oEPA
United States
Environmental Protection
Agency
Office of Pesticide Programs
Washington, DC 20460
EPA 540/9-86-133
June 1986
Hazard Evaluation Division
Standard Evaluation Procedure
Non-Target Plants: Vegetative Vigor -
TierS 1 and 2 Support Document #35.
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EPA 540/9-86-133
June 1986
HAZARD EVALUATION DIVISION
STANDARD EVALUATION PROCEDURE
NON-TARGET PLANTS: VEGETATIVE VIGOR - TIERS 1 AND 2
Prepared by
Robert W. Hoist, Ph.D.
Standard Evaluation Procedures Project Manager
Stephen L. Johnson
Hazard Evaluation Division
Office of Pesticide Programs
United States Environmental Protection Agency
Office of Pesticide Programs
Washington, D.C. 20460
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TABLE OF CONTENTS
Page
I. INTRODUCTION
A. Purpose of the Standard Evaluation
Procedure 1
B. Background Information . 1
C. Objectives of Vegetative Vigor Tests 1
1. Tier 1 Test 1
2. Tier 2 Test 2
II. INFORMATION TO BE SUPPLIED 2
III. DATA INTERPRETATION 2
IV. THE DATA EVALUATION PROCESS
A. Identify Data Gaps 3
B. Assess the Appropriateness and Adeguacy of
the Data 3
C. Report Preparation 4
D. Conclude if the Reguested Action
is Supportable 4
V. APPENDICES
Appendix 1: Information Reguested of the
Registrant 5
Appendix 2: Specific Questions for the
Reviewer 9
Appendix 3: Sample Standard Format for
Preparation of Scientific
Reviews 12
REFERENCES 13
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NUN-TARGET PLANTS-. VEGETATIVE VIGOR - TIERS 1 AND 2
I. INTRODUCTION
A. Purpose of the Standard Evaluation Procedure
This Standard Evaluation Procedure is designed to aid Ecologi-
cal Effects Branch (EEB) data reviewers in their evaluations of
preliminary (Tier 1) laboratory vegetative vigor studies submitted
by registrants in the assessment of pesticide effects on non-target
plants. This document is also designed to aid EEB reviewers in
their evaluations of laboratory/greenhouse/smal1 field plot (Tier 2)
vegetative vigor"S"tudies submitted by registrants for the same
purpose.
B. Background Information
Vegetative vigor studies are designed to provide phytotoxicity
data on a pesticide. These phytotoxicity data are needed to evalu-
ate the effect of the level of pes.ticide .exposure to non-target
terrestrial plants and to assess the impact of pesticides on endan-
gered and threatened plants as noted under the Endangered Species
Act. The preliminary level (Tier 1) study evaluates the effect of
the maximum exposure level while the greenhouse/laboratory/small
field plot (Tier 2) study evaluates the effects of differing exposure
levels. Where a phytotoxic effect is noted in one or more plants,
further terrestrial vegetative vigor studies may be required. These
studies are required by 40 CFR § 158.150 to support the registration
of any pesticide intended for outdoor use under the Federal Insecti-
cide, Fungicide and Rodenticide Act (FIFRA), as amended.
Pesticides with outdoor use patterns that do not readily release
the pesticide to the environment do not have to be evaluated using .
this phytotoxicity test. These use patterns include tree injection,
subsurface soil applications, recapture systems, wick applications,
and swimming pool uses. If any of these use patterns do readily
expose non-target plants to the pesticide, as through vapors, the
pesticide phytotoxicity potential may need to be evaluated.
C• Objectives of Vegetative Vigor Tests
1. Tier 1 Test
The objective of the Tier 1 vegetative vigor test is to deter-
mine if a pesticide exerts a detrimental effect to plants during
the vegetative growth period of their development as a result of
post-emergent foliar or soil applications. The test is performed
on species from a cross-section of the non-target terrestrial plant
population that have been historically used for this type of testing
and, therefore, have known types of responses. This is a maximum
dose test designed to evaluate the phytotoxic effects of the pesti-
cide quickly at the one dose.
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2. Tier 2 Test
The objective of the Tier 2 vegetative vigor test is to deter-
mine if a pesticide exerts a detrimental effect to plants during
the vegetative growth period of their development. The test is per-
formed on species from a cross-section of the non-target terrestrial
plant population that have been historically used for this type of
testing and, therefore, have known types of responses. This is a
multiple dose test designed to evaluate the phytotoxic effects of
the pesticide over a wide range of anticipated pesticide quantities
as may be tound in the environment.
II. INFORMATION TO BE SUPPLIED v -~~~
The registrant's report on vegetative vigor studies should
include all information necessary to provide: 1) a complete and
accurate description of the laboratory/greenhouse treatments and
procedures, 2) sampling data and phytotoxicity rating, 3) data on
storage of the plant material until analysis, if so performed,
4) any chemical analysis of the plant material as to chemical con-
tent, if so performed, 5) reporting of the data, rating system and
statistical analysis, and 6) quality control measures/precautions
taken to ensure the fidelity of the operations.
A guideline of specific information that should be included in
the registrant's report on vegitative vigor studies is provided in
Appendix 1 of this document. The lists of requested i rif orroat ion
and reviewer aids are derived from the Pesticide Assessment Guide-
lines, Subdivision J; Hazard Evaluation of Non-Target Plants,
which is complemented by this Standard Evaluation Procedure.
III. DATA INTERPRETATION
The acceptability of the study results will depend upon whether
the test requirements/standards are followed. If a deviation is
made, a determination must be made as to whether the deviation has
changed the quality of the results in such a manner that the results
cannot t>e extrapolated to the natural environment. There should be
little or no deviation from the liberalized standards prescribed in
this study.
The results of the phytotoxicity test of the chemical with re-
spect to the quantity applied post-emergent to the foliage or soil
are important. The concentration of the chemical in the carrier is
important in that stronger concentrations than normally used can
lead to burning and necrosis or undesired growth. Subtoxic concen-
trations, on the other hand, may cause unwanted rapid growth.
Plants can recover from certain types of injury that will have
little or no effect on the esthetic or economic value of the plant(s)
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tested or to wnich an evaluation is made. Therefore, it is important
that a minimum of two weeks of observations be made after application
of the pesticide; three to four weeks are preferable.
A decision point to proceed on to the next higher test is a 25%
detrimental effect, i.e., a 25% reduction or over-enhancement in
plant growth or injury as compared to the untreated controls. This
level is considered to be that point at which the plants will not
recover to their full esthetic value, economic value, or reproductive
potential as in the case of the maintenance of the endangered or
threatened species.
IV. THE DATA EVALUATION PROCESS ^ -~~
Upon careful examination of the information/data supplied by
the registrant in his submission to the Agency, the reviewer shall
evaluate the data as follows.
A. Identify Data Gaps
Using Appendix 1 of this document as a guide, the reviewer
should look for data gaps - omissions in the information supplied
by the registrant in his report. These should be duly noted in the
reviewer's report, and a judgment made as to which are considered
significant enough to adversely affect the review process. Those
so identified should be communicated back to the registrant by the
Product Manager for corrective action.
B. Assess the Appropriateness and Adequacy of the Data
The data reviewer then considers the appropriateness, i.e., the
intended use pattern, and adequacy of the data/information that has
been supplied. Appendix 1 of this document is a useful guide to.the
various parameters that need to be considered. Appendix 2 provides
specific questions that should be answered by the reviewer during
the study evaluation process. Statistical treatments of the data
should be independently verified and the quality control precautions
noted.
As an adjunct to these, the reviewer should draw upon the tech-
nical guidance in the reviewer aids materials that are available.
(See also the recommended references in Subdivision J - Hazard Eval-
uation; Non-Target Plants.) A listing of additional source materials
is located in the References section of this document.
In addition to the data gaps noted above, any perceived defici-
encies in the data/information supplied should be identified. A
statement as to these deficiencies should be made in the reviewer's
report and corrective action to resolve them should be provided.
This information can be relayed to the registrant by the Product
Manager for appropriate action.
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C. Report Preparation
The Agency reviewer prepares a standard review report following
the standard format for preparation of scientific reviews as provided
in Appendix 3 of this document. All important information provided
by the registrant including the methodology and results should be
summarized in order that future evaluations can be made. The
results may be expressed in the form of tables where specific values
are related. Figures (graphs) may be provided but are not to be
the sole source of the values needed for future evaluations.
D. Conclude if the Requested Action is Supportable
Lastly, the reviewer considers the results of the vegetative
vigor studies and makes a judgment as to whether they support the
requested registration action of the data submitter. If the data
are not supportive, possible alternative action(s) that may be taken
by the registrant, such as label modifications, are suggested. If
deficiencies/omissions exist in the submitted data, the reviewer
may have to defer judgment until such time as appropriate corrective
action has been rendered by the registrant.
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APPENDIX 1
INFORMATION REQUESTED OF THE REGISTRANT
The registrant's report on preliminary vegetative vigor studies
should include all information necessary to provide: 1) a complete
and accurate description of the laboratory/greenhouse/field treat-
ments and procedures, 2) sampling data and phytotoxicity rating,
3) data on storage of the plant material if so performed, 4) any
chemical analysis of the plant material as to chemical content,
5) reporting of the data, rating system, and statistical analysis,
and 6) quality control measures/precautions taken to ensure the
fidelity of the operations.
Specifically, each laboratory/greenhouse/small field plot vege-
tative vigor report should include the following information.
I. General
0 Cooperator or researcher (name and address), test location
(county and state; country, if outside of the U.S.A.), and date of
study;
0 Name (and signature), title, organization, address, and tele-
phone number of the person(s) responsible for planning/supervising/
monitoring;
0 Trial identification number;
0 Quality assurance indicating: control measures/precautions
followed to ensure the fidelity of the phytotoxicity determinations,
record-keeping procedures, and availability of logbooks; skill of
the laboratory personnel; equipment status of the laboratory or
greenhouse; degree of adherence to good laboratory practices; and
degree of adherence to good agricultural practices in maintaining
healthly plants; and
0 Other information the registrant considers appropriate and
relevant to provide a complete and thorough description of the test
procedures and results.
II. Test Substance (Pesticide)
0 Identification of the test pesticide active ingredient (ai)
including chemical name, common name (ANSI, BSI, ISO, WSSA), and
Company developmental/experimental name;
0 Active ingredient percentage in the technical grade material
or in the manufacturing-use product, if the technical grade material.
is unavailable for test purposes;
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0 Solvent used to dissolve and apply the pesticide if the
pesticide is insoluble in water or other intended carrier;
0 Dose rate(s) in terms of active ingredient per area of land
or of leaf (if leaf-area-index is provided);
0 For Tier 1, dose rate(s) in terms of the maximum label
rate, or if the registrant has shown that the maximum quantity that
will be present in the non-target area is significantly less than
the maximum label rate, the dose equal to or no less than three
times that maximum environmental quantity;
0 For Tier 2, dose rate(s) in terms of less than the maximum
label rate, with dosages in a geometrical progression of no more
than two-fold and with subtoxic (< £059 level) and non-toxic (no-
observable -effect- level ) concentrations;
0 Method of application including equipment type; and
0 Number of applications.
III. Plant Species
0 For Tier 1, identification of the six dicotyledonae species
and four monocotyledonae species with family identification. The
six dicots are to be of at least four different families and the
moncots of at least two families. Soybeans, corn, and a dicot root
crop like carrot are the required species. The proposed species
and families as originally provided in Subpart J of the proposed
guidelines [FR notice of 3 November 198U] are given below and are
acceptable for the laboratory/greenhouse vegetative vigor test.
Family
Solanaceae
Cucurbitaceae
Compos itae
Leguminosae
Cruciferae
Umbelliferae
Gramineae
Gramineae
Gramineae
Amaryllidaceae
Species
Lycopersicon esculentum
Cucumis sativus
Lactuca sativa
Glycine max
(Innoculated with Rhizobium japonicum)
Common
Tomato
Cucumber
Lettuce
Soybean
Brassica oleracea
Daucus carota
Avena sativa
Lolium perenne
Zea mays
Allium cepa
Cabbage
Carrot
Oat
Perennial Ryegrass
Corn
Onion
0 For Tier 2, identification of the plant species tested
including those phytotoxically affected in the Tier 1 test;
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0 Identification of the cultivar(s) of the plant species or
assignment of an identification number of the cultivar used and the
plant source;
0 Identification of the number of replicates and the number of
plants per replicate per dose; and
0 Identification of the date of planting, date of pesticide
application, and date of phytotoxicity rating or harvest.
IV. site of the Test
0 Site description of the vegetative vigor study such as the
type of growth chamber, greenhouse, or field (small field plots);
0 Location of the test site;
0 Climatological data during the test (records of applicable
conditions for the type of site, i.e., temperature, thermoperiod,
rainfall or water regime, light regime .- .intensity and quality,
relative humidity, wind speed);
0 Field lay-out (for small field plots), e.g., size and number
of control and experimental plots; number of plants per plot/unit
area;
0 Pot, plate or row density of plants;
0 Cultural practices such as cultivation and irrigation; and
0 Substrate characteristics (name/designation of soil type and
its physical and chemical properties, including pH and percent
organic matter). .
V. Results
0 Phytotoxicity rating (including a description of the rating
system) for each plant or population in the test;
0 Reporting of weight, height, or other growth parameters that
may have been measured to ascertain toxic effects of the pesticide
upon the plants with dates of observations; and
0 Statistical analysis of the results including an environmen-
tal or effective concentration effect (EC) value. (Note, with only
one dose level in Tier 1, there will be only a percent effect
level at a specific concentration which is then compared to 25% of
the growth [mass or rate] of the control.)
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VI. Evaluations
0 For Tier 1 studies, determination as to whether Tier 2
studies would be required due to phytotoxic effects noted in one or
more of the tested species.
0 For Tier 2 studies, determination as to whether Tier 3 tests
(terrestrial field study) would be required due to phytotoxic effects
noted on one or more of the tested species.
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APPENDIX 2
SPECIFIC QUESTIONS FOR THE REVIEWER
The following questions are provided to aid the reviewer in
performing the standard evaluation procedure in a scientific manner
and in acquiring the necessary information to complete a standard
format for preparation of scientific reviews.
I. General
0 Was the name of_ the cooperator or researcher (name and
address), test location (county and state; country, if outside of
the U.S.A.), ana date of study provided?
0 Was the name Jand signature), title, organization, address,
and telephone number of the person(s) responsible for planning/super-
vising/monitoring and, for small field plot studies, applying the
pesticide provided?
0 Was the trial identification number provided?
0 Were quality assurance control measures/precautions indicated?
II. Test Chemical
0 Is the test chemical being used the technical grade, or if
not available, the manufacturing-use product with the highest
percentage of active ingredient?
0 Is the active ingredient percentage or degree of purity of
the chemical given?
0 If a solvent was used, was it used at concentrations that
were not phytotoxic and was a solvent control used?
0 Is the dose given in quantity per unit area (of plant or
land surface) or in tank concentration?
0 For Tier 1, was the dose equal to or greater than the maxi-
mum label rate, or if registrant has shown that the maximum quantity
that will be present in the non-target area is significantly less
than the maximum label rate, was the dose equal to or no less than
three times that maximum environmental quantity?
0 For Tier 2, was the maximum dose less than the maximum label
rate?
0 For Tier 2, were the additional dosages of a geometric pro-
gression of no more than two-fold, e.g., 0.1, 0.2, 0.4, 0.8, 1.6 kg/ha?
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0 Were a subtoxic (£€59 level) and a non-toxic (no-observable-
_ef feet-level) concentration evaluated?
III. Test Species
0 For Tier 1, were at least ten different species tested?
0 For Tier 1, were the ten species split between monocots and
dicots, four and six, respectively?
0 For Tier 1, were the ten species from six different f-amilies
and the family names provided? __
0 For Tier 1, were two of the species tested soybeans and corn
and was the third species a dicot root crop?
0 For Tier 2, were at least those species that were phytotox-
ically affected in Tier 1 tested?
0 Where various cultivars could be used such as in the case
of most agronomic and horticultural plants, were cultivar or varietal
names provided?
0 Were the plant sources provided?
0 Were there at least three replicates with five plants per
replicate for each dose level?
0 Were the plants at least one week to four weeks post-emergent
and in the exponential stage of growth at the time of application?
0 Were the plants healthy and not in a state of stress?
0 Were endangered or threatened plant species not used?
IV. Test Procedures
0 Was the test site specified, i.e., greenhouse, growth cham-
ber, or small field plot?
0 Were the environmental conditions that prevailed during the
test (temperature and thermoperiod, light regime - intensity and
quality, relative humidity, rainfall or watering regime, wind)
provided as appropriate for the site?
0 Were the environmental conditions that prevailed during the
test those most favorable and most typical to the growth of the
plants used? Were these conditions referenced?
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0 Was the test durat.ion at least two weeks in length?
0 Were observations taken at least weekly?
0 Was the method of pesticide application including the type
of application equipment employed given?
V. Reporting
0 Were the detrimental effects reported as severity of phyto-
toxicity, growth inhibition, etc. (rating or percentage)?
0 If a rating system was used, was an explanation provided?
0 Were abnormal changes in growth, development and/or morpho-
logy reported as compared to the controls or "normal" jplants?
0 Though not required, were direct measurements of height
and weight of the plants provided?
0 Were the results statistically analyzed? Note that care
should be taken in interpreting the statistical results where the
sample size is small.
VI. Evaluation
0 Were the results tabulated to indicate a percentage effect
level (EC value) for each species as compared to the untreated con-
trol plants?
0 For Tier 1 studies, was a determination made as to whether
Tier 2 tests should be performed if any of the Tier 1 species were
detrimentally affected (greater than 25% detrimental effect on
growth)?
0 For Tier 2 studies, were 25 and 50 percent detrimental
effect levels determined for those plant species of Tier 1 that
showed a phytotoxic effect to the chemical?
0 For Tier 2 studies, was a determination made as to whether
Tier 3 tests (terrestrial field study) should be performed if any
of the Tier 2 species were detrimentally affected (greater than 25%
detrimental effect on growth)?
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APPENDIX 3
SAMPLE STANDARD FORMAT FOR PREPARATION. OF SCIENTIFIC REVIEWS
The following format shall be used in documenting the review of
the Subdivision J - Hazard Evaluation: Non-Target Plants - Tier 1
and Tier 2 Vegetative Vigor Studies.
Chemical: (Common Name)
Formulation: (Percent Active Ingredient)
Study/Action: (Purpose of the Submission)
Study Identification:
(Subdivision J Test Title)
(Reference or Registrant Data Information with
Study Number) .•
(EPA Accession Number)
Reviewer: (Name and Address of Reviewer; Date of Review)
Approval: (Quality Control Reviewer)
Conclusions: (Summary and Conclusion of Tests)
Acceptability and Recommendations:
(Decide as to (1) the scientific validity of the study
and (2) compliance to the Subdivision J - Vegetative
Vigor Tier 1 and Tier 2 Studies.)
Background: (Introductory Information and Directions for Use)
Discussion: 1. Study Identification
2. Materials and Methods
3. Reported Results
4. Reported Conclusions
5. Reviewer's Interpretation of Results and Conclusion
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REFERENCES
Truelove, B., ed. 1977. Research Methods in Weed Science. Southern
weed science Society. Auburn, AL: Auburn Printing, Inc.
Other scientific articles on vegetative vigor may be found in
the following journals:
Agronomy Journal
Environmental Science and Technology
Journal of Environmental Quality
Soil Science and Plant Nutrition
Weed Science
ft US. GOVERNMENT PHINTDra OFFICE: 1986 - 621-735 - 1302/60519
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