, and
679
those with scleroderma or malignancies. After zinc was depleted by L-histi-
dine administration, anorexia uniformly developed as the first symptom associated
with zinc loss. Appetite returned to normal after oral zinc administration. *
In some patients with malignancy and cachexia, treatment with zinc ion has been
useful in obviating some of the more severe effects of anorexia; in others with
idiopathic hypogeusia and anorexia, oral administration of zinc ion is useful in
..... --it-t. -t.i_ j 681. 1420b _ ,
obviating anorexia if the hypogeusia has been corrected. In the rat t
anorexia and a marked cyclic pattern of food intake also developed as the first
symptoms of zinc deficiency and these symptoms were the first corrected even with
270
the administration of very small amounts of zinc.
615,617,620a,621,679,1522
Acute or chronic zinc deficiency or loss in humans
1004
and animals has been accompanied by anorexia, hypogeusia and hyposmia. As
noted above, anorexia is the initial symptom in all species. The symptoms of
678
hypogeusia, hyposmia, dysgeusia and dysosmia have been shown to develop later.
Oral administration of zinc to human patients with acutely induced zinc deficiency
abolished the dysgeusia and dysosmia and restored taste and smell acuity to or
toward normal in each patient so affected. Changes in human taste acuity were
related to the diminution of the recently isolated par otic zinc protein, gustin,
which has a molecular weight of 37,000 daltons and contains 8% histidine and 2
moles of zinc I mole of protein. This protein is present in subjects with
370
-------
normal taste acuity but has been shown to be decreased in patients with various
types of taste dysfunction.674 Because this protein contains zinc and is the
major zinc-containing protein in saliva,674 measuring salivary zinc excretion
from the parotid gland may be a convenient way to obtain information about the
C6ntration of this protein in parotid saliva. '^?&'° Decreased zinc
concentration in parotid saliva often reflects decreases in salivary gustin
674
concentration; this change in turn is commonly associated with anatomic ab-
normalities in the taste buds.
The pore area of the bud is disrupted by several pathologic processes, but
they may be observed systematically only with the use of transmission electron
microscopy. Pathologic conditions may be reflected by several anatomic and
functional abnormalities. In patients who lack saliva, e.g., patients with xero-
stomia from Sjogren's syndrome or fatty parotid syndrome, there is a decrease
in taste acuity (hypogeusia), and in those taste buds that can be found, the pore
area of the bud is disrupted. Treatment of these patients with drugs or x-irra-
diatioa» in whom salivary flow was restored to or toward normal, was accompanied
by the restoration of normal taste bud anatomy and taste acuity to or toward nor-
684 . .*•
mal. Patients with untreated Sjogren's syndrome and hypogeusia do not
exhibit decreased levels of salivary zinc; rather they have normal levels of
parotid salivary zinc but apparently too little salivary secretion to support
normal taste bud function.
The salivary flow rate of patients with idiopathic hypogeusia (i.e., taste
loss related to unknown or unexplained factors) is normal, but taste acuity is
impaired. ' * Their taste bud anatomy is altered in a manner grossly
similar to that observed in patients with xerostomia — the pore area of the
taste bud is markedly disrupted and the normal cell types exhibit cytologic
pathology. In some patients with idiopathic hypogeusia, analysis of parotid saliva
revealed lower than normal levels of zinc and gustin. 674,678
371
-------
Treatment with zinc ion in a single-blind study restored taste acuity and
taste bud anatomy to or toward normal in some patients with idiopathic hypogeusia,
and in both single-and double-blind studies, concomitant increases in parotid sali-
673a,674,678,681
vary zinc and parotid gustin concentrations were observed. For
other patients not only was zinc ion ineffective in restoring taste acuity and taste
bud anatomy to or toward normal, but also their parotid salivary gustin concentrations
673a
did not increase. Placebo treatment was also effective for some patients, as
noted later, and ineffective for others in restoring taste acuity to or toward normal
673a
as observed in a double-blind study. No systematic studies have yet been pub-
lished in which the nature of metabolic or other changes in these patients improved
by placebo have been ascertained.
The rationale for treating patients with idiopathic hypogeusia with zinc
ion lay in empirical observations of taste and smell dysfunction and abnormalities
682a,683
of zinc metabolism among patients with acute viral hepatitis, and
in women during the first trimester of their pregnancies. In both of
these groups, taste and smell dysfunction returned to or toward normal
when zinc metabolism returned to or toward normal with the waning of the
hepatitis or the progression and subsequent termination of the pregnancy.
In a single-blind study of the influence of zinc ion and placebo in treating
patients with taste dysfunction of several etiologies, a statistically
681
significant effect of zinc ion was measured. In addition, hypozincemia
was observed among these patients, reinforcing the.hypothesis of an
abnormal zinc metabolism. In a subsequent double-blind study in a
similar group of patients, both placebo and zinc ion were equivalent in '
restoring taste acuity to or toward normal. Reevaluation of the double-blind
study, carried out in the same manner as the single-blind study, suggested that
the order in which the drugs were given affected the results of the study.
Thus, zinc therapy is clearly not effective in the treatment of unselected patients
with taste disorders.
372
-------
Although taste dysfunction has been described as a common accompani-
ment of zinc deficiency states and treatment with zinc ion has been shown
to correct the abnormalities of the deficiency, including those of taste
and smell dysfunction, patients with changes in taste and smell function
do not always exhibit abnormalities of zinc metabolism. As noted, available
data indicate that placebo therapy cannot be differentiated from therapy
with zinc in the correction of taste and smell dysfunction of diverse
etiologies.
The confusion between zinc deficiency in which taste loss is a
symptom, and taste loss, which may be related to a variety of factors,
is compounded by the lower than normal mean serum levels of zinc observed
in patients with taste and smell dysfunction when measured as a •-••• .
673a,681,1420b .
group. Before the double-blind study carried out by
£ 7O
Henkin and his associates, it was assumed that most patients
with taste and smell dysfunction exhibited an abnormality of body zinc
metabolism because of their low serum values. This hypothesis was
buttressed by the finding of decreased alkaline phosphatase in leukocytes
of patients with hypogeusia participating in the double-blind study.
However, these findings may not reflect total body zinc loss. In
a study of the oral absorption of zinc-65 in 13 patients with hypogeusia
and hyposmia, three groups of patients were tentatively identified. '
Six patients absorbed 65± 4%(mean - I SEM) of the zinc-65, a value
950a 1345c
similar to that reported ' for normal subjects. Five patients
absorbed 22 t 5% of the dose given, a value similar to that reported as
indicative of zinc malabsorption. This value is similar to that absorbed
by patients with untreated acrodermatitis enteropathica. Two patients
373
-------
absorbed 99 - 1% of the administered dose. Athough the mean level of
serum zinc in all 13 patients was lower than normal, only 30%
,i m n
exhibited apparent zinc malabsorption.
The role of zinc in the taste system has been investigated
in several studies. Taste bud-enriched membranes have been isolated
from bovine circumvallate papillae. As these membranes were purified,
the activity of alkaline phosphatase, a zinc-dependent enzyme, was
significantly enhanced and found to be the major enzyme of the purified
membranes. In addition, the specific binding of several tastants to
these purified membranes was inhibited by EDTA but rose toward normal
QC QU
by adding zinc ion. The saliva and mucus secretions from the
small gut of zinc-deficient sheep and rats are also different from
their normal counterparts and these secretions return to normal
when zinc ion is provided.
Color Blindness
Michaelson noted that in patients with acroderraatitis enteropatuica
whose color blindness was induced from optic atrophy after diiodohydroxquin
treatment, zinc restored color vision to normal. The visual cones
may be sensitive to zinc deficiency and thus zinc might influence the
visual process.
Treatment of Laryngeal Granulomas
Granulomas of the larynx arise from several etiologies and often are
difficult to manage. Since repeated surgical removal may be compli-
cated by repeated regrowth of this tissue, narrowing of the laryngeal
airway, and other surgical problems, a chemical method of treatment
has been sought. Oral administration of zinc has been reported to be
374
-------
useful therapy. °5' 1305a» 13<>5b The specific relationship between
zinc therapy and the occasional complete disappearance of these lesions
is not clear at present, although the rationale for its use is based
on the hypothesis that zinc accelerates the healing of wounds.1*18
Skin and Epithelial Tissue
Locally applied and orally ingested zinc has been used to treat several
skin disorders. Topical application of calamine, an impure zinc oxide, was
described as early as 1550 BC. Calamine is still widely used as a pro-
prietary preparation to treat many skin disorders and zinc oxide itself has
been prescribed by dermatologists for many years. Zinc is chief constituent
of many common proprietary agents, including talcum powders and skin creams.
Zinc stearate, oleate, and sulfate solutions
are main ingredients in talcums and creams. Zinc has also been claimed
296
to rejuvenate hair .and skin. Dilute solutions of zinc sulfate and
acetate are components of eye washes prescribed to treat conjunctivitis;
and they are sometimes applied as ancillary aids in the management of urethritis.
Zinc peroxide pastes are commonly-used in the treatment of skin ulcers,
especially sores that are complications of varicose veins or vascular insuffi-
ciency of the lower extremities. 1
Cardiovascular Disorders
Abnormalities of zinc metabolism in several disorders, including
hypertension, arteriosclerosis, and Raynaud's phenomenon have been directly
685b,686
or indirectly related to the pathogeneses of those disorders. Treat-
ment with zinc ion has been used to reverse the abnormalities observed in
686a, 1261,1557a
atherosclerotic cardiovascular disease. The validity of these
impressions remain to be clearly established, because clinical trials have
claimed that zinc
not been controlled. It has been / improves local blood flow to ischemic
375
-------
686
areas in patients with impaired circulation to the leg; this hypothesis
has not been rigorously tested yet. However, zinc-65 appears to deposit
preferentially in rapidly proliferating tissue, particularly areas undergoing
685a,843a
rapid healing.
Routes of Zinc Administration*
lu general, when zinc is administered to animals or humans, it is given
1262,1263,1265
by the oral route. Effectiveness of oral absorption depends upon
the many factors noted in Chapters 7 and 8. At present, sulfate is the only
approved, easily available counterlon for pharmacologic zinc therapy in the United
1647
States. Nevertheless, this compound is poorly absorbed and one of the more toxic
of 1'ie available zinc compounds.
Oral administration of zinc sulfate may be associated with gastrointestinal
toxicosis in some subjects. The Food and Drug Administration has approved the use
of zinc only as an emetic for humans. Thus, administration of significant amounts
of oral zinc is limited by this side effect.
The pharmacology of oral zinc administration and subsequent blood and tissue
levels have not been studied systematically. Similarly, pulmonary inhalation
of zinc and subsequent changes in blood and tissue levels have not been carefully
investigated. Since zinc is used to treat diverse disorders, its pharmacology is
of immediate and practical interest to physicians and veterinarians.
* Because of the limited state of knowledge and the preference for oral
administration, intravenous and other kinds of parenteral administration
of zinc will not be discussed. However, these routes have become more
important in treating some clinical conditions, particularly in patients
undergoing long-term hyperalimentation.615 '
376
-------
CHAPTER 11
TOXICITY OF ZINC
HUMANS
Ingestion, Inhalation, and Absorption
Acute effects. Apart from effects of inhaling zinc fumes encountered
during some types of industrial exposure, it is apparent that zinc, especially
from oral ingest ion, is not very toxic to humans. There are a few sparse
reports of accidental exposure of humans to food or drink contaminated with high
levels of zinc. However, these reports are problematic because it is difficult
to determine whether the alleged zinc toxicosis was caused by the zinc ger s_e or
by some impurity associated with the metal, particularly cadmium. In addition,
the literature is old and lacks present day confirmation.
Acute onset of gastrointestinal distress and diarrhea was reported to follow
ingest ion of lemonade prepared in galvanized iron garbage cans for military per-
222
sonnel stationed in Panama. Zinc absorbed into the acidic beverage was sus-
pected of being responsible for this outbreak of diarrhea, and use of galvanized
iron utensils for preparing or serving food or drink was subsequently prohibited
by the Army Medical Research Board. Another incident was reported in which several
people became ill, apparently with food poisoning, from food eaten which had been
173
cooked and/or held in galvanized containers. Storing food, particularly acid
foods such as citrus punches, in galvanized containers may bring about consider-
able zinc contamination.
An even less well documented report was a letter which described a couple who
drank water containing 40 ppm zinc from a galvanized pipe. They became
irritable, and suffered difficulty in concentrating, drowsiness, cloudiness of
thinking, mental fatigue, and headaches. A diagnosis of zinc toxicosis was only
suggested by the nonspecific symptomatology and the couple was said to have
improved upon change in water supply.
377
-------
More recent reports suggest that ingested zinc is toxic to humans
only in very high dose levels that break down the homeostatic mechanisms
124
controlling zinc uptake and excretion. Eighteen patients were given a
daily dose of 660 mg zinc sulfate for 16-26 wk for
treatment of venous leg ulcerations. No evidence of hematologic, hepatic,
or renal toxicosis from zinc was found in any of these patients. It was
reported that a 16-yr-old boy ingested 12 g of elemental zinc in 2 days
to hasten healing of a minor laceration. In 3 days he developed a lethargy
that progressed for 5 more days. He complained of light-headedness, and
exhibited a slightly staggering gait and difficulty in writing. These
effects were accompanied by an elevated blood zinc concentration and increased
1094
serum amylase and lipase, suggesting a pancreatic effect.
A patient on home hemodialysis experienced nausea, vomiting and
fever, allegedly from zinc toxicosis that developed when the metal contaminated
the water stored in the galvanized tank of her dialysis unit. Plasma and
520
red blood cell zinc concentrations were elevated.
Chronic effects. Whether long terra or chronic ingest ion of excessive
zinc in humans is associated with adverse health effects is debatable.
A syndrome consisting of gastrointestinal symptoms which include anorexia,
nausea, vomiting, epigastric discomfort, and weight loss was described
in workmen in a galvanizing plant as characteristic of excessive exposure
, . 1006
or zinc. However, simultaneous exposure to other potentially toxic
substances also occurred, so that identification of zinc as the principal
toxic substance is uncertain.
378
-------
Another report suggests occupational exposure to zinc oxide fumes at
concentrations ranging from 3-15 mg/m3 for 2-35 yr may not affect health
84
adversely. Increases in blood zinc and zinc content of gastric secretions
were found in furnace operators who were chronically exposed to zinc
622
oxide fumes in a brass foundry in Egypt. The increased zinc
concentration in gastric secretion might account for the reported gastric
complaints, notably epigastric pain after food intake. Radiography
detected no pathologic changes in stomach or duodenum in the four workers
studied by this technique.
On the basis of a single case report, it has been suggested that
chronic inhalation of zinc stearate powder may produce chronic pneumo-
coniosis.1145
Some observations of children have suggested that excessive oral intake
of zinc might be more widespread than previously suspected. For example,
toy cars made with zinc are often placed in warm bath water along with
soap. Because of the high pH, this environment would be conducive for high
concentrations of zinc to leach out of the toy into the bath water. Children
do drink bath water, and this chain of events has been reported to precipi-
tate the lethargy, fatigue and acute hemolytic anemia of some children
284
observed in hospital. Their serum zinc concentrations were elevated,
as was the amount of zinc excreted in their urine.
Excessive intake of zinc in the form of zinc supplements given to
aid healing of wounds has produced lethargy in children, along with high
levels of blood zinc.10^* A severe porotic effect of oral zinc on bone
also may result, especially if dietary calcium is low.463 Other investigators
251
have noted that oral zinc supplementation is not without medical consequences.
379
-------
Metgl Fume Fever^
Metal fume fever is defined as an acute disability of short duration
that occurs when fume is inhaled from metal heated to a temperature
above its melting point. Clinical features as well as the pathogenesis
of the syndrome have been summarized.177,751,829,1555 ^e disorder has
been most commonly associated with inhalation of zinc oxide fume, but it
may be seen after fumes of other metals are inhaled, particularly magnesium,
iron, and copper. But metal fume fever is most severe among brass founders
and the higher the proportion of zinc, the more severe are the symptoms.
Clinical signs and symptoms occur within 4-8 h of exposure. They
are characterized by hyperpnea, shivering with fever, profuse sweating,
pain in the chest and legs, and general weakness. An attack is usually
sliort--lasting only 24-48 h--and accompanied by leukocytosis. One of
v
the more intesting aspects of this disorder is that a second exposure
to the metallic fumes within 48 h will produce little or no response,
but if the second exposure comes after a lapse of 48 h or more, an attack
is likely. This is best demonstrated in workmen who suffer fresh attacks
on a Monday following a weekend away from exposure. However, if the
individual continues to be exposed daily, he does not become ill again.
The word "tachyphylaxis," meaning "quick immunity," has been applied to this
phenomenon.
Although metal fume fever was said to be recognized as long ago as
1822 by the French physician, Potissier,751 complete understanding of the
role of zinc in the pathogenesis of the syndrome is still not available.
McCord refers to metal fume fever as the foremost scientific enigma
380
-------
of industrial toxicology. Several theories have been advanced to explain
the observed features of the disorder but few of these notions have received
experimental support.
One appealing hypothesis is that inhaled zinc fume has a direct
chemical toxicity to alveolar and capillary epithelium, which brings
about an acute inflammatory response and consequent exudation. Fume
fever, the systemic reaction, may occur because protein produced by breakdown
of polyraorphonuclear neutrophils pours into the circulation. This concept is a
slight variation of an earlier proposal*1^ that the inhaled zinc produces pulmonary
bacteriolysis. Proteins released from the lysed bacteria are pyrogenic.
A more recent study suggests that protein released from cell breakdown
serves as an endogenous pyrogen. Extracts prepared from tracheal mucosa
and lungs of animals with experimentally induced metal fume fever have
T631
produced similar symptoms when injected into other animals.
McCord suggested that the effects of zinc on respiratory tract
tissues result in the formation of an allergen and eventually to an antigen-
antibody, or hypersensitivity reaction. The thesis is intriguing but it
has not been proved. Also, one might expect eosinophilic polymorphonuclear
leukocytes to increase significantly as part of the peripheral leukocytesis
occurring in the disorder, but apparently no such process occurs.
Another experimental approach has been to reproduce the disorder in
animals by spraying blood serum into a zinc oxide cloud, and collecting
and injecting this material into animals. It also was claimed that the
syndrome could be induced by the subcutaneous or intravenous administration
OnT
of zinc salts, but this method has not been repeated or verified. In fact,
there is little or no evidence to suggest that metal fume fever is a direct
result of systemically absorbed zinc jger_se«
381
-------
Some of the most detailed clinical and experimental studies of the
pathogenesis of metal fume fever here conducted by Drinker and his associates
398 399 402
between 30 and 40 years ago. ' ' It was demonstrated that an initial
respiratory exposure to zinc fume at 52 mg zinc/m air was capable of
inducing metal fume fever in human subjects, although later exposure to higher
concentrations within 24 h had no further effect. They believed then
and it was more recently contended by some ' that freshly formed fumes
composed of very small particles from 0.05-0.5 ym are necessary to cause
the disorder. As fumes age, they tend to agglomerate and become less
reactive. Larger particles tend to settle out or become entrapped higher in
the respiratory tract. 2
399
Drinker and Drinker also produced clinical metal fume fever in cats,
rats and rabbits by exposing them to high concentrations (600 mg zinc/nr air)
of zinc oxide fumes. Within 1 or 2 h the animals exhibited an initial decrease
in body temperature followed by fever. They also demonstrated that the highest
concentrations of zinc in these animals after pulmonary exposure were
in pancreas, liver and gallbladder, a phenomenon similar to that found after
the feeding of a single dose (0.175-1.0 g) of zinc oxide. They also learned
that elevated atmospheric carbon dioxide, perhaps because it increased depth
of respiration, increased the severity of experimentally induced metal fume
fever.
It has been shown that the inhalation toxicity of catalytically
active zinc oxide dust is increased markedly by. ultraviolet radiation.9^1
876
Krause demonstrated that inhaling zinc inhibited red blood cell
carbonic anhydrase activity; the level of inhibition correlated with dose
of zinc. Similar inhibition of carbonic anhydrase followed inhalation of
382
-------
magnesium oxide, a metal that also produces metal fume fever. He
suggested that inhibition of carbonic anhydrase was the actual cause of metal fume
fever, but beyond demonstration of the association, the rationale for this
notion was not discussed.
Reasons for the transient tolerance or immunity of experimental animals
and exposed workers to metal fume fever also remains unexplained. Drinker
observed that the period of immunity or tolerance seems to parallel the period
of the leukocytosis.
It also must be considered whether or not the symptoms of metal fume
fever that seem to be caused by the metallurgy of zinc are actually arising
from the presence of a metal contaminant, such as arsenic, cadmium, manganese,
lead, chromium, or silver. Apart from the knowledge that the syndrome may
be contracted from exposure to other forms of metal fume, particularly
manganese oxide, there is no other information implicating these
697
metals. Moreover, a similar syndrome (cotton-mill fever, or byssinosis)
occurs among cotton mill workers, presumably from exposure to a foreign
QC/:
protein in cotton dust.
Therefore, it can be said that exposure to finely divided zinc dust
will produce a disorder described as metal fume fever. The pathologic
mechanisms Involved In the expression of the syndrome are not yet defined.
The syndrome may be produced by other forms of metal fume or protein-containing
dusts.
ANIMALS
Animals are quite tolerant of high levels of zinc in the diet. Levels
100 times that required in the diet usually do not cause any detectable
383
-------
symptoms of toxicosis. Part of any observed toxicosis is caused by the
decreased food consumption that accompanies the feeding of high zinc diets;
interference with copper and iron absorption and metabolism is probably
also responsible for toxicosis.16^0 High zinc ingestion frequently affects
the joints of animals fed such a diet; this effect is probably not from inter-
ference with food intake or copper and iron metabolism, and may be related
to the increased level of zinc in the bones of animals fed excess zinc. The
adverse effects of high levels of dietary zinc on bone have been reported
'els of zii
1051,1198
464
to be aggravated in the presence of low calcium intake. Levels of zinc
as high as 600 ppm in the diet have not been reflected in hair.'
Elevated plasma zinc coupled with low plasma copper has been suggested as
a reliable indicator of chronic zinc intoxication.
Horse
Reports of lameness and death In young horses raised in the vicinity
of lead-zinc smelters prompted a study of lead and zinc toxicosis in young
1786
horses. Growth rates of animals fed zinc decreased after intake exceeded
90 mg/kg body weight/day. Enlargement at the epiphyseal region of the long bones
was the first clinical sign noted, and it occurred in all animals between
20-30 wk. These swellings were followed by stiffness, reluctance to walk
and turn, lameness, and reluctance to stand. The growth of animals receiving
lead was similar to that of controls until signs of illness appeared. Animals
that received both lead and zinc had lower levels of lead in bones but higher
levels in liver and kidney than did those fed only lead. They did not,
384
-------
however, have the neurologic dysfunction associated with the high levels
of lead.
Another report of illness in foals in the vicinity of a smelter was
diagnosed as lead toxicosis, 31 but the symptoms are more like those
described for the horses given either high levels of zinc or of zinc and
lead. Since zinc in grasses was as high as 1,100 ppm and 3,500 ppm in
overwintered grass, it is likely that the condition resulted from the high
levels of zinc as well as lead. Since a foal affected the summer before
the study was undertaken was still stiff the next summer, it would appear
that horses do not recover once they have been affected, although here the
animal was still eating the contaminated forage. Older horses and cattle
in the area were not affected.
A German study in which similar involvement of bone joints was
found in foals pasturing near metalworks with large lead and zinc emissions
754
indicated that cattle were also affected at times, although not as severely.
If not too severe, damage to the joint was apparently reversible; but both
horses and cattle were likely to suffer other complications which decreased
their usefulness for breeding.
Swine
A zinc-lead interaction has also been demonstrated in growing pigs.
Zinc (0.4% of the diet) increased lead (0.1% of the diet) toxicosis when
728
fed over 15 wk. The toxicity was greater with low levels of
calcium-phosphorus in the diet. (Zinc interferes with development and
mineralization of bone by decreasing calcium and phosphorus content as
well as reducing the calciumiphosphorus ratio.1391) Enlargement and
385
-------
softening of epiphyseal ends of the humerus and femur were reported in young
pigs receiving milk piped through galvanized iron piping and attributed to
568
high levels of zinc in the milk. In swine overdosed with dietary zinc
(0.4%), zinc levels in the liver increased, whereas hepatic iron decreased
markedly and copper remained unchanged.328 Pigs fed levels of zinc alone
up to 500 or 1,000 ppm for several weeks showed no significant difference
in weight gain.61'1640
Ruminants
Calves fed 600 ppm zinc for 7-21 days showed no sign of toxicosis
although zinc levels in pancreas, liver, and kidney were strikingly
elevated.1051'1060'1544 Lactating dairy cows receiving 1,300 ppm zinc
for 6 wk did not have a greater increase of zinc in their milk than cows
on a 700 ppm zinc diet. Moreover, feed containing 1,300 ppm did not
produce any discernible changes in health or milk production. Levels
around 1,000 ppm appear to be approaching toxic levels for lambs and feeder
cattle, however. Levels of this order have been found in mixed pasture
herbage downwind of possible zinc sources. Grasses grown in an area
in which a smelter previously had operated had levels of 600 ppm. ^ The
greater toxicity to ruminants may be accountable to adverse effects on
rumen microorganisms, since cellulose digestion by rumen bacteria in vitro
825 981
was reduced by zinc concentrations of 10-20 yg/ml. ' The effect varied
825
to some extent with the protein source.
Cows that accidentally ingested a very large dose C20,000 ppm) of
zinc oxide contracted severe enteritis and prostration, and some fatalities
followed.20
386
-------
Poultry
Mallard ducks fed high levels of zinc suffered severe paralysis; some
ducks were unable to walk after 30 days.52* The lowest level fed them was
3,000 ppm, but even at this level food consumption decreased. At higher
levels consumption was reduced to the point that none of the groups received
as much zinc in a 30-day period as did those on the 3,000 ppm intake. The
high mortality in the groups receiving over 3,000 ppm zinc may have been
caused primarily by decreased feed intake.
Addition of 3,000 ppm zinc to a corn-soybean meal ration significantly
reduced growth in chickens, too. Only a slight reduction occurred in
growth with 2,000 ppm, >1351 and taking chicks off the high zinc diets
brought about weight gains during the next 6 wk equal to that of the
796
controls. When given in a sucrose-fishmeal diet, 2,000 ppm
zinc did impair growth, but this diet did not support normal growth even
96
with control levels of zinc. Mortality from Salmonella galUnarum was
702
not affected by 2,000 ppm zinc in the diet. Growth in turkey poults was
1700
reduced slightly by 4,000 ppm zinc but not by 2,000 ppm.
Cat and Dog
Cats fed daily doses of zinc oxide between 150-600 mg and dogs given
400
500-1,000 mg daily for periods from 3 wk to 1 yr showed no sign of damage.
Although in this experiment cats did not reject the food until the zinc
oxide concentration was greater than 600 mg, other investigators found
that cats either rejected or vomited a diet containing more than 300 mg
zinc oxide. After 12-16 wk on a diet containing 300 mg zinc oxi^p, cats
lost weight and exhibited marked fibrotic changes in the pancreas as well
, 1458
as a decrease in pancreas size.
38,7
-------
Dogs given 4 rag zinc g]ucc-nate/kg body weight intravenously developed
•it CO
lassitude, enteritis, and paresis of the hind legs. Their electrocardio-
grams recorded changes similar to those charted in cases of potassium
intoxication.
Rat and Rabbit
The rat can tolerate very high levels of dietary zinc; the median
223
lethal dose of ingested zinc for rats is 350 rag/kg body weight. A level
of 2,000 ppm in the food during gestation and lactation had little effect
on either dams or offspring. ' Feeding 5,000 ppm during gestation did
not affect maternal weight or number of viable offspring, although fetal
gOQ
weights and weights of 14-day-old pups were decreased. If females were
fed 5,000 ppm from weaning, however, their litter size was reduced, and
most of the offspring were born dead. Feeding 4,000 ppm for 3 wk before
mating also reduced litter size. In another study, however, females that
received 7,000 ppm zinc from weaning were able to maintain pregnancy, although
457
litter size was reduced and only 50% of the fetuses were viable. If 4,000
ppm were fed during lactation, the pups had increased zinc and decreased iron
281
and copper in the liver. To sorae extent the hepatic condition was a reflection
of the higher zinc level (approximately three times normal) and lower copper and
280
iron levels in the milk; but since the pups had access to the dam's diet,
the pups' ingestion of the high zinc diet probably was responsible for much of
the change in mineral levels.
Consumption of high zinc diets accounted for decreased copper con-
centration in some tissues, particularly liver. Interference with copper
metabolism was reflected in lessened activity of cytochrome oxidase and
1687
catalase. Enzymatic activities returned to normal in both liver and
407
heart if the high zinc diets were supplemented with copper. In young rats
388
-------
liver copper was reduced on a 2,000 ppm zinc diet more than it had been
1761
on a 1,000 ppm diet. Zinc concentration in liver, kidney, and spleen
was much higher with 2,000 ppm zinc in the diet than with 10-1,000 ppm.266
Intake .of a high zinc:copper ratio (40:1) has been reported to cause
858
hypercholesterolemia In rats. High zinc:copper ratios were therefore
suggested to be of concern in human diets because of the association of
hypercholesteroleraia with heart disease. ' The hypercholesterolemia
in rats, however, may have been caused by copper deficiency because the level
of ingested copper was quite low. Since zinc and copper are antagonistic
in several aspects of metabolism, a a high zinc:copper ratio may induce
copper deficiency when copper levels are low, but have no effect when they
are higher. For instance, zinc:copper ratios of 10:1 and 20:1 reduced
1099
weight gain in weanling rats at a copper level of 1 ppm but not at 2 ppm.
Even with a low level of copper in the diet, a zinc:copper ratio of approxi-
mately 20:1 did not Increase serum cholesterol in weanling male rats fed the
442
diet for 4 wk. Supplementing infant formula with a zinc:copper ratio of
1799
17:1 for 6 mo did not increase serum cholesterol. Zinc sulfate injected
1560
subcutaneously reduced serum cholesterol in adult rabbits. Zinc was
given in drinking water (35-50 mg/day) to rabbits on control diets and on
atherogenic (cholesterol-supplemented) diets. Rabbits fed the atherogenic
diet showed decreased serum zinc levels and.compared to controls, zinc
472
concentration in their aortas was reduced. High levels of zinc in the diet
decreased fat content in the liver of-young rats on a high fat-low protein
diet.1391
Cardiovascular lesions have been linked to copper deficiency in the rabbit.
In rats, excess zinc decreases cytochrome oxidase activity of the heart, but
389
-------
969
copper supplementation will restore cardiac oxidase activity to normal
or greater than normal.^07 Similarly, high dietary levels of zinc markedly
decrease liver catalase and cytochrome oxidase activities, but they return
1 687
to normal with addition of copper. °°* However, copper deficiency in Americans
appears to be extremely rare. Since no evidence exists of marginal
copper deficiency in the general population of this country, there is no
reason to expect high zinc: copper ratios in the diet to cause hypercholestero-
lemia.
In pregnant rats and their offspring, high levels of dietary zinc
281
also reduced iron concentrations in some tissues, ' ' but that effect
was less marked and more variable "»"'» than the effect on copper. In
young rats, 4,000-7,500 ppm zinc produced a condition resembling iron
deficiency anemia and reduced hepatic iron stores. -*^*» 1516 Supplements of
iron plus copper increased hemoglobin concentration to normal levels,
969
although growth remained depressed. A level of 1,200 ppm zinc in the
diet of young rats was reported to cause decreased hematocrit, although
weight gains were not affected. It would be wise to check animals on high
zinc intakes for signs of anemia or copper deficiency even if growth does
not appear to be affected.
Reports of the effects of zinc on the kidney are sparse. One study
suggested that rats given intraperitoneal injections of zinc chloride
on alternate days for 10 doses of 2.4 and 4.8 mg/kg body weight would develop
histologic changes in renal tubular lining cells. *5/* The nuclei enlarge,
and the formation of intranuclear inclusion bodies is reminiscent of the
changes that occur in lead or zinc intoxication. Repetition of this
experiment In another laboratory failed to produce those changes (R. A. Goyer,
personal communication).
390
-------
Walters and Roe found no indication that feeding high levels of zinc for
as long as 1 yr accounted for any increased incidence of tumors in mice.1726
SUBCELLULAj^ EFFECTS
Mitochondria
Zinc inhibits respiration of isolated liver mitochondria at concentra-
—5 752
tions of about 10~;J M. Smaller concentrations induce mitochondrial
o / c
swelling. Effects of zinc ions on respiratory chain enzymes are complex
and depend upon the particular concentration. The most sensitive reactions
appear to be nicotinaraide adenine dinucleotide (NAD) reduction with succinate
and the electron transfer between cytochromes b_ and c±. Higher concen-
i o
trations of Zn further inhibit respiration by impairing flavin and
cytochrome oxidase activity. ^"
Brierly has demonstrated that Zn stimulates energy-linked
I n
accumulation of Mg in heart mitochondria, an action ordinarily not
associated with any irreversible membrane change, implying that the uptake
may be a physiologic effect.
Lysosomes
In an attempt to determine how zinc may beneficially influence various
286
tissue injuries, Chvapil et al. found that zinc stabilizes lysosomal
membranes by a mechanism restricted to the surface of the membrane.
CARCINOGENESIS. TERATOGENESIS. AND MUTAGENESIS
Zinc and Cancer
Tumor induction by zinc in experimental animal^. That zinc may induce
tumors was first reported by Michalowsky, who produced testicular
391
-------
teratomas in adult roosters by injecting a 5% zinc chloride solution directly
beneath the testicular capsule. Tumors developed in only a small percentage
of cases. However, spontaneously occurring testicular teratomas occur only
rarely.982 Zinc was found to induce testicular tumors in fowl only if the
testes were Injected during the months (January-March) when the gonads were
active, but they could be induced at other times of the year if the testes
were stimulated by prolonged injection of gonadotropins.
Injecting testes of Japanese quail with zinc chloride caused
few teratomas in organs where gonadal growth was stimulated by manipulation
CQQ
of the photoperiod. Zinc will not induce tumors in immature testes.
64 2^2
Zinc-induced teratomas do not metastasize, ' and are believed to be
derived from anlagen originating in germ cells and resembling reticulum cells.
These rests may become trapped in the zinc-induced scars.150" Zinc-induced
teratomas from testes of fowl have been transplanted successfully
39
into subcutaneous tissues and intra-abdominal cavities of other fowl.
The transplanted tumors grow rapidly and may invade adjacent skeletal muscle.
Injection of other fibrosing substances or irritants have not produced testicular
teratomas,1038'10*0 so that zinc per se applied in this manner somehow must
provide the tumorigenic stimulus to these cells. Similar tumors may be
induced in mammals. Testicular tumors have been produced in rats by
intratesticular injection of zinc chloride.1350
Except for the ability to induce testicular teratomas, no experimental
evidence exists that zinc administered orally or parenterally is tumorigenic.15
Mice fed diets containing 5,000 ppm zinc as zinc oleate for
3 mo, reduced to 2,500 ppm for 3 mo and then 1,250 ppm for a total time of
1 yr were studied for tumor incidence at 45 wk of age.1726 Dosage was reduced
392
-------
because of onset of anemia, weight loss, and death. Hepatoma, malignant
lymphoma, and lung adenoma were found in control and experimental groups.
Only the number of hepatomas exceeded the number of similar tumors in the
control group (7 out of 23:3 out of 24), but these numbers were not believed
to be significantly different. Addition of zinc sulfate to drinking water at
5,000 ppm and 1,000 ppm for the same period of time did not increase the
incidence of tumors
mor growth by zinc. Several studies of experimental animals
and observations of humans suggest that the administration of zinc may inhibit
tumor growth. The initial suggestion that zinc might have such an effect
arose from studies by Bishchoff and Long,*^ who injected virgin Marsh-Buffalo
mice aged 2 mo with a predictably high occurrence of spontaneous mammary
adenocarcinoma. Seventy-three percent of the control mice in this study
developed tumors at age 15 mo, whereas tumors occurred in only 38% of the
zinc-injected mice. Similarly, supplementing drinking water with zinc sulfate
reduced the incidence of 9,10 dimethyl-l,2-benzanthracene -induced
1 268
tumors in the cheek pouches of golden hamsters. Zinc chloride paste has
been found useful in controlling local effects of inoperable breast cancer.152*
Tissue zinc levels in humans with malignant disease. There is no available
evidence at the present time to suggest that zinc deficiency per se has any
etiologic role in human cancer. However, a number of studies have been
conducted which indicate change in tissue content of zinc in persons with
malignant disease, but it is not possible now to interpret the significance
of these changes. Furthermore, results from many of the studies to date
appear contradictory. Reasons for the present uncertanties are manifold,
393
-------
but variations in analytic methods and cytologic criteria guiding selection
of material to be analyzed may be responsible for some of the difficulties.
An early report suggested that some neoplasms contain particularly high
concentrations of zinc,336 but it is clear now that this does not apply to all
cancers. In fact, the converse Is more generally true. Tumors usually
contain lower levels of trace metals such as zinc than do non-neoplastic
tissues in the same patients,1616'1807 but exceptions may exist for specific
tumors. For example, scirrhous carcinoma of the breast and bronchogenic
carcinoma may contain increased amounts of zinc. But measuring zinc
ope
content of pleural fluid cannot distinguish benign and malignant lung diseases. J
Prostatic tissue has a particularly strong affinity for zinc,*5 4»965»994«1652
and hyperplastic prostate gland significantly increases in zinc content, whereas
carcinomatous portions of prostate decrease in zinc.138'355'593»709,993,1368,1443
In 1959, it was shown that the zinc level of blood from patients with
cancer generally is subnormal; and it has been suggested that the lower serum
zinc levels reflect lower levels of red cell carbonic anhydrase. Davies
found that 75% of all patients with carcinoma of the bronchus had plasma
zinc levels below the normal range. In a larger study of more than 100 patients
with bronchial carcinoma of all histologic types, he concluded that most
patients with bronchogenic carcinoma have persistent low serum zinc and that
25% of patients with malignancies that do not metastasize have lower than
normal plasma zinc. Patients with cancer at other sites do not have low
plasma zinc unless they have experienced marked weight loss or have low serum
protein levels.360 Others warned that zinc metabolism is inhibited by
excessive calcium, cadmium, copper and probably other elements, so that
* See also Chapter 7,
394
-------
lower plasma zinc levels can only be interpreted when amounts of other trace
metals in plasma are determined simultaneously.1556
Lower than normal plasma zinc levels have also been observed In children
with untreated leukemia. Plasma copper levels are increased but no correla-
tion was found between copper:zinc ratios and total white cell or peripheral
blast cell counts. The zinc content of noncancerous portions of liver
containing malignant tumor is higher than normal;1171'1807 in one case of
acute lymphatic leukemia with hepatic infiltration, the liver showed a
258% increase in zinc, a 301% increase in iron, and a 233% increase in cobalt.1171
Elevation of hepatic zinc levels in persons dying of malignant disease not
involving the liver has been found in more recent studies.5^' Morgan
et al. reported increasefll liver and kidney zinc in patients with carcinoma
of the lung. Three explanations for the increase in hepatic zinc that
accompanies malignancy have been postulated:
• the rise in liver zinc could reflect widespread preraalignant change
not peculiar to liver;
• the rise in liver zinc could be related to the poor nutritional
state of patients with cancer; and
• the rise in liver zinc could be a feature of a chemical defense
566
reaction of normal liver to invasion by malignant cells.
Table 11-1 shows the results of the study by Griffith et al.566
that confirmed the increase of liver zinc in apparently normal tissue in
subjects dying from cancer. No comparable elevations occurred in kidney,
heart, spleen, or pancreas* thus the authors suggested that the first two
explanations were Improbable.
395
-------
TABLE 11-1
Tis_sue
wJ All
Malignant Dis^ease_
Standard
jCojnjLr o_l_ Series
Standard
Tissue8
Liver
Kidney
Heart
Spleen
Pancreas
Mean D<
837
502
301
169
263
sviation
204
154
98
64
104
Mean
538
505
364
196
291
Deviation
95
106
69
42
126
rrobabiiity
<0.05
>0.49
>0.25
>0.25
>0.30
- Data from Griffith et al.
566
- Zinc values in mg/100 g ashed tissue.
Other explanations of the relationship between zinc metabolism and
cancer involve the influence of zinc on RNA and DNA metabolism and
chelating ability of chemical carcinogens.
Zinc Teratogenesis a_nd_Mu_ta_g_ene_si£
No evidence exists that excessive zinc produces any teratogenic effect,
It is of interest, however, that simultaneous administration of zinc with
cadmium salts will dramatically reduce the teratogenic effect of cadmium
in golden hamsters.
No literature was found to suggest that zinc is mutagenic.
396
-------
CADMIUM
Introduction
The close relationship between zinc and cadmium makes it necessary to
discuss both metals whenever one of them is under survey. This imperative
was recognized early. In an extensive study published in 1926 on workers
exposed to zinc8^ the authors pointed out that they had chosen a smelter
treating zinc ore with low content of cadmium and lead to avoid significant
effects from these other metals. However, this aspect has often been
neglected since. Reports on effects of zinc inside or outside zinc smelters
have sometimes not taken into account that significant exposure to cadmium,
lead, or arsenic was also likely. Similarly, food poisoning reported to have
been caused by zinc actually may have been caused by cadmium leaking from
zinc-plated materials. Zinc compounds often contain relatively large amounts
of cadmium, and since reports on animal exposures to large amounts of zinc
seldom state the concentration of cadmium or lead, it cannot be ignored that
some effects may have been caused by other metals. Aspects of relationships
1451
between zinc and cadmium have been discussed by Schroeder ejt a]..
Metabolism and Effects of Cadmium
Because the metabolism of zinc has been discussed elsewhere, only the
metabolism and effects of cadmium will be mentioned. For further details,
two recent reviews ' should be consulted. The following section is based
on those two reviews.
The systemic absorption of ingested cadmium in healthy human beings is
about 6%. Data from experiments on rats indicate that calcium deficiency
may cause higher absorption of cadmium. The placenta constitutes an effective
397
-------
barrier to cadmium and thus the newborn is virtually free from it.
Absorbed cadmium will mainly accumulate in liver and kidneys; in normal
people, the kidneys will contain about one-third of the total body burden.
The main part of the cadmium is bound to a low molecular weight protein,
metallothionein, which contains both cadmium and zinc. The synthesis of
this protein is stimulated by exposure to cadmium. It has been claimed
that zinc also may stimulate the synthesis and that metallothionein is important
to zinc metabolism. *
The accumulation of renal cadmium may eventually cause renal tubular
dysfunction. The critical level, that is, the concentration at which the
first signs may appear in sensitive individuals, has been estimated to be
about 200 yg cadmium/g wet weight in renal cortex. Present mean concentra-
tions in European and North American populations at age 50 are 15-30 yg cadmium/g
wet weight, whereas in parts of Japan regarded as nonpolluted by cadmium,
higher values have been reported.
The first known sign of renal dysfunction in human beings is increased
excretion of low molecular weight proteins. This sign may be followed by
more advanced proteinuria, glycosuria, aminoaciduria, and disturbances in renal
handling of calcium and phosphorus. Itai-itai disease,* found in Japan, is
thought to be caused by disturbed mineral metabolism; women with calcium
deficiency and renal damage caused by cadmium after prolonged ingestion of
rice contaminated by the metal have exhibited the disorder.
Excretion of cadmium is very slow and thus the biologic half-life
is extremely long: estimates from between 10-30 yr have been postulated.
*Itai-itai or "ouch-ouch" disease got its name from the severe pain caused by
the multiple fractures of bones that occur. It is resistant to treatment with
vitamin D^, except in extremely large doses. The disorder was reported in
Toyama, Japan, where rice fields had been contaminated by river water used for
irrigation. The pollution source was a zinc mine many miles upstream.
398
-------
INTERACTIONS .BETWEEN ZINC AND CADMIUM
This section will mainly deal with in vivo interactions between zinc and
cadmium, as a vast literature on in vitro effects of cadmium on zinc, especially
in enzyme systems, is easily available. For accounts of such experiments, a
1663
review by Vallee and Ulmer may be consulted.
Animals
Major work in this field was initiated by Parizek,1206 who observed
that a large dose of a zinc salt could prevent the action of a large single
579
dose of injected cadmium on the testes. His finding has been confirmed. '
581-583,817,818,983 Also> the teratogenic and carcinogenic effect of large
injected doses of cadmium can be unequivocally prevented by zinc. '
However, these were all acute experiments, involving large amounts of injected
cadmium and zinc and with little application to long-term toxicity problems.
That dietary cadmium could accentuate zinc deficiency symptoms was first
shown in turkeys. "* The adverse effect of cadmium could be reduced by
1574
increasing zinc intake. Similar findings were then obtained in chicks.
This research had great heuristic value. Groups of chicks were fed diets
with varying amounts of zinc and cadmium (and effects of copper and iron were
studied as well). The basic diet contained 25 pg zinc/g and was supplemented
with 200 pg zinc/g and/or 100 pg cadmium/g. It was found that cadmium depressed
the growth rate and caused pathologic gizzard changes and that both these
changes were reversed by zinc. Cadmium brought about hypochromic anemia,
nf\ -I
which was not reversed by zinc. Bunn and Matrone studied groups of rats
and mice. Some groups were pretreated so that they were copper-deficient.
Zinc was added to the diet, which contained 9 pg zinc/g and 2 pg copper/g
in a concentration of 200 or 400 pg zinc/g diet and/or cadmium in a concentra-
tion of 100 pg/g diet for 5 wk.
399
-------
Whereas supplemented zinc did not increase zinc concentrations in liver
or testes, ingested cadmium caused zinc concentrations to increase significantly
in these organs in normal and copper-depleted mice on both basic and supplemental
zinc diets. Also, supplemental copper increased zinc concentrations, especially
in the testes. In rats the results were not so consistent. Cadmium exposure
actually decreased liver zinc in copper-depleted animals on the basal diet,
but hepatic zinc markedly increased in animals pretreated with a commercial
diet.
Even in zinc-supplemented and copper-depleted animals, there was only a
slight increase in liver zinc, whereas animals treated with copper plus zinc
increased their zinc levels markedly after cadmium exposure. Cadmium caused
anemia, which was partly reversed by copper, but not by zinc. Indeed, anemia
was more pronounced in mice given both zinc and cadmium.
Banis et _al. made similar studies on rats. Zinc (200 Vg/g), cadmium
to a basic diet.
(100 yg/g), and iron (68 yg/g), alone or in combination, were added/ Cadmium
depressed weight gain, and although zinc or iron alone did not reverse this
effect, zinc plus iron did. Cadmium decreased hemoglobin concentration and
cadmium plus zinc further reduced it. Another experiment demonstrated that
cadmium intake increased liver zinc in animals on basal diets.
Miller et al. found increased fecal zinc in calves given a diet with
40 yg zinc/g and 350 yg cadmium/g. Powell et al. found that in calves given
640 yg cadmium/g in a diet that also contained 27 yg/g of zinc, kidney and
liver levels of zinc increased. No tissues examined were found with zinc
levels below those of the controls.
400
-------
That there was an Increase in fecal zinc after ingestion of cadmium seems to
^ 7 ft
conflict with the findings of Gunn e£ al., who found that injecting cadmium
in rats caused reduced excretion of zinc-65 via feces. However, the high amount
of dietary cadmium given by Miller et al..1059 may well have reduced zinc absorp-
578
tion. The results of Gunn et al. will be discussed in detail later.
1259
Pond et. al. gave 154 yg cadraium/g diet to pigs on diets high (74 yg/g)
and low (22 yg/g) in zinc for 6 wk. Whereas low zinc alone did not depress
weight gain, ingestion of cadmium did cause a significant drop, although the
effect was partially overcome by the higher zinc intake. Fox et al.504a
found that in young Japanese quail given a diet with cadmium (75 yg/g) and
zinc (75 yg/g), cadmium markedly reduced the zinc content of the tibia.
All the above experiments were performed using high dietary levels of
12^6
cadmium and/or zinc. In contrast, Petering et al. gave groups of weanling
rats a zinc-deficient diet ( <2 yg zinc/g) and zinc in drinking water in
concentrations of 0, 2, 8, and 32 yg/g. Optimum growth was obtained with
8 yg/g, whereas at 2 yg/g, growth was marginal. After 4 wk, 50% of the groups
on 2 and 8 yg zinc/g water were then given cadmium in the water at a
concentration of 3.4 yg/g for about 7 wk. The molar ratios of zinc:cadmium
were 1:1 and 4:1. The cadmium exposure was considerably lower than those
used earlier, and it is more realistic. It was found that in the group on
2 yg/g water, cadmium decreased the growth rate, and was responsible for
some pathologic changes, such as corneal keratinization, which are seen in zinc-
deficient animals. Gonads did not seem to be affected. Rats on higher zinc
intake were not influenced by cadmium.
The most significant finding with regard to distribution of zinc was
that the zinc concentrations in testes markedly decreased in rats given
marginal zinc and cadmium (molar ratio, 1:1), whereas no such change was noted
401
-------
in rats given zinc: cadmium at a ratio of 4:1. Therefore, cadmium can cause
changes in distribution of zinc at marginal intakes. Zinc will still be stored
in organs such as liver and kidney, where cadmium will accumulate. However,
zinc may be depleted in other organs where cadmium does not accumulate as
greatly.
Most studies on long-term interactions between zinc and cadmium are ingestion
1691
experiments, and few studies use injection techniques. Vigliani reported
on rabbits given subcutaneous injections of 0.25 mg cadmium/kg body weight, 5
days a week for several months. Some animals also were given injections of
equiroolar amounts of zinc. Morphologic changes and degree of proteinuria
revealed that cadmium alone caused more severe renal tubular damage than cadmium
1691
plus zinc.
That cadmium can alter the metabolism of zinc has also been shown in
322
isotope experiments using zinc-65. Cotzias et al. gave 63 uM of
cadmium, zinc, copper, or mercury as the sulfate or citrate in a single
intravenous injection to rabbits, alone, simultaneously with, or after an injection
of zinc-65 as the chloride. Cadmium as well as stable zinc changed the
clearance curves of zinc-65 in blood similarly, whereas the other metals
had no effect. And, there were differences in the distribution of zinc-65
activity in organs after the administration of zinc or cadmium. The authors
interpreted the results to mean that some degree of interchange between cadmium
and zinc occurred in vivo, but zinc-zinc and cadmiura-zinc interchanges were not
identical.
When zinc or cadmium was given some time after the zinc-65 injection,
differences'appeared in the plasma zinc-65 curves, indicating a difference
in exchange. The doses administered were high, but nevertheless these data
indicate a possible interaction between cadmium and zinc.
402
-------
Gunn et. _al. found that cadmium decreased the uptake of zinc-65 in the
testes and dorsolateral prostate. They conducted the following four studies on
rats and mice:
• Rats were given an intracardial dose of zinc-65 4 mo, 1 wk, and 1 day
after a subcutaneous injection of 3.4 rag cadmium/kg body weight. After
24 h, the zinc uptake of organs was measured. Increased zinc uptake
was noted in liver, pancreas, and kidney, whereas no difference was
found.in skeletal muscle,
• After an intracardial injection of zinc-65 to rats, an intracardial
/
injection of 2 mg cadmium/kg body weight was given 24 h later. Liver
content of zinc-65 was measured 5 and 48 h after the cadmium injection.
Five hours after injection, concentration of zinc-65 decreased in
cadmium-treated animals compared to saline-treated controls whereas
after 48 h the reverse was found. This initial displacement of zinc
322
from the liver had been noted before.
i
• Zinc-65 was given to rats in an intracardial injection and feces
were collected daily. Four days after the zinc dose, half of the rats
were given a subcutaneous injection of 3.4 mg cadmium/kg body weight.
Feces were collected for another 3 days. Compared to controls, zinc
excretion decreased in the animals given cadmium and it was found that
zinc-65 concentrations were higher in the cadmium-treated animals.
0 Three days after mice were injected subcutancously with 3.4 mg cadmium/kg
body weight, they were given a subcutaneous dose of zinc-65. Animals
were killed 1, 2, 3j and 7 days after the zinc-65 injection. Activity
was measured in liver, kidney, pancreas, and carcass. Whole body retention
was 15-20% greater in cadmium-treatod rats than in controls.
403
-------
The authors interpreted the results to mean that cadmium was interfering
with the gastrointestinal excretion of zinc, thus causing zinc levels to
rise in liver, kidney and pancreas. The present knowledge of cadmium
metabolism suggests the opposite, that is, the decrease in excretion was
caused by increased zinc retention.
Studies also have been conducted on more specific physiologic problems.
Whereas large doses of injected cadmium bound to cysteine
increased renal tubular reabsorption of sodium, zinc had no effect at all.
Cadmium, either after injections or long-term, low-level ingestion, can
cause hypertension in experimental animals; it occurs at a molar cadmium:zinc
into
1445
1445
ratio of > 0.4. When a zinc chelate was injected into animals made hyper-
tensive by cadmium, blood pressure decreased rapidly.
There are a number of zinc-dependent enzymes which could be susceptible
in vivo to cadmium interference. Alkaline phosphatase activity decreased
in renal cortex of rabbits exposed to cadmium for 6 mo by subcutaneous
injections. This decrease still existed 6 mo after Che last injection.
It was postulated that leucine aminopeptidase would be a kidney enzyme
1.691
susceptible to cadmium. 7A This enzyme is important for protein catabolism
in the kidney and interference with that system could be influential in the
development of tubular proteinuria. Cousins e£ aU showed that in
cadmium-exposed pigs the activity of this enzyme in renal cortex was decreased
after 150 pg cadmium/g in the diet (80 pg zinc/g) for 6 wk, resulting in a
mean renal concentration of 78 pg cadmium/g wet weight, and about 100 pg/g wet
weight in renal cortex.
Because different levels of cadmium were given and both cadmium and zinc
were determined in organs^ the research also dealt with distribution of zinc
404
-------
in relation to cadmium. Zinc levels increased ir whole kidney, and on a molar
basis there was a distinct change between 150 and 450 pg cadmium/g diet.
Exposure to 50 and 150 pg cadraium/g resulted in molar cadmium:zinc ratios of I
0.55 and 0.71 respectively, whereas 450 ^g/g made a molar cadmium:zinc ratio of
2.73. The renal concentrations of cadmium were 41, 78, and 276 pg/g wet weight.
These results indicate that the critical level in this experiment must have
been between 100 and 300 pg/g wet weight in renal cortex. The decreased
leucine aminopeptidase activity indicates that the level could well be close
to 100 Pg/g wet weight.
Liver enzymes such as glutaraic-oxaloacetic transaminase (GOT), alkaline
phosphatase, aldolase, and succinoxidase, as well as oxidative phosphorylation,
1539
were studied by Sporn et al. Young rats were given diets supplemented with
10 pg cadmium/g and/or 80 pg zinc/g. Growth rate was not affected by cadmium.
Cadmium caused a decrease in phosphorylation, but zinc counteracted this effect.
Cadmium also reduced the activity of GOT, alkaline phosphatase, aldolase, and
the succinoxidase system, but zinc did not affect this change. The authors
emphasized that the zinc-cadmium interaction was not a case of general antagonism,
but an effect limited to certain enzymes and metabolic sequences. Original
cadmium and zinc levels were not reported. It was calculated by using data
from similar exposures that the amounts of liver cadmium could only have been
a few pg/g wet weight.
1449
Long-terra effects of cadmium were studied in two generations of mice.
Cadmium was given in drinking water at a concentration of 10 pg/g from the
end of the weaning period until the end of the experiment. The metal was
found to be toxic for breeding mice to such an extent that there was no
405
-------
survival beyond the second generation. Congenital abnormalities appeared
at a much higher frequency than in controls. Cadmium does not traverse the
placenta and it is conceivable that a teratogenic effect was caused by
a secondary zinc deficiency in the fetuses. However, cadmium and zinc were
not determined in the organs.
1249a
A recent paper on normal levels of cadmium and zinc in horse kidneys
reported that cadmium concentrations in renal cortex varied from 5-250 yg/g wet
weight in 37 horses aged 1-25 yr. At lower cadmium concentrations (5-70 yg/g)
an equiroolar increase in zinc occurred, but at higher cadmium concentrations
zinc did not increase to the same e.xtent as cadmium.
Humans
Information on the relationship between cadmium and zinc in humans has been
1451
obtained by autopsy studies. Schroeder et al. showed that parallel increases in
1250
zinc and cadmium levels in kidney occurred with age. Piscator and Lind
found the same relationship and demonstrated that the increase in zinc was
equiraolar to the increase in cadmium. This discovery was thought to be
connected to the presence of raetallothionein in the renal cortex, because
this protein normally has equiroolar amounts of the two metals. Furthermore,
it was shown that if the amount of zinc equiraolar to the cadmium were
subtracted from the total zinc, the physiologic amount of zinc in the cortex
could be determined. The concentration of this zinc fraction was found to
be about 160 yg/g dry weight (34 yg/g wet weight), piscator and Lind's
data were obtained from members of the general population. They were
not victims of occupational exposure, and had concentrations of cadmium below
levels that could cause renal damage.
406
-------
CHAPTER 12
STANDARDS FOR ZINC LEVELS
Air quality standards for zinc and its compounds have been established
in many countries for occupational exposures, but a thorough search of the
literature did not reveal any standards for public exposure.
Table 12-1 presents the current standards for industrial situations,
all of which are maximum allowable concentrations (MAC). Their variations
may arise partly from historical reasons. Before 1962, the U.S. standard for
Q
zinc oxide fume was 15 mg/ra and was based primarily on the work of
402a 451a
Drinker and Fairhall. Subsequent experience showed that zinc fume
3 28
fever occurred from exposures to less than 15 mg/m . No adverse effects
have been reported where this standard is observed.
EXISTING STANDARDS FOR WATER QUALITY281*' 1011b' 1645b» 1806a
Standards for drinking water quality for zinc are based primarily on
esthetic characteristics such as taste and cloudiness. Most standards
recommend 5 mg soluble zinc/ 1 water, which is the threshold for the
astringent taste. At 30 mg/1 the water becomes cloudy, and at 40 mg/1 it has
829
a metallic taste. Acute gastrointestinal distress will occur from concen-
trations in the range of 280 mg/1 and above.
Z_inc_JLnJ[)rinMn£ Water
The levels at which zinc is found most frequently in drinking waters
cannot be considered to be detrimental to human health. The current recom-
mended standard is 5 mg/1. Zinc is an essential trace element in human and
animal nutrition with a recommended daily allowance of 15 mg/day for adults
1119
and 10 mg/day for growing children.
407
-------
TABLE 12-1
Occupational Standards £or_ Zinc and Zinc Compounds in Air-
Country .
Bulgaria
Czechoslovakia
Finland
Germany (entire)
Hungary
789b
Japan
Poland
Romania
Romania
„ . 1790a
Swed en
United Arab Republic
Compound
Zinc oxide
Zinc oxide
Zinc oxide fume
Zinc oxide fume
Zinc oxide
Zinc oxide fume
Zinc oxide
Zinc oxide fume
Zinc pentachlorothlophenate
Zinc oxide
Zinc oxide fume
Leyel_
10 mg/m
3
5 mg/m
3
15 mg/m
5 mg/m
5 mg/m
5 mg/m
c / 3
5 mg/m
3
10 mg/m
5 mg/m
5 mg/m
15 mg/m
Coniments
b
MAC
MAC
MAC
MAC
MAC
MAC
MAC
MAC
MAC
MAC
Syrian Arab Republic
USA, entire
USA, entire
USA, Pennsylvania
USA, Massachusetts
USSR, entire (1975)
USSR, entire (1972)
1363a
World Health Organiza-
tion
Yugoslavia
Zinc chloride fume
Zinc oxide fume
Zinc oxide fume
Zinc chrornate
Zinc oxide
Zinc pentachlorothiophenate
Zinc oxide
Zinc oxide
1 mg/m MAC
3
5 mg/m MAC
10 mg/m3 MAC-
0.2 mg/m3 MAC
3
6 mg/m MAC
2 mg/m3 MAC
3
5 mg/m MAC
5 mg/m MAC
408
-------
TABLE 12-1 - continued
^Unless otherwise noted, data from ILO/WHO Committee on Occupational Health
and the American Conference of Governmental Industrial Hygienists.
-Maximum allowable concentrations, the equivalent of threshold limit value,
are the time-weighted average concentrations to which nearly all workers may
be exposed for eight hours a day, five days a week, for their working life-
time without expectation of any adverse health effects.
—At this level, the maximum exposure could be no more than 30 min.
409
-------
In a survey by EPA, a nationwide water analysis of 591 samples failed
to find any sample that exceeded 4 mg/1. Craun and McCabe reported
that the concentration of zinc at treatment plants was well below the current
-------
Cadmium and lead are common contaminants of zinc used in galvanizing.
Assuming that zinc is dissolved from galvanized water pipes no less than
cadmium, dissolution of zinc to produce 5 mg/1 would be accompanied by some-
thing less than the allowable 0.01 mg cadroium/1 when cadmium contamination
of the zinc is as high as 0.03%. The cadmium may contribute to the acute
gastrointestinal distress following ingestion. Similarly, lead concentrations
would likely be increased by something less than the allowable 0.05 mg/1
when lead contamination of the zinc reaches as high as 0.6%. Based on the
available evidence, the current drinking water standard of 5 mg/1 provides
sufficient protection for consumers.
Zinc _in_Wate_rways_
869
Kopp and Kroner reported that in 1,207 positive tests for zinc on
samples from U.S. waterways, the highest observed value was 1,138/ig/l (from
the Cuyahoga River at Cleveland, Ohio) and the mean was 64 jsg/1. Soluble
zinc was measured in over 76% of all water samples tested. The highest mean
zinc value, 205 A»g/l, was found in the Lake Erie basin, whereas the lowest
mean zinc value, 16>ug/l, was observed in the California basin. In seawater,
the highest concentration of zinc has been found to be about 10 >ig/l.
The toxicity of zinc compounds to aquatic animals Is modified by several
environmental factors, particularly hardness, dissolved oxygen, and tempera-
ture. Skidmore. in a review of the literature on the toxicity of zinc to
fish, reported that salts of the alkaline earth metals are antagonistic to
the action of zinc salts, and salts of certain heavy metals are synergistic
in soft water. Elevations in temperature and reductions in dissolved oxygen
increase the toxicity of zinc. Toxic concentrations of zinc compounds can
affect eh* morphology and physiology of fish adversely. Acutely toxic con-
centrations induce cellular breakdown of the gills, and possibly clog the
411
-------
gills with mucus. Conversely, chronically toxic concentrations of zinc com-
pounds bring about general enfeebleraent and widespread histologic changes to
*
many organs, but not gills. Growth and maturation are retarded.
See Chafer 6, "Zinc in Aquatic Animals," for a detailed discussion of the
effects of zinc on fish.
412
-------
CHAPTER 13
SAMPLING AND MEASUREMENT TECHNIQUES FOR ANALYZING ZINC
SOLID AND LIQJJID,, CAMPLES
Although the analytic chemical literature pertaining to zinc is
quite extensive, only a limited number of techniques have enjoyed the bene-
fit of extensive evaluation by various organizations of analysts and, where
possible, these procedures are emphasized. Attention is paid to the evalu-
ation of zinc in physiologic fluids and other biologic media, potable and
polluted waters, foodstuffs, and soils and plants.
Samples
Levels of zinc in most media of direct interest to environmental
health occur at the trace level and regardless of measurement technique,
require that maximum care be exercised to minimize both loss of zinc from
the sample and the contamination of the metal by zinc-containing reagents,
vessels, etc. Contamination is especially worrisome in the case of zinc
because of its ubiquitous presence in the general environment. Details of
sampling procedure to minimize the above problems have been fairly well
1606
standardized and one is directed to the discussion by Thiers for
specific information.
In most procedures for zinc analysis, some prior treatment of the
sample is necessary and involves destruction of the zinc-containing sample
matrix. In practice, this involves wet-, dry-, and low-temperature ashing.
A detailed discussion of the merits and disadvantages of the various ashing
551
techniques may be found in a recent book by Gorsuch.
Zinc in Biologic Media
Unlike procedures for foodstuffs, soils, plants and waters, standard-
ized official methods for zinc analysis in physiologic media have not evolved.
413
-------
Consequently, the limits of accuracy, precision and detection which have
been developed in representative methods will be noted.
With liquid samples, deproteinization is usually carried out by
using denaturants such as trichloroacetic acid, hydrochloric
1283,1369,1753
acid, or a combination of both. Mineralizing techni-
ques involving wet or dry ashing may also be carried out and these pre-
1140
treatments are invariably necessary for most zinc analyses of bone,
642 661,809,1656
hair, and soft tissue. A newer, highly efficient mode of
541
dry ashing is low-temperature ashing, in which organic material is
removed from the sample by an oxygenated high-energy plasma generated in
a combustion unit by a radio-frequency discharge.
Most spectrophotometric procedures for zinc determination in biologic
media have involved the use of chelating agents which form colored coordi-
1044
nation complexes with divalent zinc. The most common is diphenylthio-
carbazone (dithizone) and zincon.
Dithizone is only moderately specific
for zinc complexation and requires the concomitant use of masking agents
and/or steps for removing interferents, whereas zincon is more selective
to zinc complexation but forms a highly labile and complex chroraophore
which necessitates rapid analysis.
Measurement of zinc in biologic media by atomic absorption
spectrometry (AAS) appears to be the current method of choice in most
laboratories. Sample handling and reagent use steps are greatly reduced
while sensitivity, accuracy, and precision remain superior. With the
373
advent of microsampling techniques in AAS, such as the Delves cup and
414
-------
811
boat techniques,* as well as flaraeless atomic absorption spectral tech-
650
niques —sampling manipulation and lower detection limits for elements
like zinc are being revised downward continually, as is apparent from the
1162,1170
literature. Fluid samples may be deproteinated and diluted or
1031
simply diluted: both approaches enjoy routine use. Because they may
enhance or repress the zinc absorption signal, chemical and physical matrix
effects should be considered in zinc assay by AAS, although appropriate
278
steps can be taken to minimize them. When bone, hair, or soft
tissue is involved, wet or dry ashing is necessary for conventional
atomic absorption spectral "analysis. Further, it is advisable to dry
ash samples in platinum crucibles, as porcelain crucibles have been shown to
624
entrap zinc on their surface. For hair samples, an analytic compromise
usually must be reached between chemical cleaning of hair surface to remove
contaminating metals and leaching of zinc from hair by vigorous cleaning.
A recent text of clinical chemistry techniques describes recommended
1750
methods for zinc evaluation in serum and urine. Serum is deproteinized
with trichloroacetic acid, and the resulting supernatant is analyzed
directly by AAS using a zinc 214 run line. To deprotelnize urine, zinc
is chelated with ammonium pyrrolidino-dithiocarbamate and
extracted with methyl isobutyl ketone^.
*The Delves cup method is a microanalytic procedure for volatile elements
such as lead, cadmium, etc. The sample is placed in a nickel crucible, which
is inserted in the flame portion of an atomic absorption spectrometer. The
burner is equipped with an absorption tube to enhance the collection of
the atomized element present. The tube is fabricated of a ceramic or quartz
material. With the boat technique, samples are placed in shallow boats con-
structed of tantalum and inserted into the flame portion of the instrument.
415
-------
followed by aspiration of the organic phase into the flame of an atomic
absorption spectrometer.
Neutron activation analysis (NAA) appears to be the method of choice where
very high sensitivity is desired and the requisite highly sophisticated in-
strumentation is not available. Unlike other popular techniques, activation
analysis permits determination of many trace metals or metalloids. Examples
of the application of this method to zinc determination in biologic media
469,1165,1210
are available.
Emission spectrography, using direct (dc) or alternating current (ac) arcs
for excitation, is a multielement technique that has been employed for
several years to measure zinc and other elements. But this technique has
been limited by its lack of precision in exacting quantitative work, although
this drawback is being offset by the use of increasingly more stable arcing
1651
accessories.
Spectrofluorometric techniques for zinc analysis, a procedure de-
scribing fluorometric analysis of zinc via the 8-quinolinol complex,
970
demonstrates a marked fluorescence at 517 nm when excited at 375 nm.
988
Anodic stripping voltammetry (ASV) is a newer method for zinc
measurement; it demonstrates excellent sensitivity but requires that con-
siderable pains be taken in sample pretreatment. More comparative data
and research on the range of media studies are necessary to help establish
ASV as a competitive alternative to atomic absorption or colorimetric
techniques.
In media such as soft tissue, in which various means of sampling
are available, the method of expressing data is a particularly vexing
416
-------
problem In zinc and other element analysis. How data are expressed has
hampered correlation of data on zinc levels from various sources. Measure-
ment of zinc on a wet weight of tissue basis is made risky because evapor-
ative loss may be incurred. And losses will vary with time the sample is
exposed to the atmosphere, removal of excess moisture with filter paper,
etc. An assessment of zinc on a lyophilic dry weight basis must consider moisture
uptake by hygroscopic residues. Dry ash analyses may suffer from incomplete
ashing, which will influence the ash weight. Some uniformity may be intro-
duced by expressing zinc content per unit weight of protein, DNA, or other
biochemical marker.
Measuring^ Zinc in JPotable_ and \laste_ Water
Of the various acceptable techniques for zinc evaluation in potable
and polluted waters, those involving spectrophotometry were standardized
and extensively employed first. They operate through colored complexes
formed by coraplexing of zinc with selected chelating agents. Representative
are the dithizone and zincon methods (see Appendix B for details) for
1311a
examination of waters. Colorimetric techniques for zinc in water
are tedious procedures with problems of interference with the chromophore
and stability, difficulties similar to those encountered in sampling biologic
media of the generated complex.
131 la
The presently accepted procedures are polarography and
319,1010,1023,1191b
AAS, the latter perhaps enjoying widest use.
Polarographic methods for evaluating zinc itself or zinc along with
1311a
other metals have been described. Sample treatment includes
preliminary removal of organic matter interferents from 100-ml samples by
successive evaporations to dryness with concentrated nitric acid
417
-------
followed by final manipulation with concentrated hydrochloric acid. Samples
relatively free of organic matter may be handled with a 1:15 ratio of hydro-
chloric acid:water. Zinc is easily measured polarographically and the
method permits routine evaluation of zinc in the 0.01-0.1 mg/1 range.
AAS is especially suited to the evaluation of zinc in
319,1010,1023,119Ib
water samples: in many cases, direct aspiration
of the samples into the burner of the spectrophotometer can be carried out.
The literature about zinc measurement in waters by AAS is extensive
and growing. One is referred to the annual surveys in the Journal
of the Water Pollution Control Federation, biennial review of
water analysis, Analytical Chemistry, and an annual bibliography of AAS
appearing in the Atomic Absorption Newsletter.
Measuring Zinc in Soils
Levels of zinc in most soils are usually in the trace quantity range and
are examined by the analytic techniques previously mentioned. In a stan-
628,1382
dardized spectrophotometric procedure for zinc in soil in which the
dithizonate complex is used, soil samples are ground, sieved, and digested
with various mixtures of concentrated acids. Hydrofluoric acid is
included when removing silicate is desired. Subsequent manipulation includes
the use of citrate to mask iron and copper. Copper is removed as the
dithizonate or is masked by carbamate or thiosulfate. The zinc dithizonate
is extracted into an organic medium at a
pH of 8.3. Polarographic measurement of the zinc content of soil digests
or extracts, prepared as for the spectrophotometric procedure above,also
have been described, although satisfactory results are offset by the
1029
tedium of the overall assay.
418
-------
37
Soil zinc levels may also be obtained by AAS. Soil samples are
either wet-ashed or dry-ashed with a mixture of perchloric,
nitric and hydrofluoric acids or extracted with either hydrochloric
acid or a chelating agent into an organic medium. Using ammonium
pyrrolidino-dithiocarbamate and methyl isobutyl ketone as agents of
chelation and extraction permits a multifold increase in overall sensi-
tivity because of the high aqueous:organic phase ratios as well as the
enhanced element signals found in organic media compared to an aqueous
milieu. An alternative to concentrating is the use of ion exchange
resins, for which a seventyfold increase was reported once.
When zinc levels are desired as part of a multielement profile for
a given sample, emission spectrography with an ac arc
628
has been employed. Samples are treated with a mixture of nitric,
sulfuric, and perchloric acids to destroy organic matter. Sili-
cate is removed by charring with hydrofluoric acid and a prepared buffered
mixture. Aliquots are then placed in the carbon electrode crater for
analysis.
477,478
Zinc has been analyzed in standard rock samples. Zinc in
478
rocks G-l and W-l was measured by optical emission spectrometry (OES),
X-ray fluorescence (XRF) analysis, NAA, ASV, polarography. AAS, and spark-
source mass spectrometry (SSMS). The values corresponded enough from the
various methods to permit nine levels to be established for G-l (45 ppm zinc)
and W-l (82 ppm zinc). To compile data for the new rock standards G-2,
GSP-1, AGV-1, PCC-1, DTS-1 and BCR-1, methods of zinc analysis included
OES, AAS, XRF, NAA, and polarography, and the following means were yielded:
419
-------
477,478
Rock Mean, ppm zinc
G-2 74.9
GSP-1 143.0
AGV-1 112.0
PCC-1 53.0
DTS-1 61.0
BCR-1 132.0
Evaluating Zinc in Foodstuffs
Two recommended methods for evaluating zinc in foodstuffs involve
725 34,1357
spectrophotoraetry and AAS.
In the coloriraetric technique, samples are wet-ashed in the usual
manner with nitric, sulfuric, and perchloric acids. Elements other than
zinc are eliminated in large measure as the sulfides, whereas
nickel and cobalt are removed with various chelating agents. Zinc is
isolated as the dithizonate complex through carbon tetrachloride
extraction and spectrophotometrically measured as such.
Ten collaborating groups have evolved an atomic absorption method
1357
for zinc in foods, and it was deemed accurate and precise. Sources of
zinc sampled in the study included sucrose, soybean meal, and white and
whole wheat flour. Both wet- and dry-ashing steps were employed in the
study. Recoveries were 98-102% with precision of 0.2-2.0% standard
deviation.
NAA has also been applied to evaluation of zinc In foodstuffs.
Of particular interest is the NAA program of the Food and
1597
Drug Administration, in which relatively thorough assessments
are being made of the elemental (including zinc) makeup
of food stuffs of major economic importance. Samples are digested with
420
-------
sulfuric acid and hydrogen peroxide and in the case of foods, radio-
chemical separations are carried out to simplify radionuclide counting.
Analyzing Zinc in Plants
The classic standard procedure for measuring zinc in plants is the
spectrophotometric method, involving a mixed or single color technique
725
measuring zinc as the dithizonate. Samples are dry-ashed at 500-550 C
and the residues taken up in dilute hydrochloric acid. After a series of
extraction steps to rid the analytic medium of interfering ions, zinc is
then measured as the dithizonate, either in the presence of (mixed color)
or in the absence of (single color) an excess of complexing agent.
AAS is rapidly becoming the most popular method of
analyzing zinc in plants. Direct analysis of plant digests may
be performed with little interference from other elements present. This
method has been recommended for plant analysis on the basis of a compara-
tive study between the Association of Official Analytical Chemists (AOAC)
1507
colorlmetric procedure and AAS. Samples—which included corn, sorghum,
wheat, and alfalfa—were wet- and dry-ashed. The accuracy of the
AAS method was found to be as good as or superior to the standard colori-
metric assay.
421
-------
AIR
Zinc is encountered as an atmospheric ingredient entirely in suspended
dust and dirt, so that determining zinc in the atmosphere becomes a matter
of collecting and analyzing atmospheric particulates. Because zinc is often
present in trace concentrations on the order of micrograms per cubic meter
or less, the sampling and subsequent analyses require careful planning.
Care is especially necessary in analytic procedures which determine zinc
and 20-30 other elements simultaneously. This chapter summarizes methods
of sampling airborne particulates and describes some of the advanced proce-
dures for determining their zinc content.
Surveys and Reviews
In recent years, airborne particles have received considerable
attention from investigators of air pollution, and this interest has
coincided with the development of new instrumental methods of chemical
analysis for zinc and other elements present in trace concentrations.
Therefore, a wealth of literature exists on the collection and analysis
of airborne particulates so that a comprehensive bibliographic review
could easily become a treatise in itself. For the scope of this report,
only the most recent and pertinent references have been cited.*
* Chemical Abstracts (1967-1974), Air Pollution Abstracts (1971-1974),
The Analyst (1970-1974), Analytica Chimica Acta (1970-1974), Analytical
Chemistry (1972-1974), Talanta (1973-1974), Staub Reinhaltung der Luft
English edition (1972-1974), Atmospheric Environment (1971-1974), Applied
Spectroscopy Reviews (1969-1974), and the Journal of Radioanalytical
Chemistry (1971-1974) were examined.
422
-------
The sampling and chemical analysis of atmospheric participates have
been the subject of surveys, reviews, and collections of papers *39, 772»
1030, 1489
which do not always describe zinc determinations specifically,
but are useful nonetheless for providing an overview of particle collection
and characterization against which the occurrence and determination of zinc
can be related.
A procedures manual by the Atomic Energy Commission (AEC)16 reviewed
air sampling and elemental determinations, including zinc determined by
1758
AAS. West reviewed the trace metal analysis of inorganic particulates
and concluded that classical methods of gravimetric and titriroetric analysis
are being replaced by instrumental methods offering advantages of selectivity
and sensitivity: colorimetric and spectrophotometric methods, ring oven
methods, emission spectrography, flame photometry, AAS, and polaro-
564
graphy. Greifer and Taylor surveyed methods for determining trace
elements (concentrations less than 100 ppra) in environmentally important
materials such as coal, fly ash, and incinerator ash that could eventually
find their way into the atmosphere as particulates. They described the
determination of zinc and 26 other elements using nuclear methods, SSMS,
XRF and X-ray emission, absorption spectrophotometry, atomic emission
spectrography, voltammetry (polarography), and potentiometry (ion-selective
815
electrodes). Kane and Larrabee reviewed trace analysis techniques
for solids and their treatment of emission spectrography, activation analysis,
SSMS, and XRF is directly applicable to particulate analysis for zinc content.
Zief and Speights 1818 reviewed the techniques of trace element analysis
and described sample handling and analysis by emission spectrography, flame
spectrophotometry, absorption spectrophotometry, neutron activation, and
coulometry. Specific information on zinc determinations was included where
applicable.
423
-------
.^rAt Sampling
To determine zinc in the atmosphere, airborne particulates must be
collected and transported to the laboratory for chemical analysis because
no good methods exist for carrying out trace element analysis in the field.
Atmospheric sampling involves careful planning to assure that the elemental
content of the samples in the laboratory is truly representative of the
atmosphere in the locations sampled. Careless sample collection and handling
may invalidate the most sophisticated and accurate chemical analyses.
The number of samples taken for analysis and the variety of locations
sampled must be sufficiently great to assure that all the trace elements,
and zinc in particular, be represented in their correct concentrations in
the collected specimens. Caution must be exercised to avoid problems
arising from inhomogeneity of the particulates, variation of composition
during sampling, sample alteration during transport, i.e., zinc losses or
zinc contamination, and inefficient collectors which do not capture
all the trace elements. A full treatment of the best places to sample,
the bast times, how big a sample to take, how often and for how long, and
how to collect and retain the zinc and -other trace elements is outsi-de the
scope of this chapter. These matters are essential to a proper inter-
pretation of the analytic data, and they may assume even greater importance
if health considerations or legal implications are involved. Many of these
principles of trace element sampling are discussed by Hendrickson, Fair
et al..A51 and the AEC.1644
There are various techniques for collecting suspended particulates
from the air, and the most widely used are filtration methods and impingers.
Other particle collectors include electrostatic precipitators, sedimenta-
tion bottles, thermal precipitators, and liquid sorption traps.665' 91°
424
-------
The sampling techniques may be divided into two broad categories: those
which collect total participates and those which fractionate the samples
into size classifications. The latter samplers are popular where the
occurrence of zinc in the atmosphere is to be correlated with particle
sizes.
Most particulate samples are obtained with filters, the most popular
filter materials being paper (randomly matted cellulose fiber), membrane
(thin plastic structures with flat, porous surfaces), and glass fiber.
Metallic silver is used at times, as when wet chemical analyses require
acid dissolution of the filter material. Membrane filters (Millipore,
Gelman, Nuclepore, etc.*) collect on their surface (as opposed to the bulk
material matrix)all particles of a size larger than the stated pore size.
They are available in a variety of materials such as cellulose acetate-
nitrate copolyroer, polyvinyl chloride, nylon, and Teflon. The 0.45-um
and 0.8-um pore sizes are widely used for air sampling.
Contrary to popular belief, filters do not work by simply straining
the air stream, although this sieving action is one mechanism by which
particles are trapped. The primary filter action is impact ion, resulting
when the air stream suffers a sudden change in velocity (magnitude and
direction) and the particles continue onward from inertia to impact with
the substrate where they remain entrapped. The relative effects of
irapaction versus sieving are shown by the ability of Whatman 41 filter
paper (widely used in air sampling) to collect about 98% of 0.2 pro particles
from an air stream of 100 cm/sec face velocity although the pore size is
* Trade names have been identified solely to help readers and do not
imply any endorsement or recommendation by the National Academy of
Sciences or the National Research Council.
425
-------
20 pm,910 or the 0.45 ym pore size membrane filter to collect 0.1-um
particles.665 Diffusion, a third filter mechanism, is of minor importance
910
with the thin filters and the relatively high air velocities used. High
volume samplers, such as those used in the U.S. National Air Sampling
2
Network (NASN) draw air through a filter at a rate of 1.1-1.7 m /min (1,100-
1,700 1/min) and are considered a standard method of outdoor particle collect-
7R9
iotx. Low volume samplers such as are used for sampling the occupational
environment operate in the range of 5-50 1/min; and personal (lapel)
samplers, limited in size and weight to small battery-operated devices,
operate at 1-3 1/min.
Impaction devices are samplers which collect particles impinging on
surfaces (as cascade impactors) or in liquid bubblers fas midget Impingers).
Cascade impactors such as the Andersen sampler are made up of a series of
stages (often six or more). The stages consist of plates containing
holes or slots of progressively decreasing size, which effect a size
separation of air-entrained particles. The particles are not sieved by the
holes but are deposited according to the air velocities in each stage; the
largest particles are deposited in the earliest stages where the air velocities
are lowest, and the finer particles are carried toward the later stages where
the air velocities increase with decreasing hole size. The overall volume
rate of flow remains constant, about 28 1/min for the Andersen sampler.
The particles are deposited on paper or membrane filters, sticky tape, glass
or stainless steel surfaces, or agar in Petri dishes.
The trace metal content of filter materials and irapaction surface
coverings themselves have been investigated extensively because of its
353
possible interference with the particulate analyses. Dams etal. tested
426
-------
10 filter materials by NAA using lithiua-drifted germanium (Ge[Li]> detectors
or their spectrometers and found high trace element concentrations in most
of the materials. Cellulose paper and membrane filters had the lowest trace
2
metal content, 7-30 ng/cm zinc, compared with filters such as polystyrene,
which had 60-515 ng/cm zinc. Bodart et al.137 analyzed finely dispersed
filter paper for 10 elements using XRF and lithium-drifted silicon (Si[Li]>
and Ge(Ll) detectors and found 0.13 + 0.04 pg/cm zinc. Birks
et al. determined 17 elements in membrane and paper filters and found
2
0.2-30 ug/crn zinc through XRF measurements, compared to Bowman et al.,
who found 0.007-0.025 jjg/cm zinc by NAA. a Birks et al. also reported
EPA figures based on OES that showed glass fiber filters to have a very
2 1353
high zinc content (160 ug/cm ). Robertson summarized contamination
problems in trace element analysis and gave the zinc content of polyethylene
filters as 25-300 ppb, membrane filters as 2.4 ppm, polystyrene as 4 ppm,
paper as 27 ppm, and tissue paper as 49 ppm; he concluded that paper and
membrane filters were suitable for collecting particulates from the
atmosphere.
Midget impingers also are mentioned as particle collectors in the
910
air pollution literature. They resemble gas scrubbers in that the air
stream from a nozzle la bubbled through a liquid in a container. The
particles are collected in the liquid by an impaction mechanism although
the liquid also has a scrubbing action on the gas stream. Various sizes of
impingers are marketed, the midget impiuger mentioned above accepting
an air flow of 2.8 1/min. These collectors have the disadvantage that
some zinc may leach out of the particles into the liquid to cause a negative
error, or else some zinc from the container walls may transfer over to the
427
-------
particulates to give a positive error (contamination). This collection
device is popular because of its simplicity and low cost.
When particles are to be collected over an extended period of a month
or so and they are large enough (5 pro or larger) to settle out as dust without
the aid of motorized blowers, then they may be sedimented into suitable
containers using standardized methods such as the Intersociety Committee's
780
tentative method for analyzing dustfall from the atmosphere, or the
procedure for collecting and analyzing dustfall proposed by the American
30
Society for Testing and Materials. The advantages of simplicity, low
cost, and unattended operation sometimes may be offset by contamination
from trees, insects, bird droppings, or curious passersby. Sedimented
particles that have settled out on soil and vegetation by the natural
action of winds and rain are sometimes studied as alternatives to collection
in sedimentation bottles. They have provided valuable information on the
198, 765, 891, 1638
amount of zinc and other elements carried in the atmosphere.
Less frequently used collectors include electrostatic precipitators
which impart an electrical charge to the particles so that they are attracted
to an oppositely charged electrode. The collector electrode may be covered
with a membrane filter for ease in handling the collected material, or with
a sample grid for subsequent instrumental analysis, as with an electron
microprobe. This type of collector has a high collection efficiencv and
does not damage the particles physically, but its awkward handling in the
field accounts for the poorer incidence of its use. Cyclone samplers which
impart a centrifugal motion to the air stream and collect the particles in
sharp size fractions above about 5 jjra through inertial effects are becoming available
and are enjoying increased use. Therraan precipitators that impart a thermal
428
-------
gradient to the air stream and collect particles on the colder of two plates
are not used often for particle collection in air pollution studies because
of snail handling capacities and the extensive cleaning required between
samples.
P_eter1Bina_t-iOD__of_Zlnc _i
Most of the work in this field has concentrated on pollution, with
interest directed toward the simultaneous determination of all the metals
present in the atmosphere rather than zinc or any other element in particular
(except for lead and mercury). Large numbers of low cost, multielement
analyses of high sensitivity and good precision and accuracy have been
needed most, and this need is being satisfied by the development of sophisti-
cated computer-assisted instrumental methods. Several reviews ' ' '
are devoted almost entirely to instrumental methods of analysis.
In contrast, classical wet methods of zinc determination as
428
exemplified by the absorption spectrophotometry of dithizone complexes,
are slow and expensive. Also, they are suitable for determining only one
element at a time and subject to interferences from the other elements present
in the particulates. This is not to say that wet chemistry has been discarded;
on the contrary, wet chemical manipulations are indispensible for the
dissolution of particulates for AAS or polarography, for chemical separations
in activation analysis, and for destruction of filter substrates when required.
Analytic techniques reported for zinc are NAA, SSMS, XRF spectrometry, OES,
AAS, and the classical techniques of absorption spectrophotometry and polar-
ography.
429
-------
A^.l.t?:°H Spectrophotometry
Although the literature is heavily oriented toward instrumental methods
which require a minimum of sample handling (and often not even sample
dissolution), this wet-chemistry technique continues to be used because of
its modest apparatus requirements, good sensitivity, and potential for high
accuracy. In absorption spectrophotometry the particulates are dissolved,
interfering elements are removed or sequestered, and the visible or ultraviolet
light absorption of a suitable colored complex is measured in solution at a
characteristic wavelength.
978
Marshall et al. described the determination of zinc oxide
particulates in air by collection on a membrane filter, dissolution in dilute
hydrochloric acid, and colorimetric determination using 4-(2'-thiazolylazo)-
resorcinol (TAR) at a wavelength of 530 nm. Zinc oxide concentrations up to
o
17.4 mg/m were measured, more than twice the threshold limit value set by
the United Kingtom. Sereda and Artemova collected particulates on a
filter, dissolved them in dilute hydrochloric acid, separated out inter-
fering metals on an anion exchange column, eluted the zinc with 0.65 M
hydrochloric acid, and determined it colorimetrically using sulfarsazen*
RBO
at 510 nm. The method detected 0.04 pg/ml zinc. Krylova sampled particulates
at a rate of 10-15 1/min, dissolved them in 1:1 hydrochloric acid, precipitated
interfering metals as dithiocarbamates, and determined zinc colorimetrically
using sulfarsazen at a wavelength of 500 nm. The sensitivity was 0.5 yg
zinc in 5 ml solution. Further information on absorption spectrophotometry
for trace metal determinations has been summarized by Weiss.
* Benzene sulfonic acid, 4-[{4-[3-(2-arsono-4-nitrophenyl)-l-triazenyl]
phenyl} azoj—, monosodiura salt. C. A. Registry No. 1772-02-07.
430
-------
Atomic Absorption Spectrometry
AAS measures the absorbance of light by a sample when it is present
as an atomic vapor at a specific wavelength that is characteristic of
the individual element being determined, here 213.9 run for zinc. A light
source often containing the element of interest incorporated in one of the
electrodes emits the specific line spectrum of this element, and a simple
monochromator isolates the wavelength to be measured. Absorbance is
proportional to the concentration of ground state atoms when the vaporized
sample is introduced into the light path in the atomic state. The production
of vaporized sample in the atomic state usually is accomplished by atomizing
a solution of the sample into a flame, or by vaporizing the solution on a
tungsten or tantalum ribbon, graphite rod, or in a graphite furnace. The
method is rapid, sensitive, and highly specific for metals, although it is
subject to some interferences from anions and organic materials. It is well
suited to accurate determination of zinc in atmospheric particulates.
Atomic fluorescence spectrometry (AFS) is a related analytic technique in
which the optical path is broken at a 90° angle and the stimulated visible
and ultraviolet emission from the atoms is observed rather than their
absorption. When the direct flame emission is observed without a separate
light source, the technique is called flame emission spectrometry (FES).
206
Burrell has reviewed these three techniques.
The analytic procedure of AAS involves collection of particulates on
a filter, acid dissolution of the sample, destruction of organic matter
including the filter, and measurement against suitable standards. In an
extensive review of the use of AAS for determining trace metals including
431
-------
zinc in atmospheric participates, Hwang noted a yearly average of about 0.001
yg/ai^ zinc in urban air samples from 22 cities with incidences up to 1.6 yg/
2
The unsuitability of glass fiber filters Q-60 yg/cm zinc impurity) and the
acceptability of silver and cellulose membrane filter materials CO.01 and
2
0.002 yg/cm zinc, respectively) were mentioned.
Thompson et al. determined zinc and 12 other metals by collection
on glass fiber filters and oxidation with a low-temperature asher to destroy
organic materials (oxygen passing through a corona discharge formed a
strongly oxidizing oxygen plasma which ashed samples at 150 C instead of
the 500 C used in conventional furnace ashing). The particulates were
dissolved in acid and analyzed by AAS at a wavelength of 213.8 nm. The
3 3
technique gave a detection limit of 0.2 ng/m for zinc based on a 2,000 m
air sample. The sample preparation procedure was stated to be suitable
3
for FES as well, for which a zinc detection limit of 120 ng/m was found.
TOT f
Ranweiler and Moyers measured 22 metals in atmospheric particulates
by collection on polystyrene filters using high volume samplers, dry ashing
at 400-425' C, and dissolution with a mixture of hydrofluoric, hydrochloric,
and nitric acids in Teflon bombs. After appropriate dilutions and matrix
corrections by adding lanthanum and cesium, zinc and 21 other elements were
determined. High zinc concentrations in the reagent blanks were attributed
to filter and reagent impurities, and urban air was reported to contain
0.15 yg/m zinc.
The usual means of producing atomic vapor by nebulizing a sample
solution into a flame requires relatively large volumes of solution, a
disadvantage in air particulate analyses in which samples are limited.
Recently, flameless devices have been introduced to improve sensitivity
and decrease sample requirements from milliliters of solution to microliters
432
-------
of solution. Matousek and Brodie167' 986 filtered air particulates directly
into perforated graphite cups containing membrane filters. The cup was
part of a carbon rod furnace in the instrument light path, and direct
determinations of lead and cadmium could be made in air samples as small
as 200 ml. Although zinc was not reported, the technique is mentioned because
it is novel, and appears to be suitable for the determination of many more
metals than lead and cadmium.
The necessity of destroying all organic matter, including membrane,
cellulose, and polystyrene filters, has aroused some controversy over
suitable procedures for accomplishing this without loss of the volatile
zinc. Morgan and Homan, in unpublished work mentioned by Hwang and
1609
Thompson et al., reported the recovery of only 39% of the zinc after
ashing samples in a furnace at 550 C, compared with 96% recovery with a
low-temperature oxygen plasma. Dissolving the membrane filter in acetone
766,767
before ashing in a furnace at 550 C has been recommended. Van
1681
Jlaaphorst et al. summarized the contradictory claims regarding zinc losses
during dry ashing and reported his results of radioactive tracer experiments,
which showed no volatilization of zinc after dry ashing in porcelain crucibles
for 20 h at temperatures to 1,000 C, and no adsorption of zinc on crucible
866
walls to 550 C. Kometani et al. studied 3ry ashing of airborne particulates
on paper and glass fiber filters as a preliminary step in analysis by AAS,
and also concluded that losses were not from volatilization but from formation
of insoluble silicates. It is inferable from these papers that difficulties
encountered with high temperature ashing had a variety of causes and volatilizing
of zinc was less important than formation of insoluble slags.
433
-------
853 861
Kleinman et al. and Kneip et al. correlated the results of
AAS analysis of atmospheric participates with .neteorologic phenomena.
The mixing height of the atmosphere, taken as the height of the inversion
layer and measured daily with balloon-borne temperature and altitude sensors,
was considered to limit the dispersion of pollutants rising into the air
from emission sources at ground level. Multiplying this height in meters
by the wind speed in meters/second yielded a product related to the zinc
2
concentration and defined as the dispersion factor, with unite of m /sec.
Marked seasonal variations for zinc and for the total particulates,
involving spring minimums and summer maximums, were attributed
to changes in the atmospheric capacity for mixing, as indexed by this
dispersion factor. For additional information on applications of AAS,
1300
attention is invited to Price s work.
Optical Emission Spectroscopy
Optical emission methods measure the wavelength and intensity of
visible and ultraviolet radiation emitted from excited electronic states
of atoms introduced into arc and spark excitation sources. They are
suitable for rapid, semiquantitative, multielement surveys of airborne
particulates as well as for accurate quantitative measurements of Individual
elements. An emission spectrographic laboratory utilizes critically aligned,
vibration-free optical and excitation instruments, computer-based data
processing, and personnel with a high degree of skill and training. Never-
theless, the speed, high sensitivity, and broad applicability of OES make
it one of the most popular analytic techniques for determining zinc in the
1390
atmosphere.
434
-------
The technique generally involves powdering or ashing the samples,
mixing them with a buffer and an internal standard, and arcing in graphite
894
electrodes. Lander et al. described the spectrographic determination
of zinc and 11 other elements in airborne particulates. Samples collected
on paper or membrane filters were analyzed without further treatment by
rolling the papers tightly into cylindric graphite sample electrodes and
burning them with high voltage spark excitation in an oxygen atmosphere.
Zinc was measured at a wave length of 213.9 nm. Calculations and tabula-
tion of results were performed by a computer. Individual filters 25 mm in
diameter, sampled at a rate of 14 1/min, contained up to 1.5 ug zinc, with
the detection limit judged to be 0.1 jjg zinc. Coefficients of variation
(the measure of precision) ranged from +9 to + 60% for the individual
elements.
1562
Sugimae collected airborne particulates on a silver membrane
filter, dissolved the filter in dilute nitric acid,precipitated the silver
with hydrochloric acid, evaporated the solution to dryness and determined
zinc and 11 other elements simultaneously by emission spectrography using
dc arc excitation. Zinc was determined in the concentration range 0.13-
3 "^
2.5 yg/m , based on a sampled air volume of 400 nr (50-1,000 ug/filter) with
a precision of + 15%, using the analytic line pair of zinc I 334.5020 nm
and indium I 303.9356 nm.
778
Imae et al. collected particulates on membrane filters for
10 days using a low volume sampler with a flow rate of 20 1/min, burned
the filters with ethanol (C2H5OH) in a quartz boat and further ignited
the carbonized ash in a low temperature plasma to remove the carbon and
residual organic materials. They mixed the ash with indium
and palladium oxides , buffered with lithium carbonate
435
-------
and graphite, and excited the sample in an ac arc to determine zinc and 12
other trace elements. The urban zinc concentration was reported as 0.14-1.02
yg/m3 with a precision of + 16.5%, using the 328.23 nm analytic line for zinc.
Metallic impurities in hydrochloric acid extracts of 105 mm diameter filter
materials also were measured: zinc content was 0.4-1.0 yg for 5 membrane filters,
10.0 vg for 1 polystyrene filter, and 73 yg-10.3 mg for 6 glass filters.
Seeley and Skogerboe^°° avoided problems of filter contamination and
disposal by using the graphite spectroscope electrodes as filters for atmos-
pheric particulates, sampling at a rate of 1 1/min. They determined zinc
and 13 other elements directly by emission spectroscopy using dc arc excita-
tion with indium as an internal standard. Their detection limit for zinc
was 3 ng when the analytic wavelength of 334.502 nm was used.
914
Lee et al. reporting on the results of emission spectrographic
analyses of particulates collected by the National Air Surveillance Cascade
Impactor Network (NASN), emphasized the variation of metal concentrations
with particle size. Particulate matter collected biweekly over 24-h periods
with a 5-stage Andersen sampler was extracted with nitric acid, buffered
with lithium chloride, and analyzed on a 2-meter spectrograph employing
a rotating disc and high-voltage spark source. Indium and yttrium were
taken as internal standards. The annual concentration levels observed for
3
zinc were 0.1-1.7 yg/m , which were intermediate between highs for iron
and lead and lows for nickel and vanadium. Particle size distributions were
correlated with metal concentrations for zinc and 11 other elements. About
70-80% of the zinc particles were smaller than 2 ym, and 40% were smaller
than 1 ym. No seasonal pattern was discerned, but a geographic pattern
hinted at possible urban sources of zinc in the atmosphere.
436
-------
X-Ray Fluorescence Spectrometry
XRF spectrometry, also known as X-ray emission Spectrometry, measures the
characteristic photon or X-ray emissions that result when higher energy
electrons fall into a K or L shell whose electron was previously ejected
by irradiation with high energy photons or particles. The energy of the
emitted X radiation is characteristic of the elements present in the
sample, and the frequency with which the transitions occur is proportional
to the quantity of element present. All elements above sodium can be
determined rapidly without any sample preparation.
The instrumentation required includes an excitation source which
may be an X-ray tube, radioisotope, or other source of electrons, protons,
or ions; a sample holder; an analyzer to resolve the emitted X rays into
an energy spectrum by wavelength dispersion (such as crystal diffraction, or
bandpass filters) or by energy dispersion; and an X-ray detector. Detectors
used with wavelength dispersion spectrometers may be scintillation detectors,
or gas-filled or gas-flow proportional detectors. Detectors used with energy-
dispersive spectrometers are the solid-state Si(Li) detectors, with an out-
put pulse height proportional to the incident energy. A great
many combinations of source, analyzer, and detector are possible.
Several very readable introductions to the principles of XRF analysis
114, 115, 531
exist. Four types of excitation (X-ray tubes, fluorescers,
radioisotopes, and high energy ions) are compared, as well as two types
of data analyzers (crystal spectrometers and energy-dispersive detectors)
for determining 10-20 elements including zinc in air particulates. A
multichannel crystal spectrometer instrument is recommended as the optimal
technique for measuring at least 12-14 elements simultaneously. If only
437
-------
one or a few elements are of interest, then energy dispersion analysis with
a low power X-ray tube or high activity radioisotope and a simple proportional
counter may be an adequate alternative to sophisticated laboratory instrumenta-
tion. Zinc detection limits for 100-sec measurements of prepared standards
2
ranged from 51 ng/cm for X-ray tube excitation and wavelength dispersion
2
to 1,400 ng/cm for isotope (iron-55) excitation and energy dispersion.
XRF results for zinc and 16 other elements were comparable to atomic
absorption measurements made on the same particulate samples. Studies of
115
elemental impurities in filter materials have been mentioned earlier.
1337
Rhodes et al. compared energy-dispersive XRF spectrometry
using radioisotope sources with AAS for determining zinc and 16 other
elements in particulates collected with high volume samplers on cellulose
3
filter paper. Air pollution data presented for urban areas averaged 0.13 ug/m
zinc, compared with a national average of 0.67 yg/m . The X-ray analyses
produced consistently lower results than the atomic absorption determinations,
and this discrepancy was thought to be an effect of particle size of the
samples.
958
Luke et al. compared XRF spectrometry with AAS for analyzing
particulates collected on filter paper. Zinc and six other elements were
determined by direct X-ray analysis, X-ray analysis of samples after wet
chemical separations, and AAS. The results, between 2-100 yg/cm2 for zinc,
were comparable for-all 3 techniques.
924
Leroux and Mahmud analyzed particulates collected in an urban
area on membrane filters and found 0.08-4.03 yg/m3 zinc. The analyses
529
took only 5 min/element. Giauque et al. used X-ray-induced XRF
spectrometry to determine zinc and other elements in particles collected over
438
-------
2-h intervals for 24 h with a 4-stage impactor. Detection limits for zinc
3 ^
were stated to be 4 ng/m on membrane filters and 2 ng/m on Mylar filters.
Diurnal charts showing changes of zinc concentrations with time were presented
230
for several cities. Cares described X-ray spectrometric procedures for
airborne dusts and included a detailed method for preparing reference
OQg
standards, but zinc was mentioned only in passing. Dittrich and Cothern
analyzed trace metals in urban atmospheric particulates using a high volume
air sampler to collect particles on filter paper. A y-ray source (americium-
241) and a bremsstrahlung excitation source (promethium-147) both were used
to produce X rays. Data for zinc and six other elements were presented
3 1071
as counts rather than as yg/m . Mitsugi et al. collected air particulates
on filter paper using a high volume sampler, and determined zinc and lead
concentrations directly; also, samples collected with an electrostatic precipita-
tor were briquetted with boric acid and analyzed. Zinc concentra-
3
tions in the range of 0.25-1.06 yg/m agreed with results yielded by AAS.
137
Bodart et al. analyzed particulates on filter papers by irradiating the
filters with 1.7 million electron volt (MeV) protons from a low energy
Van de Graaff generator, and detecting XRF with Si(Li) and Ge(Li) detectors.
Standards were prepared from pure salts and powdered paper filters. Findings
were presented for 10 elements: results for zinc fell in the 1.40 + 0.14
2
yg/cm range.
Mizohata and Mamuro ' used americium-241 excited, energy-
dispersive XRF spectrometry to determine zinc and 11 other elements in
airborne urban dust samples. Corrections for sample loading on membrane
filters were made by comparing the X-ray data with determinations from instrumental
NAA. (The measured X-ray emission of zinc decreased about 20% as the mass
2
loading of the filter increased from 0.2 to 7.5 mg/cm because of self-
439
-------
absorption in the sample.) Actual dust samples collected with a low
2
volume air sampler were found to contain 1.1-48.7 yg/cm zinc, as measured
by the two techniques.
625
Hammerle et al. compared XRF spectrometry with NAA for determining
10 elements in atmospheric particles. The zinc analyses for both techniques
3
agreed within 6.7% of the average of 0.28 yg/m found in urban particulates.
314
Cooper compared particle- and photon-excited XRF determinations of
zinc and about 20 other elements in urban aerosols collected on filter
paper, and in rural aerosols collected on Mylar film in an impactor. The
samples were cemented as thin samples or briquetted as thick samples, and
analyzed by excitation with either high energy a-particles (80 MeV and 30
Me'V) or protons (4 MeV and 2 MeV). Cooper also studied photon excitation
with a molybdenum target X-ray tube, and with high intensity radioisotope
excitation using iron-55 and cadmium-109 sources, and measured the energy
spectrum with a Si(Li) detector. Zinc was reported at a concentration of
o
1-5.6 ng/m in typical urban particulates. Photon excitation methods were
concluded to be significantly more sensitive than high energy a-excitation,
with the additional advantages of small size, easy portability, and applica-
bility to a wide variety of samples for trace element analysis of environ-
mental particulates.
This brief survey of the application of XRF spectrometry
to the determination of zinc in air particulates shows that it is a popular
analytic technique because 20 or so elements may be determined simultaneously
and directly on filters without prior sample manipulations such as acid
dissolution, extraction, or ashing. What cannot be emphasized strongly
440
-------
enough is the necessity of having reliable calibration standards because
the technique is substrate-sensitive. Dried solutions of pure salts on filters
may not simulate collected particulates closely enough, because the latter
are mixtures of many different inorganic and probably organic materials with
different particle sizes and shapes. It seems reasonable to use a uniformly
deposited, exhaustively analyzed natural material for an X-ray calibration
standard to supplement the data obtained with pure salts.
Neutron Activation Analysis
Activation analysis is similar to XRF or emission spectrometry except
that it is the atomic nucleus rather than an extranuclear electron which
is energized by irradiation with energetic particles or photons. The
emitted radiation of the nucleus as it decays radioactively is measured. The
energy and type of the emitted radiation as well as its rate of decay identify the
nucleus, and the intensity of the radiation is a measure of the amount of
element present. The technique generally involves irradiation with thermal
neutrons, chemical separation of the activated species, and measurement of
the emitted radiation, preferably with solid-state detectors coupled with
multichannel analyzers and computer-controlled data processing to convert
spectral counts and energies to elemental concentrations. Although a variety
of neutron sources is available (neutron generators and isotopic neutron
sources such as californium-252), the trace metal concentrations in air-
borne particulates are so low that high thermal neutron fluxes of the order
12 13 2
of 10 or 10 n/sec/cm are required and these fluxes are available only
from nuclear reactors. Chemical separations of activated species may be
required if the energies of emitted radiation from many elements overlap;
-------
however, the high resolutions available from the new solid-state Ge(Li) y~
detectors make direct sample assays possible without the need for wet
01c 83s
chemistry. Two reviews ' give a concise introduction to the subject.
352 354 1311
Dams et al. ' and Rahn et al. have described the determina-
tion of 33 elements in air pollution particulates directly on filters without
prior chemical separations. Particulates collected on polystyrene filters
were irradiated for 2-5 h (for long-lived isotopes such as zinc) in a
nuclear reactor at a thermal neutron flux of 1.5 x 10^ n/sec/cm2 together
with a standard mixture of elements in a polyethylene bottle. After
irradiation, the samples and standards were counted with a Ge(Li) detector
coupled to a 4,096 channel pulse height analyzer. The spectra were recorded
on seven-track magnetic tape for subsequent data reduction by computer. Zinc
was determined as the 13.8-h zinc-69 metastable radioisotope, using the
438.7 kiloelectron volt (KeV) y-ray radiation. The detection limit for zinc
was 0.2 yg, which corresponded to an urban air concentration of 0.02 ug/m^
for a 24-h sample. The authors commented that emission spectrography would
o
detect about 0.24 ug/m zinc in such a sample. Suspended particulates
o 3
contained 180-1,690 ng/nr zinc (standard deviation, 10-300 ng/m ). Dams and co-
353
workers used this analytic technique to evaluate filter materials and
study airborne trace element distributions. ' 1311 2inc and antimony
exhibited similar well-defined distribution patterns which were correlated
to geographic areas.
887
Kuykendall et al. used automated instrumental NAA to determine
zinc and 39 other elements in air filter samples. Particulates collected
on filter paper with a high volume sampler operating for 24 h at a flow
rate of 1,223-1,631 m3/day were irradiated in a neutron flux of 5 x 1012
f\
n/sec/cm for 8 h (for long-lived radioisotopes such as zinc), and counted
442
-------
directly with a Ge(Li) detector coupled to a 3,200 channel pulse height
analyzer with computer-compatible magnetic tape readout, sample changer,
and system controller which permitted continuous unattended counting of
many samples. Zinc was reported in the range of 48-1,453 ng/m3, with
3
a mean of 410.5 ng/m , which was well above the stated detection limit of
120 ng/m3.
1440
Schramel et al. determined zinc and 11 other elements in dustfall
samples collected for 1 mo periods in polyethylene bottles, thereby avoiding
the limitations of filter blanks. The particulates were sealed in quartz
ampoules and irradiated for 24 h in a thermal neutron flux of 1013 n/sec/cm .
They were cooled for 3 days, and the zinc group was chemically separated
from the other elements and counted with a Ge(Li) detector connected to a
4,096 channel pulse height analyzer. High zinc concentrations were noted
in areas of heavy vehicular traffic (16.26 mg/g), which were thought to
be caused by airborne rubber particles from automobile tire wear.
1788
Winchester discussed the significance of the results of NAA as
they related to occurrence and transport of pollution and natural substances
in the air. He recommended that sampling from aircraft be seriously consid-
ered for acquisition of improved data. ,
Particulates have been collected with excellent size fractionation
49 539 548
on polyethylene films in a cascade impactor ' * and analyzed for
18 elements by instrumental NAA. Zinc occurred predominantly in the finest
particles, in 100 times the concentrations found in the coarsest
particles. Actual particle sizes were not determined. Statistical
correlations with elemental abundances from known sources such as the
earth's crust, marine aerosols, coal and oil combustion, and automobile
443
-------
exhausts did not explain the zinc enrichment in the atmosphere. Thr- fact that
the greatest amounts of xin< were found In I he smallest particles seemed to
vaporization followed by condensation, that Is, a combustion source, ft
539
was suggested that tire dust generated by abrasion would most likely
consist of large particles, so that the hypothesis of tire wear as a source of
zinc ' would not be consistent with measured particle size distributions.
1247
Pi Hay and Thomas described the analysis of airborne participates
using sequential air sampling onto special high-purity filters, high flux
13 2
(3 x 10 n/sec/cm ) thermal neutron Irradiation, and high-resolution Y-ray
spectrometry to determine zinc and 1.8 other elements. Instrumental neutron
activation for multielement analysis of airborne particles has been described
252, 351, 547, 556, 795
by other investigators.
49
Aras et al. determined 14 elements Including zinc by an Instrumental
photon activation technique. Atmospheric particulates were Irradiated
with bremsstrahlung from 35 mev electrons produced with a linear accelerator,
and the emitted y-ray radiation was counted with a Ge(Ll) detector.
Zinc was determined by the zinc-68 (y,p) copper-67 reaction, and was reported
3
In the range of 0.09-0.22 jjg/m at several sampling locations. The detection
3 -53
limit for zinc was reported to be 3 ng/m , compared with 4 x 10 ng/m for
3
Instrumental NAA, and 0.0025 ng/m for flame methods. Therefore, it was
concluded that Instrumental photon activation analysis was a useful technique
for analyzing particulates. Instrumental NAA Is the method of choice for
many Investigators who determine zinc and other elements simultaneously in
air particulates. The wide applicability of this analytic technique has
379
been thoroughly demonstrated.
444
-------
Mass Spectroroetry
Ma0e spectrometry of airborne particulars in usually taken to mean
S8MS, in which samples introduced as electrodes in an evacuated chamber
are pulsed with a 1 megahertz (MHz) radio frequency (rf) voltage of 20-80
keV to yield ions of all the elements present. The ions are accelerated
with a 24-kV dc potential, focused In an electrostatic sector, and dispersed
according to mass;charge ratio in a magnetic sector for photographic or
electronic detection and measurement. The technique Is highly sensitive:
it can detect all the elements simultaneously including adsorbed gases.
Furthermore, the spectrum is simple (consisting mainly of singly-charged
positive ions), and sample preparation is minimal, requiring only 10-100
mg of solids for analysis. Some limitations include spark-sourre instability
leading to nonuniform ion production) and the relatively poor sensitivity
and reproduclbility of photographic plates used for detection and measure-
ment,
172 U6
Brown et al. and Brown and Vossen reviewed research on solid-
state mass spectrometry and presented analyses of particulars on membrane-
filters. Table 13-1 lists 46 elements Including zinc, which were determined
172
in 2.65 mg particulates collected on a membrane filter.
445
-------
TABLE 13-1
Analysis of Particulate Matter from Mine
Interior for Trace Elementsa
Element
Uranium
Bismuth
Lead
Samarium
Neodymium
Praseodymium
Cerium
Lanthanum
Barium
Iodine
Tellurium
Cadmium
Silver
Molybdenum
Niobium
Zirconium
Yttrium
Strontium
Rubidium
Bromine
Selenium
Arsenic
Germanium
Concen t rat ion
0.006
0.02
0.07
0.01
0.12
0.03
0.12
0.06
1.2
0.01
0.01
0.01
0.02
0.04
0.02
0.16
0.02
0.77
0.02
0.03
0.02
0.07
0.05
Element
Gallium
Zinc
Copper
Nickel
Cobalt
Iron
Manganese
Chromium
Vanadium
Titanium
Scandium
Calcium
Potassium
Chlorine
Sulfur
Phosphorus
Silicon
Aluminum
Magnesium
Sodium
Fluorine
Boron
Lithium
Concentration
0.18
1.1
2.3
2.0
0.03
210.0
0.12
0.45
0.44
42.0
0.08
35.0
44.0
0.61
8.3
0.81
780.0
750.0
2.1
2.0
0.005
0.01
0.02
1 79
aData from Brown et al.
Weight retained on filter in yg; total sample weight 2.65 mg on Millipore filter.
446
-------
In an interesting variation, Perry1233 mentioned that trace metals on
filters might be converted to volatile metal oxinates or S-diketonates,
extracted into an organic solvent, and identified and determined quantita-
tively using conventional high-resolution mass spectrometers of the types
used to analyze organic materials. Standardization with pure metal alkyls
or chelates would be required. Only findings for lead tetraethyl
were presented. The use of such metal-organic complexes has been proposed
for gas chromatographic analysis. •"•' Since zinc forms volatile chelates with
a variety of 6-diketones such as 2,4-pentanedione, hexafluoro-
2,4-pentanedione , 2,2,6,6-tetramethyl-3,5-heptanedione,
917
and 2-thio-2,4-pentanedione,
it may be possible to determine zinc by mass spectrometry. Such determina-
tions apparently have not been reported yet.
Despite its broad capabilities, SSMS does not appear to have been used
extensively to analyze airborne particulates.
Voltammetry
In common with other techniques requiring sample dissolution, buffering,
and other wet-chemical processing, electroanalytic methods are not described
in the air pollution literature as frequently as multielement techniques
that require few sample manipulations. With such attractive features as
high sensitivity and selectivity, low cost, and a modest multielement
capability, voltammetry should be used more extensively in environmental
analysis than it is.
In voltammetry (polarography), current-voltage relationships measured
during electrolysis are used to identify and quantitate the ions reacting
447
-------
at one of the electrodes (generally a dropping mercury electrode). Many
variations of the basic technique have been developed, and instruments are
available for conventional dc polarography, derivative dc polarography,
cathode ray polarography, ASV, and ac, square wave, and pulse polarography.
The various names refer to the types of voltage applied to the electrodes
and the forms of data presentation. The highest sensitivity is afforded by
ASV, in which the elements are plated from solution onto a cathodic electrode
and then oxidized into solution by applying a voltage scan in the anodic
direction. The peak currents arising from the oxidation of the ions of
interest are measured to yield results down to nanogram and picogram
concentrations in solution.
895
Landry described early work on polarographic zinc determinations
307
in the atmosphere. More recently, ASV has been used to determine
trace amounts of zinc in airborne particulates: particles were collected
on a membrane filter, which was ashed to destroy the filter and organic
materials; the residue was digested in nitric and hydrofluoric acids. The
solution was diluted and analyzed by stripping analysis, using 0.1 M
potassium nitrate as supporting electrolyte, by plating at -1.2 V
for 3 min, then scanning from -1.2 v to + 0.1 V against a standard calomel
3
electrode. Results for zinc were 0.020-0.93 yg/m ; they were
compared with atomic absorption analyses carried out on the same samples.
989
Matson et al. used ASV to determine zinc in tissue, blood, urine, and
hair below the nanogram level and found excellent agreement with results
from AAS and NAA. Their technique employed perchloric acid digestion,
addition of 0.5 M salt (Nad), 1 M sodium acetate, and
0.5 M ethylenediamine, plating for 30 min at -U3 V, and then stripping
448
-------
at 66 mV/sec to a potential of +0.1 V against a standard calomel electrode.
The method should be suitable for analysis of particulates.
To sum up, the variety of analytic methods and sampling procedures
that have been used complicates comparing results from different investigators
that might allow the testing of theories about zinc sources, atmospheric
823
transformations, or removal mechanisms. Katz has suggested the initiation
of collaborative testing programs to evaluate the various analytic methods
for specificity, selectivity, sensitivity, range, precision, and accuracy;
however, very often these programs evaluate the capability of the
operators rather than the analytic methods used. Von Lehmden et al.
attempted to evaluate methods of analysis for coal, fly ash, fuel oil, and
gasoline: 9 laboratories determined concentrations of zinc and 27 other elements
in the same samples using SSMS, OES, INAA, AAS and ASV. Table 13-2 presents
interlaboratory results obtained for these methods of analyzing zinc. Because the
wide variations reported are a consequence of differences in sample handling
and instrument operating procedure, results are difficult if not impossible
to interpret. Comparisons of data from different laboratories (for example,
the different literature cited in this chapter), could lead to erroneous
conclusions.
449
-------
TABLE 13-2
a
Collaborative Test Results for Zinc Determinations in Various Materials
Laboratory
1
3
6
1
3
2
3
4
5
8
3
9
1
7
Method of
Analysis
b
SSMS
SSMS
SSMS
OESC
OES
INAAd
INAA
INAA
INAA
INAA
AAS6
DRES^"
AAS
ASV9
Fly
Coal, Ash,
ppm ppm
100 200
5 1,000
6.6 330
100 100
50 200
-
-
100
-
-
600
350
-
•— turn
Fuel Premium
Oil, Gasoline,
ppm ng/ml
0.5 0.2
1.0
-
0.4 0.2
2.0
1.4 0.36
1.3
3.0
-
-
-
-
2.0 1.0
0.12
Low-Lead
Gasoline
US/ml
-
4.0
-
-
-
0.43
-
-
-
-
-
-
-
0.096
a 1705
Derived from von Lehmden et al.
& SSMS - spark-source mass spectrometry
Q OES - optical emission spectrography
" INAA - instrumental neutron activation analysis
e AAS - atomic absorption spectrometry
J" DRES - dissolution followed by dc arc excitation in emission spectroscopy
^ ASV - anodic stripping voltammetry
450
-------
CHAPTER 14
SUMMARY
PROPERTIES AND USES OF ZINC
As the fourth most widely used industrial metal, zinc serves the consumer
in a form not easily recognized in the end product. It is a corrosion protec-
tion for steel (galvanizing); an alloying metal with aluminum, magnesium, and
titanium in mass-produced precision parts (die-casting); an alloy with copper
for brasses; a chemical for white paint pigment; and a vital ingredient in com-
pounding rubber products. Zinc compounds have long been used pharmaceutically,
although some newer applications in physiology and medicine are still being
evaluated.
NATURAL SOURCES AND DISTRIBUTION OF ZINC
Zinc is found in most soils, but some areas are deficient in it. Differ-
ences of zinc in soil and water can influence the zinc content of
plants and animals found in these areas and in the products derived from them.
MAN-MADE SOURCES OF ZINC
Metallurgic operations, primarily mining and smelting of lead and zinc ores,
contribute appreciable zinc contamination to air, water, and soil. However,
this contamination is normally limited to areas near point sources. Few data are
available on airborne concentrations of zinc near metallurgic operations. How-
ever, evidence of elevated soil and vegetation zinc concentrations near these
operations implies that airborne levels of zinc in these areas are or were
appreciably elevated. The closing down of many older primary zinc operations
has reduced environmental contamination and presumably new operations will have
better particulate controls.
451
-------
Available data, although scanty, indicate that airborne zinc concentrations
2
in the United States away from point sources are generally low (less than 1 /ig/m ).
Zinc in drinking water in the United States rarely exceeds the drinking water
standard of 5 mg/1. Data are conflicting on the contribution of mobile sources
to zinc contamination near roadways. If mobile sources do contribute markedly
to the zinc roadside dust, the zinc probably is generated from zinc compounds in
tires and motor oil. Sewage sludges also may contain high concentrations of zinc
and other metals. Such sludges may not be suitable for indiscriminate use on
agricultural land.
Few data are available on the zinc content of solid wastes and the contribu-
tion such waste may make to total environmental zinc levels. Information that
exists on the quantity of zinc-65 released into waters from nuclear power plants
would seem to indicate that levels are within acceptable limits.
ZINC IN PLANTS
Aquatic Plants
Trace amounts of zinc are essential for normal growth and development of
aquatic plants. Uptake of zinc is accomplished by ion exchange and metabolic
assimilation processes. Aquatic plants can accumulate much more zinc by sorp-
tion than is needed for metabolic activity. Rooted plants may take up zinc from
sediments as well as from the ambient water, but zinc is usually accumulated by
the plant in relative proportion to zinc in the ambient water. Zinc concentra-
tion factors in plants as compared to the ambient water solution may range from
a few orders of magnitude to 19,000. Therefore, zinc concentration of aquatic
plants can range from a fraction of a ppm to several thousand ppm. Zinc con-
centrations in aquatic plants vary seasonally, and the variation is probably
caused by differences in availability and content of zinc in ambient waters as
well as factors such as growth rate, supply of other nutrients, and temperature.
452
-------
Excessive levels of zinc in waters can bring on zinc toxicoses in aquatic plant
communities. Tissue sampling and analysis techniques are being developed to
establish threshold values for zinc toxicity to aquatic plants. The same
analytic data may be used to determine the overall zinc nutritional status in
aquatic plants. A primary function of zinc in aquatic plants is as a cofactor
for several metalloenzymes and possibly as a stabilizer of the integrity of
plant ribosomes. Aquatic plants (mainly algae) have been used extensively to
elucidate the metabolic functions of zinc in plants.
Terrestrial Plants
The recognition of zinc deficiency and an understanding of essential func-
tions of zinc in terrestrial plants have been commonplace since 1900, and zinc
deficiency is now the most common micronutrient deficiency in the United States.
Plants vary widely in sensitivity to zinc deficiency, toxicosis, and tolerance.
Climatic and soil factors affecting zinc availability to plants are: amount of
soil reserves; soil pH; extensiveness of root zones; microbial and soil organic
matter content; soil temperature and moisture; and interactions with other
elements. Most soil zinc moves to plant roots by diffusion—convection plays a
minor role. Soil intensity and capacity as well as availability of natural and
synthetic chelating agents influence the movement of zinc to plant roots in soil.
Zinc is taken up by plants as the divalent cation, and both active and passive
uptake mechanisms are present. The active metabolic component is most important
for continued zinc uptake by plants. Zinc is translocated in the xylem of
plants as the divalent cation or as a weak metal-organic complex. Some zinc is
redistributed from older to newer tissues through the phloem. Evidence exists
for considerable interaction of zinc with phosphorus, iron, copper, and other
elements in the plant. These interactions influence the rate and degree of zinc
translocation. Zinc deficiency is often observed in plants that contain less
than 20 ppm zinc; the normal plant concentration is 25-150 ppm zinc. Toxicosis
453
-------
often occurs at concentrations above 400 ppm zinc. Zinc deficiency can be
overcome by foliar or soil fertilizer applications of soluble zinc salts. Zinc
is intermediate in toxicity to plants among the heavy metals. Several species
of plants that are extremely tolerant of high zinc concentrations in soil have been
identified and provide an opportunity to establish vegetation on zinc-contaminated
soils. The best defined role for zinc in plants is as an enzyme cofactor in
various metalloenzymes. The greatest amount of research has been conducted on
carbonic anhydrase, but the necessity of zinc to several metalloenzymes is well
established. There is also evidence that zinc may function in stabilizing
ribosomes.
ZINC IN AQUATIC ANIMALS
Although zinc is ubiquitous in aquatic organisms, the environmental and
physiologic mechanisms controlling the biologic availability and accumulation
of zinc in tissues are not well understood. In fresh fish muscle, zinc normally
occurs in a concentration range of 3-30 ppm. Similar concentrations are observed
in muscle tissue of many crustaceans and mollusks, particularly in those from
open ocean waters. Organisms from coastal estuarine waters, however, tend to
exhibit higher and more variable concentrations of zinc. Oysters have a partic-
ularly high affinity for zinc, occasionally accumulating the element to more than
1,000 ppm. The highest concentrations in oysters are usually found in specimens
from environments low in salinity. In mussels, scallops, and freshwater bivalve
mollusks, zinc is strongly localized in certain organs, whereas in oysters the
metal is rather uniformly distributed throughout all tissues. Fish, decapod
crustaceans, and certain species of polychaete worms appear able to regulate
tissue concentrations of zinc.
Since aquatic organisms spend their entire lives immersed in a complex
chemical milieu, it is difficult to distinguish dietary sources, integumentary
exchange processes, and excretory and secretory mechanisms. An understanding of the
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physicochemical behavior of zinc in the aquatic environment is therefore a
requisite for metabolic studies of aquatic organisms. Numerous studies have
suggested that various chemical forms of zinc (including organic complexes)
occur in natural waters, but their role in biologic accumulation and metabolism
of zinc in organisms has not yet been defined.
The disposal of industrial and municipal wastes in the aquatic environment
has made zinc toxicosis a potential problem for aquatic organisms. Short-term
acute toxicosis experiments have indicated that the median lethal concentration
(LCc0) for some species may be in the range of 100-300 ng zinc/1. In fish,
acute metal toxicosis has been attributed to precipitation of mucus on gills
or to other cytologic damage to gill tissue. Systematic studies of the effects
of environmental variation on toxic or sublethal effects of zinc have not been
performed for most aquatic organisms.
ZINC IN HUMANS
Zinc is an essential element found in every human tissue and tissue fluid.
trace only to iron.
Among the/transition elements, the concentration of zinc in the body is second/
Approximately
/90% of total body zinc is found in muscle and bone, but the highest concentration
of zinc is found in endocrine glands, particularly the gonadal system, and in
sensory receptors, particularly the retina of the eye. Only a small amount of
total body zinc is carried in the blood, with the active fractions bound
primarily to either albumin or amino acids, particularly histidine. Zinc is
normally excreted in the urine bound to amino acids, but only a small fraction
of the circulating zinc is excreted daily. Most dietary zinc is excreted in the
feces as a result of lack of absorption or because of resecretion from bile.
Zinc is a critical constituent of DNA polymerase. without it, protein
synthesis does not proceed normally and cell division appears to be
abnormal. Zinc deficiency in humans is commonly associated with abnormalities
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in those systems in which rapid cell division occurs; hence, growth retardation,
hypogonadism, and abnormalities of the gastro-intestinal tract are common com-
plaints. Zinc also appears to be particularly important for growth and develop-
ment in utero and in early life.
The specific manner by which zinc is transported across the gut is not known,
but the process may involve the formation of a low molecular weight organic
zinc chelate which moves across the gut primarily in several areas of the small
bowel. Studies with zinc-65 suggest that about 65% of this isotope is absorbed
by normal humans; however, there is an extremely broad range of absorption of
this isotope, a phenomenon quite different from the absorptive characteristics
of other trace metals. Zinc malabsorption, which occurs in several gastro-
intestinal diseases, is associated with absorption of less than 30% of the
zinc-65 presented orally.
Although the specific functions of zinc in various organ systems are unclear,
most systems appear dependent upon its presence. In the endocrine system, zinc
is associated with the release of several pituitary hormones, the prolongation
of action of adrenocorticotropin and insulin and an effect on prostate and testes
that appears important in spermatogenesis. Many hormones influence zinc
metabolism, and the bases of these changes are related to effects not fully
understood.
Zinc is involved in muscle function. Muscle represents the largest single
body pool of zinc and the metal may be associated with potentiation of contract-
ibility. Zinc is associated with key enzymes involved with the function
of several organ systems, including liver and retina, and with vitamin and hematopoetic
function. The association of zinc with alkaline phosphatase and the prominent
role of this enzyme in the function of receptor membranes suggests a ubiquitous
role for zinc.
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In sensory systems, the enzyme needed to convert retinol to retinal in
the retina is zinc-dependent; hence visual processes are, in part, dependent
upon this metal. In saliva, the zinc protein, gustin, may be important in the
growth and differentiation of taste buds. With the isolation of taste bud
receptor membranes from the cow, the zinc-dependent enzyme alkaline phosphatase
appears to be the most highly concentrated enzyme; demonstration of specific bind-
ing of sugars and other tastants to the isolated taste bud membrane are zinc-
dependent. Zinc depletion in humans or animals is almost uniformly associated
and taste loss,
with anorexia conditions usually reversed following administration of zinc.
Although zinc is the fourth most prevalent cation in all brain tissue,
little is known of its function. Its location in the cerebellum and in the limbic
system and the production of cerebellar dysfunction and mental dysfunction with
acute zinc depletion in humans are well established. Severe mental aberrations in
acutely zinc-depleted human subjects and poor performance in learning situations
by animals made zinc-deficient suggest that zinc is associated with higher
brain function as well as several neurophysiologic functions.
ZINC IN THE DIET
Zinc Deficiency
Feeding animals diets low in zinc has decreased growth in all species tested.
Other signs of zinc deficiency frequently observed in young animals are para-
keratosis, hyperkeratinization, and impaired testicular development. Animals in
less rapid stages of growth may have no signs of deficiency other than poor
appetite and decreased growth. Wound healing has been impaired in adult animals
on a low zinc diet; and reproduction, at least in the female rat, has been
severely affected. Since food consumption is consistently reduced in zinc
deficiency, some of the problems apparently caused by a low zinc intake are
a secondary effect of decreased food consumption.
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Sources of Zinc and Amounts Required in the Diet
Zinc in the diet primarily comes from protein. Meat, legumes, and whole
grains are good sources of zinc. Zinc intakes considered adequate for various
animals range from 20-50 mg/kg diet. Factors other than the actual content of
zinc in the diet affect its adequacy, however, since deficiencies have developed
in animals eating diets with presumably adequate amounts of zinc.
The recommended dietary allowance (RDA) for adult men and women is 15 mg/day.
Low protein diets are apt to have significantly less zinc than this. A higher
intake is recommended during pregnancy and lactation.
Since zinc deficiency in pregnant rats has caused congenital malformations,
concern has been expressed that congenital malformations in humans might be a
consequence of zinc deficiency. Malformations have only been obtained with diets
extremely low in zinc and so far only in rats. Whether the results with rats
would be obtained in other species is presently unknown.
Interrelationships with Other Components of the Diet
Phytic acid was one of the first constituents of plants to be linked to the
decreased availability of zinc from plant sources. Other factors in plants
affect zinc availability, but there is little information on them.
High calcium levels in the diet aggravate symptoms of zinc deficiency,
particularly in pigs; calcium does not have much effect, however, unless most
of the protein in the diet is derived from plant sources, particularly seeds.
Although interrelationships between zinc and vitamins such as biotin and
minerals such as cobalt have been suggested, no vitamin or mineral additions
have alleviated the symptoms of zinc deficiency. Additional histidine or
histamine has alleviated the swollen hock condition that develops in zinc-
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deficient chicks/ although other symptoms of zinc deficiency in chicks were not
affected.
Occurrence of Zinc Deficiency
Severe zinc deficiency caused by low zinc intake is rare. The incidence
of marginal deficiency states is largely unknown because of the lack of a good
method for assessing zinc status. Low hair zinc levels reported in some
American school children suggest that marginal zinc deficiencies do occur.
Apparent zinc deficiencies have also been reported in animals fed diets appear-
ing to be adequate in zinc.
Assessment of Zinc Deficiency
Because the chief effects of low zinc intake — poor appetite and slow
growth — may arise from many causes, a specific test is needed to diagnose zinc
deficiency. Plasma and hair zinc levels have been used most frequently to assess
the zinc status of humans and animals. Plasma zinc is limited in that it is
affected by many things other than zinc intake. Hair zinc reflects zinc status
only over fairly long times. Although both measures probably will be low in
cases of severe deficiency, they may not be lowered in states of marginal
deficiency.
Metabolic Lesions in Zinc Deficiency
Zinc deficiency results in decreased growth of most body tissues, although
the effect does not seem to interfere with DNA synthesis per se. Indeed,
mitotic activity increases in cells in the esophagus and buccal mucosa. Why
mitotic activity should increase in a few tissues and decrease in most others
is not known. Disturbances in sulfur metabolism, glucose metabolism, bone growth,
and reproduction have also been reported. However, the extent to which these
differences are caused by altered food intake or reduced growth of the deficient
animal rather than the lack of zinc per s£ is unclear.
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Zinc in Wound Healing in Animals
Although zinc has been implicated in more rapid healing of wounds,
studies with animals indicate that additional zinc is of value only if the
animal is zinc-deficient. There is no evidence that supplementation of zinc
for zinc-adequate animals improves wound healing.
ZINC IN METALLOPROTEINS
Since 1940, when Keilin and Mann discovered the first zinc metallo-
enzyme, carbonic anhydrase, the list of enzymes in which the functional or
structural role of Zn(II) has been documented has increased considerably.
Data on approximately 50 enzymes suggest that zinc may be involved as a
necessary cofactor. Several metabolically important reactions are catalyzed
by these enzymes, including hydrolysis, hydration, oxidation-reduction, and
group transfer reactions. Detailed physicochemical data on several of the
enzymes catalyzing hydrolysis or hydration reactions, e.g., carboxypeptidase
A and carbonic anhydrase, show the zinc to function in its capacity as a
Lewis acid by withdrawing electrons from a group of the substrate directly
coordinated to the metal ion at a site initially occupied by solvent water.
Zinc may also generate active, coordinated hydroxide ions at enzyme-active
sites. In each case, the function of the Zn(II) as a Lewis acid is clearly
only part of a concerted mechanism involving reactive side chains of the
proteins in addition to the metal ion. Whereas such interaction may be the
role of Zn(II) in the catalytic mechanism of a number of enzymes, the Zn(II)
in superoxide dismutase and aspartate transcarbamylase does not appear to
the
interact directly with/substrate. Thus zinc may also function by maintaining
the required conformation of a protein or by participating in the binding of
effector molecules to allosteric enzymes. If, in addition to its functional
role in we11-characterized zinc metalloenzymes, zinc is an absolute requirement
460
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for the function of the nucleotidyl transferase enzymes basic to DNA replica-
tion and transcription, then the metal's fundamental role in molecular
biology and at least some of the molecular reasons for the severe physiologic
effects of zinc deficiency become clear.
CLINICAL ASPECTS OF ZINC METABOLISM
Because zinc is a cofactor in protein synthesis, it plays an active role
in many disease processes.
Severe liver disease is commonly associated with loss of total body zinc.
In animals, zinc pretreatment has been associated with some protection from
liver damage with several hepatotoxins. However, the role of zinc therapy in
human liver disease is unclear. Gastrointestinal malabsorption from any cause
is associated with decreased gut absorption of zinc and with the subsequent
production of zinc deficiency. Although rare, acrodermatitis enteropathica is
a severe systemic disease in which zinc malabsorption plays a role in patho-
genesis. The symptoms of this disease disappear following exogenous zinc
administration.
Serum concentrations of zinc vary with several disease states, including
several cancers, blood dyscrasias, infectious processes, and renal diseases.
Various drugs alter zinc levels in blood and urine, as do peritoneal and blood
dialyses of patients with uremia. Parenteral hyperalimentation is associated with
decreased serum zinc concentration and increased urinary zinc excretion, a con-
dition consistent with the production of zinc depletion.
Zinc deficiency, as evidenced by total body loss of zinc, has been produced
acutely in man following the oral administration of L-histidine; the symptoms
produced by this body depletion of zinc are quickly obviated following exogenous
oral zinc administration, even in the face of continued histidine administration.
In animals, several studies have suggested that zinc may inhibit the growth
of specialized tumors. There are no applicable data for humans, but changes in
serum zinc concentrations accompany many malignant processes at various stages,
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Preliminary reports of the usefulness of zinc in the treatment of various
medical disorders have appeared but have yet to be substantiated by controlled
clinical trials. Claims have been made that schizophrenia, atherosclerotic
cardiovascular disease, laryngeal granulomas, gastric ulcers, and some forms of
color blindness can be improved by oral administration of zinc ion.
Zinc has been used as a therapeutic preparation for several centuries. It
has been applied topically to treat skin disorders since ancient times and is
still used for this purpose. Its capacity to produce gastrointestinal irritation
is consistent with its use as an emetic. Oral administration of zinc has been
used to assist in the more rapid healing of wounds of various types, but this
therapy appears to be useful only in patients with signs and symptoms of zinc
deficiency.
Zinc has also been used to treat patients with taste and smell
dysfunction of various types and, as in the wound healing studies, it is
apparently of little value except in those patients with zinc deficiency. Be-
cause zinc deficiency may be difficult to ascertain reliably, identification of
patients at risk poses a difficult practical problem.
TOXICITY OF ZINC
Humans
Zinc is not a highly toxic substance. Zinc toxicosis may occur only when
very high dose levels overwhelm the homeostatic mechanisms controlling zinc
uptake and excretion. Reports of zinc tolerance as well as toxicosis in humans
are sparse, but they do suggest that 500 mg to 1 g or more may be ingested on a
daily basis without adverse effects. Ten or more g taken as a single oral dose
may produce gastrointestinal distress, including nausea, vomiting, and
diarrhea. There are also suggestions in the literature that even higher dosage
may produce dizziness and perhaps increase blood levels of pancreatic enzymes.
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None of these observations of acute disorders have been rigorously studied
under controlled conditions nor are they confirmed experimentally. Chronic
zinc toxicosis in humans is even less well documented.
Inhalation of zinc has been related to metal fume fever, an acute
disability of short duration that can occur when fume is inhaled from metal
heated to a temperature above its melting point. It is most commonly associated
with inhalation of zinc oxide fume and is most severe among brass founders.
It is characteristized by hyperpnea, shivering accompanied by fever, profuse
sweating, pain in chest and legs, and general weakness beginning 4-8 h after
exposure and lasting about 24-48 h. With repeated exposures, some degree of
tolerance may be built up, but it will be lost when exposure to fume ceases for
a period as short as two days. The pathogenesis of this disorder, including
the role of zinc in it, is not understood.
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Animals
Although animals have a high tolerance for zinc, problems have been
reported in animals (particularly horses) grazing near lead-zinc smelters.
Zinc intake of the order of 3,000 ppm (mg/kg body weight) was required to
induce the symptoms experimentally. Most animals appear to tolerate levels
up to 1,000 ppm in the diet without ill effects if the diet contains adequate
copper and iron. High levels of zinc interfere with metabolism of these
minerals; therefore anemia and increased serum cholesterol are likely if
copper or iron intake is low.
Interactions Between Zinc and Cadmium
Zinc and cadmium have some physiochemical properties in common. There
are, however, large differences between these two metals in biologic sys-
tems: zinc is essential and has a short biologic half-time, and cadmium
is not essential and has an extremely long biologic half-time. Zinc is
found in high concentrations in most tissues, whereas cadmium mainly accu-
mulates in kidney and liver. The cadmium- and zinc-binding protein, metal-
lothionein, is the main storage protein for cadmium. Zinc easily crosses
the placental barrier, whereas cadmium is practically excluded. Experiments
with animals, in which cadmium concentrations in the diet often have been
similar to or higher than the zinc concentrations, have shown that cadmium
may accentuate symptoms brought on by zinc deficiency. Whereas zinc can
counteract some actions of cadmium (such as weight loss), it does not
influence anemias caused by cadmium.
Exposure to cadmium will increase zinc levels in kidney and liver.
When zinc intake is marginal, exposure to cadmium may cause some tissues,
such as testes, to become depleted of zinc. Cadmium has been shown to
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interfere with many enzymes both In vivo and jLn vitro. Zinc may prevent
some of these interferences, but not all.
Limited data exist on zinc-cadmium interactions in human beings. The
normal accumulation of cadmium in renal cortex will be accompanied by an
equimolar increase in zinc, probably reflecting the metal content of metal-
lothionein.
From the available data, it can be concluded that for human beings who
ingest enough zinc and who are not excessively exposed to cadmium, the main metal-
metal interaction will take place in the kidneys, where about one-third of
the total body burden of cadmium is stored. The cadmium concentrations in
other parts of the body are probably too low to interfere greatly with
zinc-dependent systems. As long as the amount of zinc necessary for normal
function is not altered in the kidneys, cadmium should not cause any functional
disturbances in that organ. Higher exposure to cadmium—from industry or
food—will elevate cadmium levels and eventually cause renal damage. The
few available data indicate that zinc stops increasing when cadmium levels
become very high in the renal cortex.
If zinc intake is marginal, then it can be postulated that zinc levels
of some tissues may become depressed if the accumulation of cadmium in kidney
and liver results in zinc storage there. A risk may be incurred for the
fetus if pregnant women are exposed to cadmium without adequate zinc intakes.
The danger lies not in a direct action of cadmium, but in the smaller amount
of zinc available for the fetus.
Usually animal products rich in protein are a good source for zinc, and
cadmium concentrations are very low in meat. If vegetable protein sources
replace meat, it can be expected that the zinc:cadmium ratios will not be so
favorable.
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The World Health Organization has recommended that the weekly intake of
cadmium in adults should not exceed 400-500 yg. This means a daily limit
of about 70 pg of cadmium. It has also recommended that the concentration
in drinking water should not exceed 5 yg cadmium/kg. If zinc supplements
are added to human food, care must be taken that excessive amounts of
cadmium are not added as well. Since not more than 10 yg (assuming 2 kg
water/day) should be allowed from drinking water, less should be allowed from
supplements. If zinc Is added to human diets, supplements should not
cause an increase of more than 5 yg cadmium/day. There is thus need for
strict control of zinc compounds used as food additives.
STANDARDS FOR ZINC LEVELS
Standards for zinc in air and water have been obtained using techniques
which do not fully reflect available advanced technology. Zinc is ubiquitous
and, as such, will be present in the environment forever. Continuous moni-
toring of the environment is important to maintain a close check on the
levels of zinc in air, water, and land.
SAMPLING AND MEASUREMENT TECHNIQUES FOR ANALYZING ZINC
Analysis of Samples from the General Environment
Methods for evaluating zinc In media such as physiologic fluids, soft
tissue, bone, hair, waters, soils, foodstuffs, and plants vary greatly in
nature as to merits and disadvantages. Techniques include absorption spectro-
photometry, atomic absorption spectrophotometry, neutron activation analysis,
anodic stripping voltammetry, X-ray fluorescence, arc-emission spectrography,
spectrofluorometry, and optical spectrography. Comparatively
reliable and standardized methods for zinc in waters, soils, food-
stuffs, and plants exist, but for biologic media, recommended routine methodology
exists only for urine and serum.
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Methods of expressing zinc levels in various media vary considerably.
They Include wet weight basis, dry weight basis, dry-ash weight basis, and
per-unit biochemical reference such as protein, DNA, etc.
Analysis of Zinc in the Air
Modern methods for determining zinc in the atmosphere include absorption
spectrophotometry, atomic absorption spectrometry, optical emission spectro-
graphy, X-ray fluorescence spectrometry, spark-source mass spectrometry,
instrumental neutron activation analysis, and voltammetry (polarography).
The inclusion of activation analysis indicates the ready availability of
nuclear reactors for carrying out routine analyses. Ring oven techniques have
not been discussed because the results obtained are semlquantitatlve.
The variety of analytic methods and sampling procedures that have been
used complicates comparing results of different Investigators necessary to
coordinate data to test theories of zinc sources, atmospheric transformations,
or removal mechanisms. Because the variations reported are a consequence of
differences in sample handling and Instrument operating procedures, results
are difficult if not Impossible to interpret. Comparisons of data from
different laboratories could lead to erroneous conclusions.
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CHAPTER 15
RECOMMENDATIONS
1. Areas of zinc deficiency and zinc excess in Soils and waters in the
United States should be identified.
2. A systematic programofmonitoring air for zinc should be established.
The sparseness of data on airborne zinc frustrates attempts to
quantify and identify sources of zinc in the environment. A study
designed to monitor airborne zinc levels both in the general community
and near known point sources would greatly assist in determining the
environmental impact of diverse zinc sources,
3. .Efforts^should bemade^to Controljzinc leveljs^ in zinc-containing was te-
waters to avoid excessive contamination of sludges that will beadded
to agricultural_J.ands.
Many industrial and sewage sludges contain high levels of zinc.
Since such sludges are sometimes used as fertilizers to provide nitro-
gen, phosphorus, and micronutrients, efforts should be made to ensure
that the addition of zinc to soil is not great enough to cause
toxicoses in plants and excessive levels in feed and food. Con-
taminated sludges should be disposed of in sanitary landfills or
incinerated; they should not be applied to land indiscriminately.
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4. Additional research should be conducted on the occurrence and characte£-
istics oforganic zinccomplexes in natural waters.
Chemical complexing of zinc may affect the biologic availability
of the metal to aquatic organisms. Naturally occurring physico-
chemical forms of zinc should be distinguished in
order to understand the mechanisms controlling zinc accumulation
and metabolism in aquatic organisms.
5. Research is needed on the relation of zinc to aquatic plants.
a. A broader range of zinc values and concentration factors are needed
for many aquatic plant species to evaluate the impact of increasing
doses and varied forms of zinc in ambient waters. Sampling and
analysis techniques need to be refined to establish critical or
threshold values for zinc toxicity and the overall zinc nutrition
status in aquatic plants.
b. The influence of biologic, chemical, and environmental factors in
sediments and waters upon zinc content and movement into ambient
waters, aquatic plants and ultimately the food chain should be
studied further.
c. The accumulated wealth of information on the functions of zinc in
aquatic plants, especially green algae, should be used as a basis
for more concentrated research on zinc in plant metabolism. Such
an approach would be the most useful for expanding understanding of
functions of zinc in plants.
6. Research isneeded on the relation of zinc to terrestrial plants.
a. The specific mechanisms of zinc uptake and translocation at the
molecular level need to be identified.
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b. Additional metabolic functions need to be characterized, because
the roles identified to date require only a small fraction of the
zinc needed for normal plant growth.
c. The physiologic basis for differential zinc requirements among
species needs to be defined.
d. The mechanism of zinc-phosphorus interactions in plants needs to
be determined.
e. The chemical forms of zinc in soils controlling zinc levels in soil
solution need to be determined. If this is done, soil solution levels
can be monitored for normal plant growth and nutrition and deficiency
and toxicosis can be more easily guarded against.
f. Specific mechanisms of zinc deficiencies, toxicoses and tolerance
in plants need to be identified.
g. The processes of zinc redistribution within plants, especially the
movement from vegetative tissue to seeds, should be determined.
h. The basis of foliar absorption and movement of zinc in plants needs
to be established.
7. Experiments on acute and chronic exposure of aquatic organisms to ele-
vated zinc concentrations in aqueous systems should be undertaken on a
systematic basis to determine the effects of zinc on vital life processes.
The effects of zinc should be determined alone and in combination
with other potential contaminants of human origin, such as
cadmium. Studies should be conducted for various critical life
stages under many combinations of important environmental interactions
(salinity, temperature, pH, dissolved oxygen) over all ranges of
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variation to which the test organism is exposed naturally. Such
a systematic approach is required to ensure adequate water quality
standards for protecting aquatic organisms.
8. Research into the physiologic role of zinc in humans should be encouraged.
Systems to be investigated should include the endocrine system,
sensory systems, including vision and taste, muscle and the nervous
system, including the central and peripheral nervous systems and
behavior.
9. The following additional research on zinc in the diet is needed:
a. A method of assessing zinc status of man and animals. There is no
currently accepted method of determining whether or not an animal is
receiving adequate zinc.
b. The occurrence of marginal zinc deficiencies. Because of the lack
of a method of determining zinc status, the prevalence of marginal
zinc deficiency is completely unknown.
c. The requirement for zinc and factors that affect the requirement.
The data available for determining the amount of zinc required,
particularly by man, are very limited. Since diets low in protein
will not generally contain the 15 mg zinc recommended daily for
adult humans, It is important to know how low an intake would be
adequate. It has been suggested that additional zinc may be bene-
ficial for both man and animals in times of stress, but more in-
formation is needed.
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d. Factors affecting the availability of zinc. Reports of zinc
deficiency in animals, particularly ruminants, fed diets con-
taining presumably adequate zinc indicate the need for additional
information on factors affecting the metal's availability. Almost
no information exists on constituents of plants, other than
phytate, that affect the availability of zinc.
e. Content of zinc in food. Attempts to calculate the dietary zinc
intake in man are hampered by the lack of adequate or accurate
information in the literature. Additional data could be obtained
readily with methods now available.
f. Zinc and cadmium. Zinc requirements should be studied in human
populations exposed to excessive cadmium. More studies should be
conducted on the effect of prolonged exposure to small amounts of
cadmium on zinc metabolism of animals on deficient, marginal, and
zinc-adequate diets. Zinc metabolism and its relation to cadmium
exposure in pregnancy especially should be studied.
10. The role, if any, of zinc in cancer should be investigated systematically.
This examination should include an evaluation of changes in zinc
metabolism that occur in various cancers and the role, if any, of
zinc as an inhibitor of oancer growth.
11. The role, if any, of zinc deficiency in human fetal wastage and con-
genital malformations should be investigated.
12. Efficacy of zinc therapy for various diseases should be evaluated
through controlled, randomized, cooperative studies.
Such trials could deal with the question of zinc treatment in wound
healing, anorexia, and taste and small dysfunction.
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13. Cadmium levels should be evaluated In preparations in zinc used
as foodstuffs.
14. Standard reference materials and standard analytic methods for
atmospheric participates should be developed.
Such measures would permit analysts to check their results.
The availability of standard materials and procedures would
provide a means not only to evaluate the reliability of
analytic methods, but also to compare data obtained in
different laboratories from different samples at different
times.
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APPENDIX A
Zinc Content of Foodg-
I.
Food and Description— Zinc, mg~
Apples, raw 0.05
Applesauce, unsweetened 0.1
Bananas, raw 0.2
Beans, common, mature, dry
Raw 2.8
Boiled, drained 1.0
Beans, lima, mature, dry
Raw 2.8
Boiled, drained 0.9
Beans, snap, green
Raw 0.4
Boiled, drained 0.3
Canned, solids and liquid 0.2
Canned, drained solids 0.3
Beef, separable lean
Raw 4.2
Cooked, dry heat 5.8
Cooked, moist heat 6.2
Beef, separable fat, raw 0.5
Beef, ground (77% lean)
Raw 3.4
Cooked 4.4
Beverages, carbonated, nonalcoholic
Bottled ^,0.01
Canned 0.08
Bran, see wheat
Breads
Rye 1.6
White . 6
Whole wheat 1.8
Food and Description— Zinc, mg—
Butter 0.1
Cabbage, common
Raw 0.4
Boiled, drained 0.4
Cake, white, without icing 0.2
Carrots
Raw 0.4
Cooked or canned, drained solids 0.3
Cheese, cheddar type 4.0
Chicken, broiler-fryer
Breast, meat only
Raw 0.7
Cooked, dry heat 0.9
Breast
Raw (81% meat, 12% skin, 7% fat) 0.7
Cooked, dry heat (89% meat,11% skin)0.9
Drumstick, thigh, back, meat only
Raw 1.8
Cooked, dry heat 2.8
Drumstick
Raw (85% meat, 13% skin, 2% fat) 1.7
Cooked, dry heat (84% meat,16% skin)2.5
Wing, meat only
Raw 1.6
Cooked, dry heat 2.4
Neck, meat only
Raw 2.7
Cooked, moist heat 3.0
Skin
Raw 1
Cooked, dry heat
0
1.2
-------
APPENDIX A - continued
Ul
I.
Food and Description" Zinc,
Chickpeas or garbanzos, mature seeds, dry
Raw 2.7
Boiled, drained 1.4
Chocolate syrup 0.9
Clams
Soft shell
Raw 1.5
Cooked 1.7
Hard shell
Raw 1.5
Cooked 1.7
Surf, canned, solids and liquid 1.2
Cocoa, dry powder 5*6
Coffee
Dry, instant 0 . 6
Fluid beverage 0.03
Cookies, vanilla wafers 0.3
Cooking oil, see oils
Corn, field, whole-grain, yellow, or 2.1
white
Corn, sweet, yellow
Raw 0.5
Boiled, drained 0.4
Corn, canned, whole kernel, yellow
Brine pack, solids and liquid 0.3
Brine pack, drained solids 0.4
Vacuum pack, solids and liquid 0.4
Corn chips 1.5
Corn grits, white, degermed, dry form 0.4
Corn flakes 0.3
Food and Description"
Cornmeal, white or yellow
Bolted (nearly whole grain)
Degermed
Dry form
Cooked
Cornstarch
Cowpeas (blackeyed), mature, dry
Raw
Boiled, drained
Crabs, blue and Dungeness
Raw
Steamed
Crackers
Graham
Saltines
Doughnuts, cake-type
Eggs, fresh
Whites
Yolks
Whole
Farina, regular
Dry form
Cooked
Fish, white varieties, flesh only
Raw
Cooked, fillet
Cooked, steak
Gizzard
Chicken
Raw
Cooked, drained
Zinc, mg~
1.8
0.8
0.1
0.03
2.9
1.2
4.0
4.3
1.1
.5
,5
0.02
3.0
1.0
0.5
0.06
0.7
1.0
0.8
2.9
4.3
-------
APPENDIX A - continued
Food and Description"
Gizzard
Turkey
Raw
Cooked, drained
Granola
Heart
Chicken
Raw
Cooked, drained
Turkey
Raw
Cooked, drained
Ice Cream
Lamb
Separable lean
Raw
Cooked, dry heat
Cooked, moist heat
Separable fat, raw
Lard
Lentils, mature, dry
Raw
Boiled, drained
Lettuce, head or leaf
Liver
Beef
Raw
Cooked
Calf
Raw
Cooked
Zinc, mg~
2.8
4.1
2.1
2.9
4.8
2.8
4.8
0.5
3.0
4.3
5.0
0.5
0.2
3.1
1.0
0.4
3.8
5.1
3.8
6.1
Food and Description"
Liver
Chicken
Raw
Cooked
Turkey
Raw
Cooked
Lobster, crayfish
Raw
Cooked or canned
Macaroni
Dry form
Cooked, tender stage
Margarine
Milk
Fluid, whole or skim
Canned, evaporated
Dry, nonfat
Oatmeal or rolled oats
Dry form
Cooked
Oat cereal, puffed, ready-to-eat
Oil, salad or cooking
Onions, mature or green, raw
Oranges, raw
Orange juice
Canned, unsweetened
Fresh or frozen
Oysters, raw or frozen
Atlantic
Pacific
Zinc, mg~
2.4
3.4
2.7
3.4
1.8
2.2
1.5
0.5
0.2
0.4
0.8
4.5
3.4
0.5
3.0
0.2
0.3
0.2
0.07
0.02
74.7
9.0
-------
APPENDIX A - continued
I.
Food and Description~ Zinc, mg~
Peaches
Raw 0.2
Canned, drained slices 0.1
Peanuts
Raw 2.9
Roasted 3.0
Peanut butter 2.9
Peas, green, Immature
Raw 0.9
Boiled, drained 0.7
Canned, drained solids 0.8
Peas, green, mature seeds, dry
Raw 3.2
Boiled, drained 1.1
Popcorn
Unpopped 3*9
Popped
Plain 4.1
Oil and salt added 3.0
Pork
Trimmed lean cuts, separable lean
Raw 2.7
Cooked 3.8
Boston butt, separable lean
Raw 3.2
Cooked 4.5
Ham or picnic, separable lean
Raw 2.8
Cooked 4.0
Food and Description"" Zinc, mg~
Pork
Loin, separable lean
Raw 2.2
Cooked 3.1
Separable fat, raw 0.5
Potatoes
Raw 0.3
Boiled, drained 0.3
Rice
Brown
Dry form 1.8
Cooked 0.6
White, regular
Dry form 1.3
Cooked 0.4
White, parboiled
Dry form 1.1
Cooked 0.3
White, precooked, quick
Dry form 0.7
Cooked 0.2
Cereal, ready-to-eat, puffed or flakes 1.4
Rolls, hamburger 0.6
Salad dressing 0.2
Salmon, canned (77% solids, 23% liquid) 0.9
Sausages and cold cuts
Bologna, beef 1.8
Braunschweiger 2.8
Frankfurters
Made with beef 2.0
Made with beef and pork 1.6
-------
APPENDIX A - continued
1^
b c
Food and Description" Zinc, mg~
Shrimp
Raw 1.5
Boiled, peeled, deveined 2.1
Canned, drained solids 2.1
Spinach
Raw 0.8
Boiled, drained 0.7
Canned
Solids and liquid 0.6
Drained solids 0.8
Sugar, white, granulated 0.06
*j Tea
oo Dry leaves 3.3
Fluid beverage 0.02
Tomatoes, ripe
Raw 0.2
Boiled, solids and liquid 0.2
Canned, solids and liquid 0.2
Tunafish, canned in oil (85% solids, 1.0
15% oil)
Drained solids 1.1
Turkey
Light meat
Raw 1.6
Cooked, dry heat 2.1
Dark meat
Raw 3.1
Cooked, dry heat 4.4
Neck meat
Raw 5.0
Cooked 6.4
Food and Description"
Turkey
Skin
Raw
Cooked
Veal
Separable lean
Raw
Cooked, dry heat
Cooked, moist heat
Separable fat, raw
Wheat, whole grain
Hard
Soft
White
Durum
Wheat flours
Whole
80% extraction
All-purpose
Bread flour
Cake or pastry flour
Wheat bran, crude
Wheat germ, crude
Wheat cereal, whole-meal
Dry form
Cooked
Zinc, mg~
1.3
2.1
2.8
4.1
4.2
0.5
3.4
2.7
2.2
2.7
2.4
1.5
0.7
0.8
0.3
9.8
14.3
3.6
0.5
-------
vo
APPENDIX A - continued
b c
Food and Description"" Zinc, mg~
.*
Wheat cereals, ready-to-eat
Bran flakes, 40% 3.6
< Flakes 2.3
Germ, toasted 15.4
Puffed 2.6
Shredded 2.8
1093a
a.
-iteproduced with permission from Murphy et^ al.
—100 g, edible portion.
•^Data are given to two decimal places if food contains less than 0.1 mg zinc per edible portion.
-------
APPENDIX A
Zinc Content of Foods"
II.
Food
Apples, raw
Applesauce, unsweetened
Bananas, raw
Beans, common, mature dry
Raw
Bo tied, drained
Beans, lima, mature, dry
Raw
Boiled, drained
Beans, snap, green
Raw, cut into 1-2 in* lengths
Boiled, drained, cut and
French style
Canned, solids and liquid
Canned, drained solids
Beef, separabe lean
Cooked, dry heat
Cooked, moist heat
Beef, ground, cooked
Beverages, carbonated, non-
alcoholic
12 fl oz (360 ml)
12 f1 oz
Breads
Rye
White
Whole wheat
Butter, 4 sticks/lb
Cabbage, common
Raw, shredded finely
Shredded, boiled, drained
Cake, white, without icing
(3 x 3 x 2 inj
Carrots
Raw
Cooked or canned, drained
solids
Cheese, cheddar
Chicken, broiler-fryer, cooked,
dry heat
Breast, cooked
Meat only
Meat and skin
Approximate
Measure
1 medium
1 cup
1 medium
1 cup
1 cup
1 cup
1 cup
1 cup
1 cup
1 cup
1 cup
3 oz
3 oz
3 oz
1 bottle
1 can
1 slice
1 slice
1 slice
1 cup
1 tbsp
1 cup
1 cup
1 piece
1 medium
1 cup
1 slice
1/2 breast
1/2 breast
Weight,
180
244
119
190
185
180
190
110
125
239
135
85
85
85
367
367
25
28
28
227
14
90
145
86
72
155
13
85
96
Zinc, mg~
0.08
0.3
0.3
5.3
1.8
5.0
1.7
0.4
0.4
0.6
0.4
4.9
5.3
3.8
0.01
0.3
0.4
0.2
0.5
0.2
0.01
0.3
0.6
0.2
0.3
0.5
0.5
0.7
0.9
480
-------
APPENDIX A - continued
II.
Food
Chicken, broiler-fryer, cooked,
dry heat
Drumstick, thigh, back, meat
only, cooked
Drumstick
Meat only
Meat and skin
Chickpeas, mature, dry
Raw
Boiled, drained
Chocolate syrup, 1 fl oz
Clams
Soft shell, cooked
Hard shell
Raw
Cooked
Surf, canned, solids and
liquids, can size
211 x 300
Cocoa, dry powder
Approx 5 1/4 tbsp
Coffee
Dry, instant
Fluid beverage, 6 fl oz
Cookies (1 3/8 x 1/4 inO
Corn, sweet, yellow
Boiled, drained
Canned, vacuum pack
Corn chips
Corn grits, dry form
Corn flakes
Cornmeal, white or yellow
Bolted, dry form
Degermed
Dry form
Cooked
Cowpeas (blackeyes)
Raw
Boiled, drained
Crabs, steamed, pieces
Crackers
Graham (2 1/2 x 2 1/2
Saltines
Doughnuts (3 1/4 in. diam)
Approximate
Measure
3 oz
1 drumstick
1 drumstick
1 cup
1 cup
2 tbsp
3 oz
Weight, g~
85
45
54
200
146
38
85
4 cherrystones or 70
5 littlenecks
4 or 5 clams 62
1 can 220
1 oz
1 tbsp
1 cup
10 cookies
1 cup
1 cup
1 oz
1 cup
1 oz
1 cup
1
1
1
1
1
cup
cup
cup
cup
cup
2 squares
10 crackers
1 doughnut
28
2.5
180
30
165
210
28
160
28
122
138
240
170
250
155
14
28
42
Zinc, mg
2.4
1.3
1.4
5.4
2.0
0.3
1.4
1.1
1.0
2.7
1.6
0.02
0.05
0.08
0.7
0.8
0.4
0.7
0.08
2.1
1.2
0.3
4.9
3.0
6.7
0.2
0.1
0.2
481
-------
APPENDIX A - continued
II-
Food
Approximate
Measure
Eggs, fresh
White
Yolk
Whole
Farina, regular
Dry form
Cooked
Fish, white varieties, fresh only
Fillet, cooked
Steak, cooked
Gizzard, cooked, drained, diced
Chicken
Turkey
Granola
Heart, cooked, drained, diced
Chicken
Turkey
Ice Cream
Lamb, separable lean
Cooked, dry heat
Cooked, moist heat
Lard
Lentils, mature, dry
Raw
Boiled, drained
Lettuce, head or leaf
Approx 1/6 head
Loose leaf, chopped
Liver, cooked
Beef
Calf
Chicken, chopped
Turkey, chopped
Lobster, cooked, cubed
Macaroni, cooked, tender
Measured hot
Measured cold
Margarine
Milk
Fluid
Canned, evaporated
Dry, nonfat
1 large
1 large
1 large
1 cup
1 cup
3 oz
3 oz
cup
cup
oz
cup
cup
cup
3 oz
3 oz
1 cup
1 tbsp
1
1
1
1
cup
cup
wedge
cup
2 oz
2 oz
cup
cup
cup
1
1
1
1
1
1
1
cup
cup
cup
tbsp
cup
cup
cup
Weight, g-
33
17
50
180
245
85
85
145
145
28
145
145
133
85
85
205
13
190
200
90
55
57
57
140
140
145
140
105
227
14
244
252
68
Zinc, mg~
<0.01
0.5
0.5
1.0
0.2
0.9
0.7
6.2
6.0
0.6
6.9
7.0
0.6
3.7
4.2
0.4
0.03
5.9
2.0
0.4
0.2
2.9
3.5
4.7
4.7
3.1
0.7
0.5
0.5
0.03
0.9
1.9
3.1
482
-------
APPENDIX A - continued
II.
Food
Oatmeal or rolled oats
Dry form
Cooked
Oat cereal, puffed
Oil, salad or cooking
Onions
Mature, chopped
Young green, chopped
Oranges, raw, 2 5/8 in. diam
Orange juice
Canned, unsweetened
Fresh or frozen
Oysters
Atlantic
Raw, drained, 12 fl oz can,
18-27 select or 27-44
standard oysters
Frozen, solids and liquid,
12 fl oz can
Pacific
Raw, drained, 12 fl oz can,
6-9 medium or 9-13 small
oyster
Frozen, solids and liquid,
12 fl oz can
Peaches
Raw, peeled, 2 1/2 in.diam
Canned, drained, slices
Peanuts, roasted
Peanut butter
Peas, green, immature
Raw or frozen
Boiled, drained
Canned, drained solids
Peas, green, mature seeds, dry
Raw
Boiled, drained
Popcorn
Unpopped
Popped
Plain, large kernel
With oil and salt
Approximate
Measure
1 cup
1 oz
1 cup
1 oz
1 cup
1 cup
1 cup
1 orange
1 cup
1 cup
1 can
1 can
1 can
1 can
1 medium
1 cup
1 tbsp
1 tbsp
Weight, g—
80
28
240
28
218
170
100
131
249
248
1
1
1
1
1
cup
cup
cup
cup
cup
1 cup
1 cup
1 cup
360
100
220
9
16
145
160
170
200
200
205
6
9
Zinc, mg—
2.7
1.0
1.2
0.8
0.4
0.6
0.3
0.2
0.2
0.05
340
360
340
254.3
268.9
30.6
32.4
0.2
0.3
0.3
0.5
1.2
1.2
1.3
6.4
2.1
7.9
0.2
0.3
483
-------
APPENDIX A - continued
II.
Food
Approximate
Measure
Pork, cooked, dry heat, separable
lean
Trimmed lean cuts 3 oz
Boston butt 3 oz
Ham or picnic 3 oz
Loin 3 oz
Potatoes
Raw, peeled, 21/2 in. diam 1 medium
Pared before cooking, boiled, 1 medium
drained
Boiled in skin, drained, 1 medium
pared
Rice
Brown
Dry form 1 cup
Cooked, measured hot 1 cup
White, regular, long-grain
Dry form 1 cup
Cooked
Measured hot 1 cup
Measured cold 1 cup
White, parboiled
Dry form 1 cup
Cooked
Measured hot 1 cup
Measured cold 1 cup
White, precooked, quick
Dry form 1 cup
Cooked
Measured hot 1 cup
Measured cold 1 cup
Cereal, ready-to-eat, puffed/flaked 1 oz
Rolis, hamburger, 3 1/2 in. diam 1 roll
Salad dressing 1 tbsp
Salmon, canned, solids and 1 cup
liquid
Sausages and cold cuts
Bologna, beef, 4 1/2 in.diam, 1 slice
1 oz
Braunschweiger, 1 oz 1 slice
Frankfurters
Made with beef, 10 per Ib 1 frank
Made with beef and pork, 1 frank
10 per Ib
Weight, g-
85
85
85
85
112
112
136
185
195
185
205
145
185
175
145
95
165
130
28
40
15
220
28
28
45
45
Zinc, mg—
3.2
3.8
3.4
2.6
0.4
0.3
0.4
3.4
1.2
2.5
0.8
0.6
2.1
0.6
0.5
0.7
0.4
0.3
0.4
0.2
0.03
2.1
0.5
0.8
0.9
0.7
484
-------
APPENDIX A - continued
II.
Food
Shrimp
Boiled, peeled, deveined,
33 per Ib
Canned, drained, solids
Spinach
Raw, chopped
Boiled, drained
Canned
Solids and liquid
Drained solids
Sugar, white, granulated
Tea, fluid beverage, 6 fl oz
Tomatoes , ripe
Raw, 2.6 in.diam
Boiled
Canned, solids and liquid
Tunafish, canned in oil
Chunk style, solids and
liquid, can size 307 x 113,
6 1/2 oz
Drained solids, can size
307 x 113, 6 1/2 oz
Drained solids
Turkey, cooked, dry heat, meat
only
Light meat
Dark meat
Veal, separable lean
Cooked, dry heat
Cooked, moist heat
Wheat flours
Whole, stirred, spooned into
cup
All purpose, sifted, spooned
Approximate
Measure
6 shrimp
1 cup
1 cup
1 cup
1 cup
1 cup
1 cup
1 cup
1 medium
1 cup
1 cup
1 can
1 can
1 cup
3 oz
3 oz
3 oz
3 oz
1 cup
1 cup
Weight, {£
84
128
55
180
232
205
200
177
123
241
241
184
157
160
85
85
85
85
120
115
Zinc, mg^-
1.7
2.7
0.5
1.3
1.5
1.6
0.1
0.04
0.2
0.5
0.5
1.7
1.8
1.8
1.8
3.7
3.5
3.6
2.9
0.8
into cup, standard granulation
Bread flour, sifted, spooned
1 cup
115
0.9
into cup, standard granulation
Cake flour, sifted, spooned
into cup
Wheat cereal, wholemeal
Dry form
Dry form
Cooked
Cooked, from 1 oz dry
1 cup
1 cup
1 oz
1 cup
96
125
28
245
216
0,3
4.5
1.0
1.2
1.0
485
-------
APPENDIX A - continued
II.
Approximate ,
Food Measure Weight, g— Zinc, mg—
Wheat cereals, ready-to-eat
Bran flakes, 40% 1 oz 28 1.0
Flakes 1 oz 28 0.6
Germ, toasted 1 tbsp 6 0.9
Puffed 1 oz 28 0.7
Shredded 1 oz 28 0.8
1093a
a
""Reproduced with permission of Murphy et al.
—Edible part of common household units; measure and weight only apply to
edible part of food.
c
•n)ata are given to two decimal places if food contains less than 0.1 mg
zinc per edible portion.
486
-------
APPENDIX B
Methods of Zinc Analysis*
Zinc is an essential and beneficial ele-
ment in body growth. However, con-
centrations above 5 mg/1 can cause a
bitter astringent taste and an opales-
cence in alkaline waters. The zinc con-
centration of U.S. drinking waters
varies between 0.06 and 7.0 mg/1, with
a mean of 1.33 mg/1. Zinc most com-
monly enters the domestic water supply
from the deterioration of galvanized iron
and the dezincification of brass. In such
cases the presence of lead and cadmium
also may be suspected, because they are
impurities of the zinc used in galvaniz-
ing. Zinc also may result from industrial
waste pollution.
1. Selection of Method
Where the equipment is available,
the atomic absorption spectrophotomct-
ZINC
ric meth'od is preferred for the determi-
nation of zinc. The dithizone and zincon
colorimetric methods are useful in the
absence of the sophisticated in-
strumentation. Dithizone method I is
intended for unpolluted waters, and II
for polluted waters or wastewatcr.
2. Sampling and Storage
Analyze samples within 6 hr after
collection. The addition of HCl will pre-
serve the metallic ion content but re-
quires that: (a) the acid be zinc-free; (b)
the sample bottles be rinsed with acid
before use; and (c) the samples be evap-
orated to dryness in silica dishes to re-
move the excess HCl before analysis.
^Copyright 1976 by the American Public Health Association.
Reproduced with permission.
1311a
487
-------
Atomic Absorption Spectrophotometric
Method
1. General Discussion
a. Principle: Atomic absorption
spectrophotometry resembles emission
flame photometry in that a sample is as-
pirated into a flame and atomized. The
major difference is that flame photome-
try measures the amount of light emit-
ted, whereas in atomic absorption
spectrophotometry a light beam is di-
rected through the flame, into a mono-
chromator, and onto a detector that
measures the amount of light absorbed
by the atomized element in the flame.
For many metals difficult to analyze by
flame emission, atomic absorption ex-
hibits superior sensitivity. Because each
metal has its own characteristic absorp-
tion wavelength, a source lamp com-
posed of that element is used; this makes
the method relatively free from spectral
or radiation interferences. Thus the
amount of energy of the characteristic
wavelength absorbed in the flame is pro-
portional to the concentration of the ele-
ment in the sample.
b. Interference: Most metals can be
determined by direct aspiration of the
sample into an air-acetylene flame. The
most troublesome type of interference is
termed "chemical" and results from the
lack of absorption of atoms bound in
molecular combination in the flame.
This can occur when the flame is not hot
enough to dissociate the molecules (in
the case of phosphate interference with
magnesium) or when the dissociated
atom is oxidized immediately to a com-
pound that will not dissociate further at
the temperature of the flame. The inter-
ference of phosphate in the magnesium
determination can be overcome by the
addition of lanthanum. Similarly, the
introduction of calcium eliminates silica
interference in the determination of
manganese. However, silicon and met-
als such as aluminum, barium, beryl-
lium, and vanadium require the use of
the higher-temperature, nitrous oxide-
acetylene flame to dissociate their mole-
cules. In addition, barium undergoes
ionization in the flame and the ground
state (potentially absorbing) population
is thereby reduced. The addition of an
excess of a cation (sodium or potassium)
having a similar or lower ionization po-
tential will overcome this problem. The
wavelength of maximum absorption for
arsenic is 193.7 nm, and for selenium
196.0 nm. Unfortunately, the air-acety-
lene flame absorbs intensely at these
wavelengths. The sensitivity of the
method for these metals can be im-
proved by the use of the argon-hydrogen
flame, in the determination of mercury
by the cold vapor (flameless) technic,
certain volatile organic materials may
absorb at 253.7 nm. If this is expected,
the sample should be analyzed by the
regular procedure and again under oxi-
dizing conditions only, that is, without
the addition of stannous chloride. The
true mercury concentration can be ob-
tained by subtracting the two values.
c. Sample handling: Before collecting
a sample, decide on the type of'data de-
sired, i.e., dissolved, suspended, total, or
extractable metals. This decision will de-
termine whether the sample is to be
acidified, with or without filtration, and
the kind of digestion required.
Acidify all samples at the time of col-
lection to keep the metals in solution and
to minimize their adsorption on the con-
tainer wall. If only dissolved metals are
to be measured, filter the sample
through a 0.45-/xm membrane before
acidification. If possible, filter and acid-
ify in the field at the time of collection.
Report the results obtained on this
sample as "dissolved." Filtration is not
necessary when total or extractable con-
centrations are required.
Acidify the sample with cone HNOa
to a pH of 2.0 or less. Usually, 1.5 ml
cone HNOa/l sample will be sufficient
for potable waters free from paniculate
matter. Such samples can be analyzed
with no further treatment. However,
samples containing suspended materials
or organic matter require pretreatment,
as described below.
488
-------
2. Apparatus
a. Atomic absorption spectropbo-
tmneter, consisting of a source of light
emitting the line spectrum of an element
(hollow cathode lamp), a device for va-
porizing the sample (usually a flame), a
means of isolating an absorption line
(monochromatoror filter and adjustable
slit), and a photoelectric detector with its
associated amplifying and electronic
"measuring equipment. Both direct cur-
rent and alternating current systems are
used in atomic absorption instruments.
The AC or chopped-beam system is
preferred because with this system flame
emission can be distinguished from lamp
emission. For waters high in salt, the use
of either a deuterium background cor-
rector or a double-beam instrument that
permits the measurement of the absorp-
tion at two different wavelengths simul-
taneously may be helpful.
b. Burner: The most common type of
burner is known as a premix, which in-
troduces the spray into a condensing
chamber for removal of large droplets.
The burner may be fitted with a con-
ventional head containing a single slot
7.6 cm (3 in.) long, which is most useful
for aspiration when organic solvents are
used; a three-slot Boling head, which is
preferred for direct aspiration with an
air-acetylene flame; or a head contain-
ing a single slot 5 cm (2 in.) long for use
with nitrous oxide and acetylene.
c. Recorder: While most instruments
are equipped with either a digital or
null meter readout mechanism, a good-
quality 10-mV recorder with high sensi-
tivity and a fast response time is needed
to record the peaks resulting from the
determination of mercury by the rold
vapor (flameless) technic and for the de-
termination of arsenic and selenium by
aspiration of their gaseous hydrides.
d. Hollow cathode lamps: Use one for
each element being measured. Multi-
element lamps are available but not rec-
ommended because they may require
the selection of different operating pa-
rameters.
e. Pressure-reducing valves: Main-
tain the supplies of fuel and oxidant at
pressures somewhat higher than the
controlled operating pressure of the in-
strument by suitable reducing valves.
Use separate reducing valves for each
gas.
/ Vent. Place a vent about 15 to JO
cm (6 to 12 in.) above the burner to re-
move the fumes and vapors from the
flame. This precaution protects the labo-
ratory personnel from toxic vapors, pre-
vents the stability of the flame from
being affected by room drafts, and pro-
tects the instrument from corrosive va-
pors. A damper or variable-speed
blower is also desirable for modulating
the air flow and preventing disturbance
of the flame.
3. Extractable Metals Analyses
"Extractable metals" include metals
in solution plus metals lightly adsorbed
on the suspended material. The results
obtained in analyses for ex tractable met-
als will be influenced by the kind of acid
or acids used in the digestion, the con-
centration of acid, and the heating time.
Unless conditions are controlled rigidly,
results will be meaningless and unre-
producible. The following procedure de-
termines metals soluble in hot HC1-
HNOs. At the time of collection, acidify
the entire sample with 5 ml cone
HNOa/1 sample. At the time of analy-
sis, mix the sample, transfer a 100-ml
portion to a beaker or flask, and add 5
ml 1 +1 redistilled HC1. Heat 15 min
on a steam bath. Filter and adjust the
volume to 100 ml. The sample is then
ready for analysis.
The data approximate the total met-
als in the sample, although something
less than the actual total is measured.
Concentrations of metal found, espe-
cially in heavily silted samples, will be
substantially higher than results ob-
tained on only the soluble fraction. Re-
port as "extractable" metals.
489
-------
Determination of Zinc by
Direct Aspiration into an
Air-Acetylene Flame
1. Apparatus
See abo**a for a description
of the required atomic absorption spec-
trophotometcr and associated equip-
ment. The three-slot Boling burner
head is recommended.
2. Reagents
a. Air, cleaned and dried through a
suitable filter to remove oil, water, and
other foreign substances. The source
may be a compressor or commercially
bottled gas.
b. Acetylene, standard commercial
grade. Acetone, which is always present
in acetylene cylinders, can be prevented
from entering and damaging the burner
head by replacing a cylinder when its
pressure has fallen to 7 kg/cm2 (100
psig) acetylene.
c. Calcium solution: Dissolve 6 JO mg
calcium carbonate, CaCCb, in 10 ml
cone HC1. Add 200 ml water, and if
necessary heat the solution and boil
gently to obtain complete solution. Cool
and dilute to 1,000 ml with deionized
distilled water.
d. Deionized distilled water: Use
deionized distilled water for the prepa-
ration of ail reagents and calibration
standards and as dilution water.
e. Hydrochloric acid, HC1, cone.
/ Lanthanum solution: Dissolve
58.65 g lanthanum oxide, LaiCb, in
250 ml cone HC1. Add the acid slowly
until the material is dissolved and dilute
to 1,000 ml with deionized distilled wa-
ter.
g. Nitric acid, HNCh, cone.
h. Standard metal solutions: Prepare
a series of standard metal solutions con-
taining 5 to 1,000 /zg/1 by appropriate
dilution of the following stock metal so-
lutions with deionized distilled water
containing 1.5 ml cone HNCh/1.
12) Zinc: Dissolve 1.000 g zinc metal
in 20 ml 1 + 1 HC1 and dilute to 1,000
ml with deionized distilled water; 1.00
ml=1.00mgZn.
3. Procedure .
a. Instrument operation-. Because of
differences between makes and models of
satisfactory atomic absorption spectro-
photometers, it is not possible to formu-
late instructions applicable to every in-
strument. In general, proceed according
to the following steps:
1) Install a hollow cathode lamp of
the desired metal in the instrument, set
the wavelength dial according to Table
301 :II, and align the lamp in accord-
ance with the manufacturer's instruc-
tions.
2) Set the slit width according to the
manufacturer's suggested setting for the
element being measured.
3) Turn on the instrument and apply
the amount of current suggested by the
manufacturer to the hollow cathode
lamp.
4) Allow the instrument to warm up
until the energy source stabilizes; this
process usually requires 10 to 20 min.
Readjust the current as necessary after
warmup.
5) Install the burner heads.
6) Turn on the air and adjust the
flow rate to that specified by the manu-
facturer to give maximum sensitivity for
the metal being measured.
7) Turn on the acetylene, adjust the
flow rate to the value specified, and ig-
nite the flame.
8) Atomize deionized distilled water
acidified with 1.5 ml cone HNOa/l, and
check the aspiration rate over 1 min.
Adjust if necessary to a rate between 3
and 5 ml/min, and zero the instrument.
490
-------
9) Atomize a standard (usually a 0.5-
mg/l standard is suitable) and adjust
the burner both up and down and side-
ways until a maximum response is ob-
tained.
10) The instrument is now ready to
operate. When analyses are finished, ex-
tinguish the flame by turning off first the
acetylene and then the air.
b. Standardization:
I) Select at least three concentrations
of each of the standard metal solutions
(prepared as in 2b above) so as to
bracket the expected metal concentra-
tion of a sample. Aspirate each in turn
into the flame and record the absorb-
ance.
2) For calcium and magnesium cali-
bration, mix 100 ml of standard with 25
ml of lanthanum solution (see 2/above)
before aspirating.
3) For iron and manganese calibra-
tion mix 100 ml of standard with 25 ml
of calcium solution (see 2c above) before
aspirating.
4) With some instruments, it may be
necessary to convert percent absorption
to absorbance by use of a suitable table
generally provided by the manufacturer.
5) Prepare a calibration curve by
plotting on linear graph paper the ab-
sorbance of the standards versus their
concentration.
6) Plot calibration curves for iron and
manganese based on the original con-
centrations of the standards before dilu-
tion with calcium solution (H2c).
7) Plot calibration curves for calcium
and magnesium based on the original
concentration of the standards before di-
lution with lanthanum solution (1f2/).
8) Recheck the calibration curve by
aspirating at least one standard after the
completion of the analysis of a group of
unknown samples.
c. A na lysis of samples :
1) Rinse the atomizer by aspirating
deionizcd distilled water containing 1.5
ml cone HNOa/1, and zero the in-
strument.
2) Atomize the sample and determine
its absorbance.
3) When determining calcium or
magnesium, dilute and mix 100 ml
sample with 25 ml lanthanum solution
fl|2/> before atomization.
4) When determining iron or manga-
nese, dilute and mix 100 ml sample
with 25 ml calcium solution (1|2f) be-
fore atomization.
4. Calculations
Calculate the concentration of each
metal ion, in micrograms per liter, by
referring to the appropriate calibration
curve prepared according to 1b 5), 6),
and 7).
491
-------
Dithizone Method I
1. General Discussion
D • • , XI . ,- ,
a. Principle-. Nearly 20 metals are ca-
, , , r . . { ,. , ...
pable of reactmg w.th diphenylthiocar-
bazone (dith.zone) to produce colored
coord,nat,on compounds. These dithi-
zonates are attractable into orgamcsol-
vents such as carbon tetrachlor.de. Most
interferences in the zinc-dithizone reac-
tion can be overcome by adjusting the
solution to pH 4.0 to 5.5 and by the ad-
dition of sufficient sodium thiosulfate.
Zinc also forms a weak thiosulfate com-
plex that tends to retard the slow and in-
complete reaction between zinc and
dithizone. For this reason, the determi-
nation is empirical and demands the use
of an identical technic in standard and
sample analysis. The duration and vigor
of shaking, the volumes of sample, so-
dium thiosulfate, and dithizone, and the
pH should all be kept constant.
b. Interference: Interference from bis-
muth, cadmium, cobalt, copper, gold,
lead, mercury, nickel, palladium, silver,
and stannous tin in the small quantities
found in potable waters is eliminated by
complexing with sodium thiosulfate and
by pH adjustment. Ferric iron, residual
chlorine, and other oxidizing agents con-
vert dithizone t Filter photometer, providing a
hght Path of 2 cm and equipped with ei-
ther a Feen filter hav'ng maximum
transmittance near 535 nm or a red fil-
ter having maximum transmittance
near °2" nm-
3> Nessler tubes, matched.
b- Separatory funnels, capacity 1 2 5
to J *° ml- Squibb form' preferably with
mert teflon stopcocks. If the funnels are
of identical size and shape, visual color
comparisons may be made directly in
them.
c- Glassware.- Rinse all glassware
with J + 1 HN°3 and zinc-free water.
d. pH meter.
„ R .
' "
a. Zinc-free water: Use redistilled or
deionized distilled water for rinsing ap-
492
-------
paratus and the preparation of solutions
and dilutions.
b. Stock zinc solution: Dissolve
100.0 mg 30-mesh zinc meral in a slight
excess of 1 + 1 HC1; about 1 ml is re-
quired. Dilute to 1,000 ml with zinc-
free water; 1.00 ml= I00 pg Zn.
c. Standard zinc solution: Dilute
10.00 ml zinc stock solution to 1,000 ml
with zinc-free water; 1.00 ml= 1.00 fig
Zn.
d. Hydrochloric add, HC1, 0.02M-
Dilute 1.0 ml cone HCI to 600 ml with
zinc-free water. If high blanks are traced
to this reagent, dilute cone HCI with an
equal volume of distilled water and re-
distill in an all-pyrex still.
e. Sodium acetate, 2N: Dissolve 68
gNaC2HjO2«3H2O and dilute to 250 ml
with zinc-free water.
/ Acetic acid, 1 + 7. Use zinc-free wa-
ter.
g. Acetate buffer solution: Mix equal
volumes of 2N sodium acetate solution
and 1+7 acetic acid solution. Extract
with 10-ml portions of dithizone solu-
tion I until the last extract remains
green; then extract with carbon tet-
rachloride to remove excess dithizone.
b. Sodium thiosulfate solution: Dis-
solve 25 g NazSzOj-JFhO in 100 ml
zinc-free water. Purify by dithizone ex-
traction as in If 3g above.
i. Dithizone solution I: Dissolve 100
mg diphenylthiocarbazone* in 1 1 CCU.
Store in a brown glass-stoppered bottle
in a refrigerator. If the solution is of
doubtful quality or has been stored for a
long time, test for deterioration as fol-
lows: Shake 10 ml with 10 ml 1+99
NH4OH. If the lower, CCU, layer is
* Eastman No. 3092 or equivalent.
only slightly yellow, the reagent is in
good condition.
/ Ditbizone solution II: Dilute I
volume of dithizone solution I with 9
volumes of CCI*. If stored in a brown
glass-stoppered bottle in a refrigerator,
this solution is good for several weeks.
k. Carbon tetrachloride, CCI<. CAU-
TION: Carbon tetrachloride is a toxic
substance. Long-continued absorption of
small amounts may be hazardous.
While the solvent can be absorbed
through the skin, the primary danger is
through inhalation of the vapor. Pre-
pare reagents and extract standards and
samples with carbon tetrachioride in a
well-ventilated hood.
/. Sodium citrate solution: Dissolve
10 g NajC6HsO7-2H2O in 90 ml zinc-
free water. Purify by dithizone extrac-
tion as in TI 3g preceding. Use this rea-
gent in the final cleansing of glassware.
4. Procedure
*
a. Preparation of colorimetric stand-
ards: To a series of 125-ml Squibb sep-
aratory funnels, thoroughly cleansed as
described in U 2c above, add 0, 1.00,
2.00, 3.00, 4.00, and 5.00 ml standard
zinc solution equivalent, respectively, to
provide 0, 1.00, 2.00, 3.00, 4.00, and
5.00 /ig Zn. Bring each volume up to
10.0 ml by adding zinc-free water. To
each funnel add 5.0 ml acetate buffer
and 1.0 ml sodium thiosulfate solution,
and mix. The pH should be between 4
and 5.5 at this point. To each funnel
add 10.0 ml dithizone solution II, stop-
per, and shake vigorously for 4.0 min.
Let the layers separate, dry the stem of
the funnel with strips of filter paper, and
run the lower (CCW layer into a clean
dry absorption cell.
tirade names have been identified solely to help readers
and do not imply any endorsement or recommendation by
the National Academy of Sciences or the National Research
Council.
493
-------
b. Photometric measurement: Meas- samples and standards at the same time.
ure either the red color of the zinc dithi- Compare the CCU layers directly in the
zonate at 535 nm, or the green color of separatory funnels if these match in size
the unreacted dithizone at 620 nm. and shape; otherwise transfer to
Set the photometer at 100% trans- matched test tubes or nessler tubes. The
mittance with the blank if the 535-nm range of colors obtained with various
wavelength is selected. If 620 nm is amounts of zinc are roughly these:
used, set the blank at 10.0% trans-
mittance. Plot a calibration curve. Run z'nc Q>|or
a new calibration curve with each set of {#
samples. 0 (blank) green
c. Treatment of samples: If the zinc ' bluc
content is not within the working range, \ blue-violet
... , . . , . t * Vl°'et
dilute the sample with zinc-free water or 4 red-violet
concentrate it in a silica dish. If the 5 red-violet
sample has been preserved with acid,
evaporate a portion to dryness in a silica 5. Calculation
dish to remove the excess acid. Do not .,, w Zn
!••••• • mg/IZn = —S6
neutralize with sodium or ammonium ml sample
hydroxide because these alkalis usually
contain excessive amounts of zinc. Using g ' precjsion and Accuracy
a pH meter and accounting for any di-
lution, adjust the sample to pH 2 to 3 A synthetic unknown sample contain-
with HC1. Transfer 10.0 ml to a sepa- ing 650 /tg/1 Zn, 500 /*g/l Al, 50 ^g/l
ratory funnel. Complete the analysis as Cd, 110 ng/\ Cr, 470 /ig/l Cu, 300
described in U 4«, beginning with the Mg/1 Fe, 70 jtg/1 Pb, 120 fig/I Mn,
words "To each funnel add 5.0 ml ace- and 150 /ig/1 Ag in distilled water was
tate buffer" and continuing to the end of analyzed in 46 laboratories by the dithi-
the paragraph. zone method with a relative standard
d. Visual comparison: If a photomet- deviation of 18.2% and a relative error
ric instrument is not available, run the of 25.9%.
Dithizone Method II
I. Principle hydroxyethyDdithiocarbamyl ion and
cyanide ion, which prevents moderate
Zinc is separated from other metals concentrations of cadmium, copper,
by extraction with dithizone and is then lead, and nickel from reacting with di-
determined by measuring the color of thizonc. If excessive amounts of these
the zinc-dithizone complex in carbon metals are present, follow the special
tetrachloride. Specificity in the separa- procedure given in 1J4£2) below.
tion is achieved by extracting from a The color reaction is extremely sensi-
nearly neutral solution containing bis(2- tive; avoid introducing extraneous zinc
494
-------
during the analysis. Contamination may
arise from water, reagents, and glass-
ware, such as beakers and separatory
funnels, on which zinc has been ad-
sorbed during previous use. Appreciable
blanks are generally found and the ana-
lyst must satisfy himself that these blanks
are representative and reproducible.
2. Apparatus
a. Colon-metric equipment: One of
the following is required:
1) Spectrophotometer, for use at 535
nm, providing a light path of 1 cm or
longer.
2) Filter photometer, providing a
light path of 1 cm or longer and
equipped with a greenish yellow filter
with maximum transmittance near 535
nm.
b. Separatory funnels, 125-ml,
Squibb form, with ground-glass stop-
pers.
3. Reagents
a. Standard zinc solution: Dissolve
1,000 g zinc metal in 10 ml 1 + 1
HNCh. Dilute and boil to expel oxides
of nitrogen. Transfer to a 1,000-ml vol-
umetric flask and dilute to volume; 1.00
ml=1.00mgZn.
b. Redistilled water: Distilled water
redistilled in all-glass apparatus.
c. tyethyl red indicator: Dissolve 0.1
g methyl red sodium salt and dilute to
100 ml with distilled water.
d. Sodium citrate solution: Dissolve
10 g Na3C6HiO7-2H2O in 90 ml wa-
ter. Shake with 10 ml dithizone solution
I to remove zinc, then filter.
e. Ammonium hydroxide\ NhhOH,
cone: Prepare according to directions in
Section 305C.3*.
/ Potassium cyanide solution: Dis-
solve 5 g KCN in 95 ml redistilled wa-
ter. (CAUTION: Toxic—take care to
avoid ingestion.)
g. Acetic acid, cone.
b. Carbon tetrachloride, CCU, zinc-
free. CAUTION.- Carbon tetrachloride is a
toxic substance. Long-continued absorp-
tion of small amounts may be hazard-
ous. While the solvent can be absorbed
through the skin, the primary danger is
through inhalation of the vapor. Pre-
pare reagents and extract standards and
samples with.carbon tetrachloride in a
well-ventilated hood.
/. Bis (2-hydroxyethyD dithiocarba-
mate solution: Dissolve 4.0 g dietha-
nolamine and 1 ml CS2 in 40 ml methyl
alcohol. Prepare every 3 or 4 days.
/. Ditbizone solution: Dilute 50 ml
stock dithizone II solution (carbon tet-
rachloride), prepared in accordance
with Section 30lC.I14£, to 250 ml with
CCU. Prepare fresh daily.
k. Sodium suljide solution /.- Dissolve
3.0 g Na2S-9H2O or 1.65 g
NazS-SHaO in 100 ml zinc-free water.
/. Sodium sulfide solution II: Prepare
just before use by diluting 4 ml sodium
sulfide solution I to 100 ml.
m. Nitric add, HNOj, 6N.
n. Hydrogen suljide, HzS.
4. Procedure
a. Preparation of calibration curve:
1) Prepare, just before use, a zinc so-
lution containing 2.0 //g Zn/ml by di-
luting 5 ml standard zinc solution to
250 ml, then diluting 10 ml of the latter
solution to 100 ml with redistilled wa-
ter. Pipet 5.00, 10.00, 15.00, and
20.00 ml, containing 10 to 40 pg Zn,
into separate 125-ml separatory funnels
495
-------
and adjust the volumes to about 20 ml.
Set up another funnel containing 20 ml
zinc-free water as a blank.
2) Add 2 drops methyl red indicator
and 2.0 ml sodium citrate solution to
each funnel. If the indicator is not yel-
low, add cone NhtaOH a drop at a time
until it just turns yellow. Add 1.0 ml
potassium cyanide solution and then
acetic acid, a drop at a time, until the in-
dicator just turns a neutral peach color.
3) Extract the methyl red by shaking
with 5 ml CCU. Discard the yellow
CCU layer. Add 1 ml dithiocarbamate
solution. Extract with 10 ml dithizone
solution, shaking for 1 min.
Draw off the CCU layer into another
separatory funnel and repeat the extrac-
tion with successive 5-ml portions of
dithizone solution until the last one
shows no change from the green dithi-
zone color. Discard the aqueous layer.
4) Shake the combined dithizone ex-
tracts with a 10-ml portion of sodium
sulfide solution II, separate the layers,
and repeat the washing with further 10-
ml portions of NazS solution until the
unreacted dithizone solution has been
removed completely, as shown by color
of the aqueous layer, which remains col-
orless or very pale yellow; usually three
washings are sufficient.
Remove water adhering to the stem
of the funnel with a cotton swab and
drain the pink CCU solution into a dry
JO-ml volumetric flask. Use a few mil-
lilitcrs of fresh CCU to rinse the last
droplets from the funnel and dilute to
the mark with fresh CCU.
5) Determine the absorbance of the
zinc dithi/.onatc solutions at 535 nm, us-
ing CCU as a reference. Plot an absorb-
ance-concentration curve after sub-
tracting the absorbance of the blank.
The calibration curve is linear if mono-
chromatic light is used.
6) Clean separatory funnels by shak-
ing several minutes successively with
HNOs, distilled water, and finally a
mixture of 5 ml sodium citrate and 5 ml
dithizone, to minimize the large or er-
ratic blanks that result from the adsorp-
tion of zinc on the glass surface. If pos-
sible, reserve separatory funnels
exclusively for the zinc determination
and do not use for other purposes.
b. Treatment of sample-.
1) Digest sample as directed under
Preliminary Treatment, Section
301C.II. Transfer a portion containing
10 to 40 /*g Zn to a clean 125-ml
separatory funnel and adjust the vol-
ume to about 20 ml. Determine the zinc
in -this solution exactly as described in
the preceding procedure for preparing
the calibration curve.
If more than 30 ml of dithizone solu-
tion is needed to extract the zinc com-
pletely, the portion taken contains too
much zinc or the quantity of other met-
als that react with dithizone exceeds the
amount that can be withheld by the
complexing agent. In the latter case, fol-
low the procedure in U 4£2) below.
2) Separation of excessive amounts of
cadmium, copper, and lead—When the
quantity of these metals, separately or
jointly, exceeds 2 mg in the portion
taken, in a 100-ml beaker adjust the
volume to about 20 ml. Adjust acidity to
0.4 to 0.5N* by adding dilute UNO3 or
NH^OH as necessary. Pass H:S into
the cold solution for 5 min. Filter off the
* The normalities of the solutions obtained in the
preliminary treatment are approximately 3-V fur the
MNO)-HiSO« digestion and approximately 0.8.V for
the HNOj-HCIO. digestion.
496
-------
precipitated sulfidcs using a sintered-
glass filter and wash the precipitate with
two small portions of hot water. Boil the
filtrate 3 to 4 min to remove HaS, cool,
transfer to a separatory funnel, and de-
termine the zinc as described in 1f
et seq.
5. Calculation
mg/l Zn =
MgZn
100
ml sample ml portion
Zincon Method*
1. General Discussion
a. Principle: Zinc forms a blue com-
plex with 2-carboxy-2'-hydroxy-5'-sul-
foformazyl benzene (zincon) in a solu-
tion buffered to pH 9.0. Other heavy
metals likewise form colored complexes.
Heavy metals, including zinc, are com-
plexed by cyanide. Chloral hydrate is
added specifically to free the zinc from
its cyanide complex. The zinc-zincon
complex is measured before other heavy
metal-cyanide complexes are destroyed
by chloral hydrate. Sodium ascorbate
reduces the interference of manganese.
The final solutions are unstable and the
procedure is designed to minimize the
effects of color fading.
b. Interference: The following ions
interfere in concentrations exceeding
those listed:
Ion
Cd (II)
Al (III)
Mn(H)
Fe(III)
Fe(H)
mg/l
I
5
y
7
9
Ion
Cr (III)
Nidi)
Cu(II)
Co (II)
CKMII)
mg/l
10
20
30
JO
50
c. Minimum detectable quantity: 1
MgZn.
2. Apparatus
Colorimetric equipment: One of the
following is required:
a. Spectrophototneter, for measure-
ments at 620 nm, providing a light path
of 1 cm or longer.
b. Filter photometer, providing a
light path of 1 cm or longer and
equipped with a red filter having maxi-
mum transmittance near 620 nm. De-
viation from Beer's law occurs when the
filter band pass exceeds 20 nm.
3. Reagents
a. Zinc-free water, for rinsing of ap-
paratus and preparation of solutions and
dilutions. Prepare as directed in Section
323C.3«.
b. Stock zinc solution: Prepare as di-
rected in Section 323C.3£».
c. Standard zinc solution: Dilute
10.00 ml stock zinc solution to 100 ml
with zinc-free water; 1.00 ml= 10.0 fig
Zn.
d. Sodium ascorbate, fine granular
powder.t
e. Potassium cyanide solution: Dis-
solve 1.00 g KCN in 50 ml zinc-free
water and dilute to 100 ml. This solu-
tion is stable for approximately 60 days.
* This method, with modifications, is identical in
.source and substance 10 ASTM D1691 -67.
t Hoffman-LaRochc or equivalent.
497
-------
CAUTION : Poison—potassium cyanide is
extremely poisonous. Observe more
than customary precautions in its han-
dling. Never use mouth pipets to deliver
volumes of cyanide solution.
f. Buffer solution, pH 9.0: Prepare
IN NaOH by dissolving 40 g sodium
hydroxide in 500 ml zinc-free water and
diluting to 1,000 ml. Dilute 213 ml IN
NaOH to approximately 600 ml with
zinc-free water. Dissolve 37.8 g KC1
and 3 1.0 g FbBOj in the solution and
dilute to 1 1.
g. Zincon reagent: Grind the entire
supply of zincon powder and make a
uniform mixture. Dissolve 130 mg
powdered 2-carboxy-2'-hydroxy-5'-sul-
foformazyl benzene (zincon) in 100 ml
methyl alcohol (methanol). Let stand
overnight or use a magnetic stirrer in a
closed flask to complete solution.
b. Chloral hydrate solution: Dissolve
10.0 g chloral hydrate in 50 ml zinc-free
water and dilun to 100 ml. Filter if nec-
essary.
i. Hydrochloric acid, HC1, cone.
j. Sodium hydroxide, NaOH, 6N.
4. Procedure
a. Preparation of colorimetric stand-
ards: To a series of thoroughly cleansed
50-ml erlenmeyer flasks, add 0, 0.25,
0.50, 1.00, 3.00, 5.00, and 7.00 ml
standard zinc solution equivalent, to
provide 0, 2.50, 5.00, 10.0, 30.0, 50.0,
and 70.0 Mg Zn, respectively. Bring
each volume to 10.0 ml by adding zinc-
free water. To each flask add, in se-
quence, mixing thoroughly after each
addition, 0.5 g sodium ascorbate, 1.0 ml
KCN solution, 5.0 ml buffer solution,
and 3.0 ml zincon solution. Add 3.0 ml
chloral hydrate solution, note the time,
and mix. Transfer to the absorption cell
and measure the absorbance at 620 nm
exactly 5 min after adding the chloral
hydrate solution. Use the treated blank
as the reference solution for initial bal-
ancing of the photometer. For greater
accuracy in the range below 10 Mg Zn,
prepare a separate calibration curve.
b. Treatment of samples: If dissolved
zinc is to be determined, filter the
sample. If total zinc is to be determined,
add 1 ml cone HC1 to 50 ml thoroughly
mixed sample and mix well. Filter and
adjust to pH 7 with 6NNaOH. Trans-
fer a 10.0-ml portion of sample contain-
ing not more than 70 Mg Zn to a 50-ml
erlenmeyer flask. Complete the analysis
as described in If 4a above, beginning
with the words "To each flask add, in
sequence . . . ," and continue to the end
of the paragraph.
Prepare as a reference solution a
sample portion treated as above, except
that 3.0 ml zinc-free water is substituted
for the 3.0 ml chloral hydrate. Use this
to compensate for color, turbidity, or in-
terference not eliminated by the proce-
dure. Prepare as nearly simultaneously
as possible with the sample portion.
5. Calculation
mg/l Zn
ml sample
6. Precision and Accuracy
A synthetic unknown sample contain-
ing 650 /ig/1 Zn, 500 Mg/l Al. 50 Mg/l
Cd, 110 Mg/l Cr, 470 Mg/l Cu, 300
Mg/l Fe, 70 Mg/l Pb, 120 Mg/l Mn,
and 150 Mg/l Ag in distilled water was
analyzed in four laboratories by the zin-
con method with a relative standard de-
viation of 13.9% and a relative error of
17.4%.
498
-------
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570
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575 Gulbrandsen, R. A. Chemical composition of phosphorites of the Phosphoria
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(
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TECHNICAL REPORT DATA
(Please read Instructions on the reverse before completing)
1 REPORT NO.
EPA-600/1-78-034
2.
3. RECIPIENT'S ACCESSION-NO.
4. TITLE AND SUBTITLE
ZINC
5. REPORT DATE
Mav 1978
6. PERFORMING ORGANIZATION CODE
7. AUTHOR(S)
Subcommittee on Zinc
8. PERFORMING ORGANIZATION REPORT NO.
9. PERFORMING ORGANIZATION NAME AND ADDRESS
Committee on Medical and Biologic Effects of
Environmental Pollutants
National Academy of Sciences
Washington, D.C. 20460
10. PROGRAM ELEMENT NO.
1AA601
11. CONTRACT/GRANT NO.
68-02-1226
12. SPONSORING AGENCY NAME AND ADDRESS
Health Effects Research Laboratory
Office of Research and Development
U.S. Environmental Protection Agency
Research Triangle Park, N.C. 27711
13. TYPE OF REPORT AND PERIOD COVERED
RTP,NC
14. SPONSORING AGENCY CODE
EPA 600/11
15. SUPPLEMENTARY NOTES
16. ABSTRACT
This report summarizes the available information on zinc as it relates to its effects
on man and his environment. Zinc is found in most soils, but some areas are deficien
in it. Metallurgic operations contribute to zinc contamination in air, water and soil
Trace amounts of zinc are essential for normal growth in plants, animals and humans,
however, excessive levels can bring on zinc toxicosis. Zinc deficiency is known to
have caused congenital malformations in pregnant rats. Severe liver disease is
commonly associated with loss of total body zinc. Zinc is not a highly toxic subs-
tance. Zinc toxicosis may occur only when very high dose levels overwhelm the homeo-
static mechanisms controlling zinc uptake and excretion. Reports suggest humans may
ingest 500 mg to 1 g or more daily without adverse effects. Ten or more g taken as a
single oral dose may produce gastrointestinal distress, including nausea, vomiting
and diarrhea. There are also suggestions in the literature that even higher dosage
may produce dizziness and perhaps increase blood levels of pancreatic enzymes.
Inhalation of zinc has been related to metal fume fever, an acute disability of
short duration that can occur when fume is inhaled from metal heated to a temperatur
above its melting point. With repeated exposure, some degree of tolerance may be
built up, but it will be lost when exposure to fume ceases for a period as short as
two days. The pathogenesis of this disorder, including the role of zinc in it, is no
17.
KEY WORDS AND DOCUMENT ANALYSIS
DESCRIPTORS
b.IDENTIFIERS/OPEN ENDED TERMS
c. COSATi Field/Group
zinc
air pollution
toxicity
health
ecology
chemical analysis
06 H, F, T
13. DISTRIBUTION STATEMENT
RELEASE TO PUBLIC
19. SECURITY CLASS (This Report)
21. NO. OF PAGES
744
20. SECURITY CLASS (Tltis page)
22. PRICE
EPA Form 2220-1 (9-73)
736
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