ASTM E1963-09(2014)
(Guide)Standard Guide for Conducting Terrestrial Plant Toxicity Tests
Standard Guide for Conducting Terrestrial Plant Toxicity Tests
SIGNIFICANCE AND USE
5.1 Terrestrial phytotoxicity tests are useful in assessing the effects of environmental samples or specific chemicals as a part of an ecological risk assessment (3-6, 12, 13).
5.2 Though inferences regarding higher-order ecological effects (population, community, or landscape) may be made from the results, these tests evaluate responses of individuals of one or more plant species to the test substance.
5.3 This guide is applicable for: (a) establishing phytotoxicity of organic and inorganic substances; (b) determining the phytotoxicity of environmental samples; (c) determining the phytotoxicity of sludges and hazardous wastes, (d) assessing the impact of discharge of toxicants to land, and (e) assessing the effectiveness of remediation efforts.
SCOPE
1.1 This guide covers practices for conducting plant toxicity tests using terrestrial plant species to determine effects of test substances on plant growth and development. Specific test procedures are presented in accompanying annexes.
1.2 Terrestrial plants are vital components of ecological landscapes. The populations and communities of plants influence the distribution and abundance of wildlife. Obviously, plants are the central focus of agriculture, forestry, and rangelands. Toxicity tests conducted under the guidelines and annexes presented herein can provide critical information regarding the effects of chemicals on the establishment and maintenance of terrestrial plant communities.
1.3 Toxic substances that prevent or reduce seed germination can have immediate and large impacts to crops. In natural systems, many desired species may be sensitive, while other species are tolerant. Such selective pressure can result in changes in species diversity, population dynamics, and community structure that may be considered undesirable. Similarly, toxic substances may impair the growth and development of seedlings resulting in decreased plant populations, decreased competitive abilities, reduced reproductive capacity, and lowered crop yield. For the purposes of this guide, test substances include pesticides, industrial chemicals, sludges, metals or metalloids, and hazardous wastes that could be added to soil. It also includes environmental samples that may have had any of these test substances incorporated into soil.
1.4 Terrestrial plants range from annuals, capable of completing a life-cycle in as little as a few weeks, to long-lived perennials that grow and reproduce for several hundreds of years. Procedures to evaluate chemical effects on plants range from short-term measures of physiological responses (for example, chlorophyll fluorescence) to field studies of trees over several years. Research and development of standardized plant tests have emphasized three categories of tests: (1) short-term, physiological endpoints (that is, biomarkers); (2) short-term tests conducted during the early stages of plant growth with several endpoints related to survival, growth, and development; and (3) life-cycle toxicity tests that emphasize reproductive success.
1.5 This guide is arranged by sections as follows:
Section
Title
1
Scope
2
Referenced Documents
3
Terminology
4
Summary of Phytotoxicity Tests
5
Significance and Use
6
Apparatus
7
Test Material
8
Hazards
9
Test Organisms
10
Sample Handling and Storage
11
Calibration and Standardization
12
Test Conditions
13
Interference and Limitations
14
Quality Assurance and Quality Control
15
Calculations and Interpretation of Results
16
Precision and Bias
1.6 The values stated in SI units are to be regarded as standard. No other units of measurement are included in this standard.
1.7 This standard does not purport to address all of the safety concerns, if any, associated with its use. It is the responsibility of the user of this standard to establish appropriate saf...
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Designation: E1963 − 09 (Reapproved 2014)
Standard Guide for
Conducting Terrestrial Plant Toxicity Tests
This standard is issued under the fixed designation E1963; the number immediately following the designation indicates the year of
original adoption or, in the case of revision, the year of last revision.Anumber in parentheses indicates the year of last reapproval.A
superscript epsilon (´) indicates an editorial change since the last revision or reapproval.
1. Scope tests conducted during the early stages of plant growth with
severalendpointsrelatedtosurvival,growth,anddevelopment;
1.1 Thisguidecoverspracticesforconductingplanttoxicity
and ( 3) life-cycle toxicity tests that emphasize reproductive
tests using terrestrial plant species to determine effects of test
success.
substances on plant growth and development. Specific test
procedures are presented in accompanying annexes. 1.5 This guide is arranged by sections as follows:
Section Title
1.2 Terrestrial plants are vital components of ecological
1 Scope
landscapes. The populations and communities of plants influ-
2 Referenced Documents
ence the distribution and abundance of wildlife. Obviously,
3 Terminology
4 Summary of Phytotoxicity Tests
plants are the central focus of agriculture, forestry, and range-
5 Significance and Use
lands. Toxicity tests conducted under the guidelines and
6 Apparatus
annexes presented herein can provide critical information 7 Test Material
8 Hazards
regarding the effects of chemicals on the establishment and
9 Test Organisms
maintenance of terrestrial plant communities.
10 Sample Handling and Storage
11 Calibration and Standardization
1.3 Toxic substances that prevent or reduce seed germina-
12 Test Conditions
tion can have immediate and large impacts to crops. In natural
13 Interference and Limitations
14 Quality Assurance and Quality Control
systems, many desired species may be sensitive, while other
15 Calculations and Interpretation of Results
species are tolerant. Such selective pressure can result in
16 Precision and Bias
changes in species diversity, population dynamics, and com-
1.6 The values stated in SI units are to be regarded as
munitystructurethatmaybeconsideredundesirable.Similarly,
standard. No other units of measurement are included in this
toxic substances may impair the growth and development of
standard.
seedlings resulting in decreased plant populations, decreased
1.7 This standard does not purport to address all of the
competitive abilities, reduced reproductive capacity, and low-
safety concerns, if any, associated with its use. It is the
ered crop yield. For the purposes of this guide, test substances
responsibility of the user of this standard to establish appro-
include pesticides, industrial chemicals, sludges, metals or
priate safety and health practices and determine the applica-
metalloids,andhazardouswastesthatcouldbeaddedtosoil.It
bility of regulatory limitations prior to use. Specific precau-
also includes environmental samples that may have had any of
tionary statements are given in Section 8.
these test substances incorporated into soil.
1.4 Terrestrial plants range from annuals, capable of com-
2. Referenced Documents
pleting a life-cycle in as little as a few weeks, to long-lived
2.1 ASTM Standards:
perennials that grow and reproduce for several hundreds of
D1193Specification for Reagent Water
years. Procedures to evaluate chemical effects on plants range
D4547Guide for Sampling Waste and Soils for Volatile
from short-term measures of physiological responses (for
Organic Compounds
example,chlorophyllfluorescence)tofieldstudiesoftreesover
D5633Practice for Sampling with a Scoop
several years. Research and development of standardized plant
E1598Practice for Conducting Early Seedling GrowthTests
tests have emphasized three categories of tests: (1) short-term,
(Withdrawn 2003)
physiological endpoints (that is, biomarkers); (2) short-term
E1733Guide for Use of Lighting in Laboratory Testing
1 2
ThisguideisunderthejurisdictionofASTMCommitteeE50onEnvironmental For referenced ASTM standards, visit the ASTM website, www.astm.org, or
Assessment, Risk Management and CorrectiveAction and is the direct responsibil- contact ASTM Customer Service at service@astm.org. For Annual Book of ASTM
ity of Subcommittee E50.47 on Biological Effects and Environmental Fate. Standards volume information, refer to the standard’s Document Summary page on
Current edition approved Oct. 1, 2014. Published December 2014. Originally the ASTM website.
published in 1998. Last previous edition published 2009 as E1963–09. DOI: The last approved version of this historical standard is referenced on
10.1520/E1963-09R14. www.astm.org.
Copyright © ASTM International, 100 Barr Harbor Drive, PO Box C700, West Conshohocken, PA 19428-2959. United States
E1963 − 09 (2014)
2.2 Code of Federal Regulations Standard: 3.2.9 toxicity endpoints, n—measurements of organism re-
CFR 49 sponse such as death, growth, developmental, or physiological
parameters resulting from exposure to toxic substances.
2.3 Other useful references have described phytotoxicity test
procedures(1-11) .
3.3 Definitions of Terms Specific to This Standard:
3.3.1 chlorotic, adj—the discoloration of shoots that occurs
as chlorophyll is degraded as a result of disease, toxic
3. Terminology
substances, nutrient deficiencies, or senescence.
3.1 General Terminology—The words “must,” “should,”
3.3.2 coleoptile, n—the protective tissues surrounding the
“may,”“ can,” and “might” have very specific meanings in this
growing shoot in a monocotyledonous plant.
guide. “Must” is used to express an absolute requirement, that
3.3.3 cotyledon, n—a primary leaf of the embryo in seeds,
is, to state that the test ought to be designed to satisfy the
only one in the monocotyledons, two in dicotyledons. In many
specified condition, unless the purpose of the test requires a
of the latter, such as the bean, they emerge above ground and
differentdesign.“Must”isonlyusedinconnectionwithfactors
appear as the first leaves.
that directly relate to the acceptability of the test (see Section
14). “Should” is used to state that the specified condition is
3.3.4 cutting, n—a vegetative segment of a plant, usually a
recommended and ought to be met if possible. Although
stem that contains several nodes and associated buds, that can
violationofone“should”israrelyaseriousmatter,violationof
be used to regenerate an entire plant.
several will often render the results questionable. Terms such
3.3.5 dead test plants, n—those individuals that expired
as“isdesirable,”“isoftendesirable,”and“mightbedesirable”
during the test observation period as indicated by severe
are used in connection with less important factors. “May” is
desiccation, withering, chlorosis, necrosis, or other symptoms
used to mean “is (are) allowed to,” “can” is used to mean “is
that indicate non-viability.
(are) able to,” and “might” is used to mean “could possibly.”
Thus the classic distinction between “may” and “can” is
3.3.6 desiccated, adj—the plant, or portion of the plant, that
preserved, and “might” is never used as a synonym for either is dried in comparison to the control plant.
“may” or “can.”
3.3.7 development, n—the series of steps involving cell
3.2 Definitions:
divisionandcelldifferentiationintovarioustissuesandorgans.
3.2.1 control, n—the treatment group in a toxicity test
3.3.8 dicotyledon, n—in the classification of plants, those
consisting of reference soil or artificial soil that duplicates all
having two seed leaves.
the conditions of the exposure treatments, but contains no test
3.3.9 dormancy, n—aspecialconditionofarrestedgrowthin
substance. The control is used to determine if there are any
which buds, embryos, or entire plants survive at lowered
statistical differences in endpoints related to the test substance.
metabolic activity levels. Special environmental cues such as
3.2.2 eluate, n—solutionobtainedfromwashingasolidwith
particular temperature regimes or photoperiods are required to
a solvent to remove adsorbed material.
activate metabolic processes and resume growth. Seeds that
3.2.3 hazardous substance, n—a material that can cause
require additional treatment besides adequate moisture and
deleterious effects to plants, microbes, or animals. (A hazard-
moderatetemperaturetogerminatearesaidtobedormant.(See
ous substance does not, in itself, present a risk unless an
quiescence.)
exposure potential exists.)
3.3.10 emergence, n—following germination of a plant, the
3.2.4 inhibition, n—a statistically lower value of any end-
early growth of a seedling that pushes the epicotyl through the
point compared to the control values that is related to environ-
soil surface.
mental concentration or application rate.
3.3.11 enhanced growth and yield, n—when a treated plant
3.2.5 leachate, n—water plus solutes that has percolated
exhibits shoot growth, root elongation, lateral root growth, or
through a column of soil or waste.
yield significantly greater than the control values, the plant is
3.2.6 test material, n—any formulation, dilution, etc. of a
“enhanced” or “stimulated.”
test substance.
3.3.12 epicotyl, n—that portion of an embryo or seedling
3.2.7 test substance, n—a chemical, formulation, eluate,
containing the shoot. It is delineated anatomically by the
sludge, or other agent or substance that is the target of the
transition zone which separates the epicotyl from the hypoco-
investigation in a toxicity test.
tyl.
3.2.8 toxicant, n—anagentormaterialcapableofproducing
3.3.13 fruits, n—the reproductive tissues derived from the
an adverse response (effect) in a biological system, adversely
ovary in the case of epigenous flowers or the ovary and
impacting structure or function or producing death.
accessory tissues in the case of hypigenous flowers.
3.3.14 germination, n—the physiological events associated
Available from Standardization Documents Order Desk, DODSSP, Bldg. 4,
withre-initiationofembryogrowthandmobilizationofreserve
Section D, 700 Robbins Ave., Philadelphia, PA 19111-5098, http://
nutrients in seeds. The emergence of the seedling radicle from
www.dodssp.daps.mil.
the seed coat defines the end of germination and the beginning
Theboldfacenumbersinparenthesesrefertothelistofreferencesattheendof
this guide. of early seedling growth.
E1963 − 09 (2014)
3.3.15 growth, n—a change in size or mass measured by casewherethetestsubstanceisevaluatedasanadditivetosoil,
length, height, volume, or mass. a range of concentrations is recommended. In tests of environ-
mental samples that already contain a putative phytotoxic
3.3.16 hypocotyl, n—that portion of an embryo or seedling
substance, the tests may be conducted with either the test soil
containing the root or radicle. It is delineated anatomically by
as collected from the field, or as diluted with a suitable
the transition zone which separates the epicotyl from the
reference soil. Another variant of the tests allows for
hypocotyl.
amendments, or spikes, of selected toxic substances to be
3.3.17 inhibited plant growth and yield, n— plant growth,
addedtoenvironmentalsamples.Finally,inthecaseoftheroot
root length and lateral root growth, or yield are “inhibited”
elongation assay, eluates, effluents, or other aqueous deriva-
whentheirmeasurementsaresignificantlylessthanthecontrol
tives of a soil sample are tested.
values.
4.2 Plants are exposed to the test substances in the form
3.3.18 lateral roots, n—roots growing off the primary roots,
described in the specific variations of the tests for a discrete
also referred to as secondary roots.
period of time that ranges from 96 h to several months. For
3.3.19 monocotyledon, n— in the classification of plants,
short tests, no nutrient additions or amendments are needed or
those having a single seed leaf.
recommended as the amendments may interact with the toxi-
3.3.20 mottled, adj—marked with lesions, spots or streaks cant and alter the toxicity response. For tests lasting more than
of different colors. This includes the discoloration of leaf
two weeks, nutrient additives may be warranted, depending on
margins. the test objectives, in order to maximize the potential for plant
growth and development. Thinning, culling, or replacing indi-
3.3.21 phytotoxicity, n—a lethal or sub-lethal response of
vidualplantsmustnotbedoneonceexposureofplantstoatest
plants to a toxicant.
substance has begun as such actions invalidate the test through
3.3.22 quiescence, n—a condition in buds, embryos, or
the introduction of bias or variable test duration among test
entire plants characterized by lowered metabolic rates and
organisms.At intermediate times, and at the conclusion of the
limitedornogrowth.Seedsthatgerminatewhensuppliedwith
exposure period, tallies of survival and measures of shoot
adequate moisture and moderate temperature are said to be
growth and development are made.
quiescient. (See dormancy.)
–2 –1
4.3 For phytotoxicity tests, 100 to 200 µmol m s of
3.3.23 radicle, n—the emerging root of an embryo during
visible light (or photosynthetically active radiation, 400 to 700
germination.
ηm) has been found to be a broadly applicable fluence rate. In
3.3.24 seed, n—the propagule of a plant derived from an
some cases, different light levels or spectral ranges (for
ovule. It consists of an embryo, a protective covering (seed
example, solar ultraviolet) may be required. Guide E1733.
coat), and may have storage tissue (endosperm).
4.4 Measured endpoints and other observational data are
3.3.25 shoot, n—the above-ground portion of a plant con-
used to calculate response levels in terms of ECxx or ICxx
sisting of stems, leaves, as well as any reproductive parts that
(where xx refers to a specified percentage response), or
may be attached.
categorical descriptions of phytotoxic effects (for example,
3.3.26 surviving plants, n—test plants that are alive at the
proportion of plants exhibiting abnormal development or other
time observations are recorded.
symptomatic indices that might be scored in qualitative terms)
relative to controls. These are interpreted to characterize
3.3.27 viable, adj—plants capable of resuming metabolic
phytotoxic effects attributed to test substances.
functions and growth are considered “viable.” Buds, embryos,
or entire plants may be dormant or quiescient and therefore
5. Significance and Use
exhibit no growth during the period of observation. Distin-
5.1 Terrestrial phytotoxicity tests are useful in assessing the
guishing dead plants from viable plants with certainty is
effectsofenvironmentalsamplesorspecificchemicalsasapart
difficult without special training and sophisticated measures of
of an ecological risk assessment (3-6, 12, 13).
metabolic function.
3.3.28 witherin
...
This document is not an ASTM standard and is intended only to provide the user of an ASTM standard an indication of what changes have been made to the previous version. Because
it may not be technically possible to adequately depict all changes accurately, ASTM recommends that users consult prior editions as appropriate. In all cases only the current version
of the standard as published by ASTM is to be considered the official document.
Designation: E1963 − 09 E1963 − 09 (Reapproved 2014)
Standard Guide for
Conducting Terrestrial Plant Toxicity Tests
This standard is issued under the fixed designation E1963; the number immediately following the designation indicates the year of
original adoption or, in the case of revision, the year of last revision. A number in parentheses indicates the year of last reapproval. A
superscript epsilon (´) indicates an editorial change since the last revision or reapproval.
1. Scope
1.1 This guide covers practices for conducting plant toxicity tests using terrestrial plant species to determine effects of test
substances on plant growth and development. Specific test procedures are presented in accompanying annexes.
1.2 Terrestrial plants are vital components of ecological landscapes. The populations and communities of plants influence the
distribution and abundance of wildlife. Obviously, plants are the central focus of agriculture, forestry, and rangelands. Toxicity tests
conducted under the guidelines and annexes presented herein can provide critical information regarding the effects of chemicals
on the establishment and maintenance of terrestrial plant communities.
1.3 Toxic substances that prevent or reduce seed germination can have immediate and large impacts to crops. In natural systems,
many desired species may be sensitive, while other species are tolerant. Such selective pressure can result in changes in species
diversity, population dynamics, and community structure that may be considered undesirable. Similarly, toxic substances may
impair the growth and development of seedlings resulting in decreased plant populations, decreased competitive abilities, reduced
reproductive capacity, and lowered crop yield. For the purposes of this guide, test substances include pesticides, industrial
chemicals, sludges, metals or metalloids, and hazardous wastes that could be added to soil. It also includes environmental samples
that may have had any of these test substances incorporated into soil.
1.4 Terrestrial plants range from annuals, capable of completing a life-cycle in as little as a few weeks, to long-lived perennials
that grow and reproduce for several hundreds of years. Procedures to evaluate chemical effects on plants range from short-term
measures of physiological responses (for example, chlorophyll fluorescence) to field studies of trees over several years. Research
and development of standardized plant tests have emphasized three categories of tests: (1) short-term, physiological endpoints (that
is, biomarkers); (2) short-term tests conducted during the early stages of plant growth with several endpoints related to survival,
growth, and development; and ( 3) life-cycle toxicity tests that emphasize reproductive success.
1.5 This guide is arranged by sections as follows:
Section Title
1 Scope
2 Referenced Documents
3 Terminology
4 Summary of Phytotoxicity Tests
5 Significance and Use
6 Apparatus
7 Test Material
8 Hazards
9 Test Organisms
10 Sample Handling and Storage
11 Calibration and Standardization
12 Test Conditions
13 Interference and Limitations
14 Quality Assurance and Quality Control
15 Calculations and Interpretation of Results
16 Precision and Bias
1.6 The values stated in SI units are to be regarded as standard. No other units of measurement are included in this standard.
1.7 This standard does not purport to address all of the safety concerns, if any, associated with its use. It is the responsibility
of the user of this standard to establish appropriate safety and health practices and determine the applicability of regulatory
limitations prior to use. Specific precautionary statements are given in Section 8.
This guide is under the jurisdiction of ASTM Committee E50 on Environmental Assessment, Risk Management and Corrective Action and is the direct responsibility
of Subcommittee E50.47 on Biological Effects and Environmental Fate.
Current edition approved March 1, 2009Oct. 1, 2014. Published March 2009December 2014. Originally published in 1998. Last previous edition published 20032009 as
E1963–03.–09. DOI: 10.1520/E1963-09.10.1520/E1963-09R14.
Copyright © ASTM International, 100 Barr Harbor Drive, PO Box C700, West Conshohocken, PA 19428-2959. United States
E1963 − 09 (2014)
2. Referenced Documents
2.1 ASTM Standards:
D1193 Specification for Reagent Water
D4547 Guide for Sampling Waste and Soils for Volatile Organic Compounds
D5633 Practice for Sampling with a Scoop
E1598 Practice for Conducting Early Seedling Growth Tests (Withdrawn 2003)
E1733 Guide for Use of Lighting in Laboratory Testing
2.2 Code of Federal Regulations Standard:
CFR 49
2.3 Other useful references have described phytotoxicity test procedures(1-11) .
3. Terminology
3.1 General Terminology—The words “must,” “should,” “may,”“ can,” and “might” have very specific meanings in this guide.
“Must” is used to express an absolute requirement, that is, to state that the test ought to be designed to satisfy the specified
condition, unless the purpose of the test requires a different design. “Must” is only used in connection with factors that directly
relate to the acceptability of the test (see Section 14). “Should” is used to state that the specified condition is recommended and
ought to be met if possible. Although violation of one “should” is rarely a serious matter, violation of several will often render the
results questionable. Terms such as “is desirable,” “is often desirable,” and “might be desirable” are used in connection with less
important factors. “May” is used to mean “is (are) allowed to,” “can” is used to mean “is (are) able to,” and “might” is used to
mean “could possibly.” Thus the classic distinction between “may” and “can” is preserved, and “might” is never used as a synonym
for either “may” or “can.”
3.2 Definitions:
3.2.1 control, n—the treatment group in a toxicity test consisting of reference soil or artificial soil that duplicates all the
conditions of the exposure treatments, but contains no test substance. The control is used to determine if there are any statistical
differences in endpoints related to the test substance.
3.2.2 eluate, n—solution obtained from washing a solid with a solvent to remove adsorbed material.
3.2.3 hazardous substance, n—a material that can cause deleterious effects to plants, microbes, or animals. (A hazardous
substance does not, in itself, present a risk unless an exposure potential exists.)
3.2.4 inhibition, n—a statistically lower value of any endpoint compared to the control values that is related to environmental
concentration or application rate.
3.2.5 leachate, n—water plus solutes that has percolated through a column of soil or waste.
3.2.6 test material, n—any formulation, dilution, etc. of a test substance.
3.2.7 test substance, n—a chemical, formulation, eluate, sludge, or other agent or substance that is the target of the investigation
in a toxicity test.
3.2.8 toxicant, n—an agent or material capable of producing an adverse response (effect) in a biological system, adversely
impacting structure or function or producing death.
3.2.9 toxicity endpoints, n—measurements of organism response such as death, growth, developmental, or physiological
parameters resulting from exposure to toxic substances.
3.3 Definitions of Terms Specific to This Standard:
3.3.1 chlorotic, adj—the discoloration of shoots that occurs as chlorophyll is degraded as a result of disease, toxic substances,
nutrient deficiencies, or senescence.
3.3.2 coleoptile, n—the protective tissues surrounding the growing shoot in a monocotyledonous plant.
3.3.3 cotyledon, n—a primary leaf of the embryo in seeds, only one in the monocotyledons, two in dicotyledons. In many of
the latter, such as the bean, they emerge above ground and appear as the first leaves.
3.3.4 cutting, n—a vegetative segment of a plant, usually a stem that contains several nodes and associated buds, that can be
used to regenerate an entire plant.
3.3.5 dead test plants, n—those individuals that expired during the test observation period as indicated by severe desiccation,
withering, chlorosis, necrosis, or other symptoms that indicate non-viability.
For referenced ASTM standards, visit the ASTM website, www.astm.org, or contact ASTM Customer Service at service@astm.org. For Annual Book of ASTM Standards
volume information, refer to the standard’s Document Summary page on the ASTM website.
The last approved version of this historical standard is referenced on www.astm.org.
Available from Standardization Documents Order Desk, DODSSP, Bldg. 4, Section D, 700 Robbins Ave., Philadelphia, PA 19111-5098, http://www.dodssp.daps.mil.
The boldface numbers in parentheses refer to the list of references at the end of this guide.
E1963 − 09 (2014)
3.3.6 desiccated, adj—the plant, or portion of the plant, that is dried in comparison to the control plant.
3.3.7 development, n—the series of steps involving cell division and cell differentiation into various tissues and organs.
3.3.8 dicotyledon, n—in the classification of plants, those having two seed leaves.
3.3.9 dormancy, n—a special condition of arrested growth in which buds, embryos, or entire plants survive at lowered metabolic
activity levels. Special environmental cues such as particular temperature regimes or photoperiods are required to activate
metabolic processes and resume growth. Seeds that require additional treatment besides adequate moisture and moderate
temperature to germinate are said to be dormant. (See quiescence.)
3.3.10 emergence, n—following germination of a plant, the early growth of a seedling that pushes the epicotyl through the soil
surface.
3.3.11 enhanced growth and yield, n—when a treated plant exhibits shoot growth, root elongation, lateral root growth, or yield
significantly greater than the control values, the plant is “enhanced” or “stimulated.”
3.3.12 epicotyl, n—that portion of an embryo or seedling containing the shoot. It is delineated anatomically by the transition
zone which separates the epicotyl from the hypocotyl.
3.3.13 fruits, n—the reproductive tissues derived from the ovary in the case of epigenous flowers or the ovary and accessory
tissues in the case of hypigenous flowers.
3.3.14 germination, n—the physiological events associated with re-initiation of embryo growth and mobilization of reserve
nutrients in seeds. The emergence of the seedling radicle from the seed coat defines the end of germination and the beginning of
early seedling growth.
3.3.15 growth, n—a change in size or mass measured by length, height, volume, or mass.
3.3.16 hypocotyl, n—that portion of an embryo or seedling containing the root or radicle. It is delineated anatomically by the
transition zone which separates the epicotyl from the hypocotyl.
3.3.17 inhibited plant growth and yield, n— plant growth, root length and lateral root growth, or yield are “inhibited” when their
measurements are significantly less than the control values.
3.3.18 lateral roots, n—roots growing off the primary roots, also referred to as secondary roots.
3.3.19 monocotyledon, n— in the classification of plants, those having a single seed leaf.
3.3.20 mottled, adj—marked with lesions, spots or streaks of different colors. This includes the discoloration of leaf margins.
3.3.21 phytotoxicity, n—a lethal or sub-lethal response of plants to a toxicant.
3.3.22 quiescence, n—a condition in buds, embryos, or entire plants characterized by lowered metabolic rates and limited or no
growth. Seeds that germinate when supplied with adequate moisture and moderate temperature are said to be quiescient. (See
dormancy.)
3.3.23 radicle, n—the emerging root of an embryo during germination.
3.3.24 seed, n—the propagule of a plant derived from an ovule. It consists of an embryo, a protective covering (seed coat), and
may have storage tissue (endosperm).
3.3.25 shoot, n—the above-ground portion of a plant consisting of stems, leaves, as well as any reproductive parts that may be
attached.
3.3.26 surviving plants, n—test plants that are alive at the time observations are recorded.
3.3.27 viable, adj—plants capable of resuming metabolic functions and growth are considered “viable.” Buds, embryos, or
entire plants may be dormant or quiescient and therefore exhibit no growth during the period of observation. Distinguishing dead
plants from viable plants with certainty is difficult without special training and sophisticated measures of metabolic function.
3.3.28 withering, v—becoming limp or desiccated, deprived of moisture; often the result of root damage.
4. Summary of Phytotoxicity Tests
4.1 The terrestrial phytotoxicity tests covered under this guide apply to a range of test conditions and test species that can be
adapted to meet project-specific objectives. Test organisms are maintained either as seeds or as cuttings until a particular test is
to be conducted. A prescribed number of individual plants are introduced into test treatments that include a negative control, a
series of positive controls, and one or more test-substance treatment concentrations. The treatment concentrations may be known
or unknown; nominal or measured, depending on the nature of the investigation. In the case where the test substance is evaluated
as an additive to soil, a range of concentrations is recommended. In tests of environmental samples that already contain a putative
phytotoxic substance, the tests may be conducted with either the test soil as collected from the field, or as diluted with a suitable
reference soil. Another variant of the tests allows for amendments, or spikes, of selected toxic substances to be added to
environmental samples. Finally, in the case of the root elongation assay, eluates, effluents, or other aqueous derivatives of a soil
sample are tested.
E1963 − 09 (2014)
4.2 Plants are exposed to the test substances in the form described in the specific variations of the tests for a discrete period of
time that ranges from 96 h to several months. For short tests, no nutrient additions or amendments are needed or recommended
as the amendments may interact with the toxicant and alter the toxicity response. For tests lasting more than two weeks, nutrient
additives may be warranted, depending on the test objectives, in order to maximize the potential for plant growth and development.
Thinning, culling, or replacing individual pl
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