ASTM D3731-20

Standard Practices for Measurement of Chlorophyll Content of Algae in Surface Waters

SIGNIFICANCE AND USE
5.1 Data on the chlorophyll content of the algae have the following applications:  
5.1.1 To provide estimates of algal biomass and productivity.  
5.1.2 To provide general information on the taxonomic composition (major groups) of the algae, based on the relative amounts of chlorophyll a, b, and c, and the physiological condition of algal communities, which is related to the relative abundance of pheopigments.  
5.1.3 To determine long-term trends in water quality.  
5.1.4 To determine the trophic status of surface waters.  
5.1.5 To detect adverse effects of pollutants on plankton and periphyton in receiving waters.  
5.1.6 To determine maximum growth rates and yields in algal growth potential tests.
SCOPE
1.1 These practices include the extraction and the measurement of chlorophyll a, b, and c, and pheophytin a in freshwater and marine plankton and periphyton. Three practices are provided as follows:  
1.1.1 Spectrophotometric, trichromatic practice for measuring chlorophyll a, b, and c.  
1.1.2 Spectrophotometric, monochromatic practice for measuring chlorophyll a corrected for pheophytin a; and for measuring pheophytin a.  
1.1.3 Fluorometric practice for measuring chlorophyll a corrected for pheophytin a; and for measuring pheophytin a.  
1.2 The values stated in SI units are to be regarded as standard. No other units of measurement are included in this standard.  
1.3 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, health, and environmental practices and determine the applicability of regulatory limitations prior to use. Specific precautionary statements are given in Section 8.  
1.4 This international standard was developed in accordance with internationally recognized principles on standardization established in the Decision on Principles for the Development of International Standards, Guides and Recommendations issued by the World Trade Organization Technical Barriers to Trade (TBT) Committee.

General Information

• Status: Published
• Publication Date: 31-May-2020
• ICS: 13.060.10 - Water of natural resources
• Technical Committee: D19 - Water
• Drafting Committee: D19.24 - Water Microbiology

Relations

• Referred By: ASTM E1218-21 - Standard Guide for Conducting Static Toxicity Tests with Microalgae
• Effective Date: 01-Jun-2020

Standards Content (Sample)

This international standard was developed in accordance with internationally recognized principles on standardization established in the Decision on Principles for the
Development of International Standards, Guides and Recommendations issued by the World Trade Organization Technical Barriers to Trade (TBT) Committee.
Designation: D3731 − 20
Standard Practices for Measurement of
1
Chlorophyll Content of Algae in Surface Waters
This standard is issued under the fixed designation D3731; 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 3.1.1 For definitions of terms used in this standard, refer to
Terminology D1129.
1.1 These practices include the extraction and the measure-
mentofchlorophyll a, b,and c,andpheophytin ainfreshwater 3.2 Definitions of Terms Specific to This Standard:
and marine plankton and periphyton. Three practices are 3.2.1 chlorophyll, n—a class of pigments involved in pho-
provided as follows: tosynthesis.
1.1.1 Spectrophotometric, trichromatic practice for measur-
3.2.2 periphyton, n—microorganisms growing on sub-
ing chlorophyll a, b, and c.
merged objects, commonly consisting of bacteria, algae,
1.1.2 Spectrophotometric,monochromaticpracticeformea-
protozoa, and rotifers.
suring chlorophyll a corrected for pheophytin a; and for
3.2.3 pheopigments (phaeopigments), n—a non-
measuring pheophytin a.
photosynthetic pigment which is the degradation product of
1.1.3 Fluorometric practice for measuring chlorophyll a
algal chlorophyll pigments.
corrected for pheophytin a; and for measuring pheophytin a.
3.2.4 plankton, n—nonmotile or weakly swimming
1.2 The values stated in SI units are to be regarded as
organisms, usually microscopic, that drift or are carried along
standard. No other units of measurement are included in this
bycurrentsinsurfacewaters,commonlyconsistingofbacteria,
standard.
algae, protozoa, rotifers, and microcrustacea.
1.3 This standard does not purport to address all of the
4. Summary of Practices
safety concerns, if any, associated with its use. It is the
responsibility of the user of this standard to establish appro-
4.1 The chlorophyll and related compounds are extracted
priate safety, health, and environmental practices and deter-
from the algae with 90% aqueous acetone. The concentration
mine the applicability of regulatory limitations prior to use.
of the pigments is determined by measuring the light absorp-
Specific precautionary statements are given in Section 8.
tion or fluorescence of the extract.
1.4 This international standard was developed in accor-
dance with internationally recognized principles on standard- 5. Significance and Use
ization established in the Decision on Principles for the
5.1 Data on the chlorophyll content of the algae have the
Development of International Standards, Guides and Recom-
following applications:
mendations issued by the World Trade Organization Technical
5.1.1 To provide estimates of algal biomass and productiv-
Barriers to Trade (TBT) Committee.
ity.
5.1.2 To provide general information on the taxonomic
2. Referenced Documents
composition (major groups) of the algae, based on the relative
2
2.1 ASTM Standards:
amounts of chlorophyll a, b, and c, and the physiological
D1129Terminology Relating to Water
condition of algal communities, which is related to the relative
D1193Specification for Reagent Water
abundance of pheopigments.
5.1.3 To determine long-term trends in water quality.
3. Terminology
5.1.4 To determine the trophic status of surface waters.
3.1 Definitions:
5.1.5 Todetectadverseeffectsofpollutantsonplanktonand
periphyton in receiving waters.
1
These practices are under the jurisdiction ofASTM Committee D19 on Water
5.1.6 To determine maximum growth rates and yields in
and are the direct responsibility of Subcommittee D19.24 on Water Microbiology.
algal growth potential tests.
Current edition approved June 1, 2020. Published June 2020. Originally
approved in 1979. Last previous edition approved in 2012 as D3731–87 (2012).
6. Interferences and Special Considerations
DOI: 10.1520/D3731-20.
2
For referenced ASTM standards, visit the ASTM website, www.astm.org, or
6.1 Pigment Extraction—The chlorophylls are only poorly
contact ASTM Customer Service at [e-mail]. For Annual Book of ASTM
extracted, if at all, from some forms of algae, such as the
Standards volume information, refer to the standard’s Document Summary page on
the ASTM website. coccoid green algae, unless the cells are disrupted, whereas
Copyright © ASTM International, 100 Barr Harbor Drive, PO Box C700, West Conshohocken, PA 19428-2959. United States
1

---------------------- Page: 1 ----------------------
D3731 − 20
other algae, such as the diatoms, give up their pigments very 6.6 Spectrophotometer Resol
...

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: D3731 − 87 (Reapproved 2012) D3731 − 20
Standard Practices for Measurement of
1
Chlorophyll Content of Algae in Surface Waters
This standard is issued under the fixed designation D3731; 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 These practices include the extraction and the measurement of chlorophyll a, b, and c, and pheophytin a in freshwater and
marine plankton and periphyton. Three practices are provided as follows:
1.1.1 Spectrophotometric, trichromatic practice for measuring chlorophyll a, b, and c.
1.1.2 Spectrophotometric, monochromatic practice for measuring chlorophyll a corrected for pheophytin a; and for measuring
pheophytin a.
1.1.3 Fluorometric practice for measuring chlorophyll a corrected for pheophytin a; and for measuring pheophytin a.
1.2 The values stated in SI units are to be regarded as the standard. The values given in parentheses are provided for information
purposes only.standard. No other units of measurement are included in this standard.
1.3 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, health, and environmental practices and determine the applicability of
regulatory limitations prior to use. Specific precautionary statements are given in Section 78.
1.4 This international standard was developed in accordance with internationally recognized principles on standardization
established in the Decision on Principles for the Development of International Standards, Guides and Recommendations issued
by the World Trade Organization Technical Barriers to Trade (TBT) Committee.
2. Referenced Documents
2
2.1 ASTM Standards:
D1129 Terminology Relating to Water
D1193 Specification for Reagent Water
3. Terminology
3.1 Definitions:
3.1.1 For definitions of terms used in this standard, refer to Terminology D1129.
3.2 Definitions:Definitions of Terms Specific to This Standard:
3.2.1 chlorophyll, n—a class of pigments involved in photosynthesis.
3.2.2 periphyton, n—microorganisms growing on submerged objects, commonly consisting of bacteria, algae, protozoa, and
rotifers.
3.2.3 pheopigments (phaeopigments), n—a non-photosynthetic pigment which is the degradation product of algal chlorophyll
pigments.
3.2.4 plankton—plankton, n—nonmotile or weakly swimming organisms, usually microscopic, that drift or are carried along by
currents in surface waters, commonly consisting of bacteria, algae, protozoa, rotifers, and microcrustacea.
2.1.2 periphyton—microorganisms growing on submerged objects, commonly consisting of bacteria, algae, protozoa, and
rotifers.
4. Summary of Practices
4.1 The chlorophyll and related compounds are extracted from the algae with 90 % aqueous acetone. The concentration of the
pigments is determined by measuring the light absorption or fluorescence of the extract.
1
These practices are under the jurisdiction of ASTM Committee D19 on Water and are the direct responsibility of Subcommittee D19.24 on Water Microbiology.
Current edition approved Sept. 1, 2012June 1, 2020. Published October 2012June 2020. Originally approved in 1979. Last previous edition approved in 20042012 as
D3731 – 87 (2004).(2012). DOI: 10.1520/D3731-87R12.10.1520/D3731-20.
2
For referenced ASTM standards, visit the ASTM website, www.astm.org, or contact ASTM Customer Service at [e-mail]. For Annual Book of ASTM Standards
volume information, refer to the standard’s Document Summary page on the ASTM website.
Copyright © ASTM International, 100 Barr Harbor Drive, PO Box C700, West Conshohocken, PA 19428-2959. United States
1

---------------------- Page: 1 ----------------------
D3731 − 20
5. Significance and Use
5.1 Data on the chlorophyll content of the algae have the following applications:
5.1.1 To provide estimates of algal biomass and productivity.
5.1.2 To provide general information on the taxonomic composition (major groups) of the algae, based on the relative amounts
of chlorophyll a, b, and c, and the physiological condition of algal communities, which is related to the relative abundance of
pheopigments.
5.1.3 To determine long-term trends in water quality.
5.1.4 To determine the trophic status of surface waters.
5.1.5 To detect adverse effects of
...