ASTM D7463-14a
(Test Method)Standard Test Method for Adenosine Triphosphate (ATP) Content of Microorganisms in Fuel, Fuel/Water Mixtures, and Fuel Associated Water
Standard Test Method for Adenosine Triphosphate (ATP) Content of Microorganisms in Fuel, Fuel/Water Mixtures, and Fuel Associated Water
SIGNIFICANCE AND USE
5.1 This test method measures the concentration of ATP present in the sample. ATP is a constituent of all living cells including bacteria and fungi. Consequently, the presence of ATP is a reliable indicator of microbial contamination in fuel systems. ATP is not associated with matter of non-biological origin.
5.2 This test method differs from Test Method D4012 as follows:
5.2.1 By providing for the rapid determination of ATP present in a fuel (petroleum) sample, a fuel and water mixture sample, fuel-associated bottom water sample, and extracellular ATP freely available in the fuel or aqueous sample matrix;
5.2.2 By providing for a method to capture, extract, and quantify ATP using self-contained test device and luminometer;
5.2.3 By providing a method of quantifying ATP present in fuel or water matrices in generally less than 10 min; and
5.2.4 By providing for the rapid separation of the ATP from chemical interferences that have previously prevented the use of ATP determinations in complex fluids containing hydrocarbons and other organic molecules.
5.3 This test method does not require the use of hazardous materials and does not generate biohazard waste.
5.4 This test method can be used to estimate viable microbial biomass, to evaluate the efficacy of antimicrobial pesticides, and to monitor microbial contamination in fuel storage and distribution systems.
SCOPE
1.1 This test method provides a protocol for capturing, concentrating, and testing the adenosine triphosphate (ATP) present in a fuel system sub-sample (that is, test specimen) associated with:
1.1.1 Microorganisms and hydrophilic particles found in liquid fuels as described in Table X6.1, or
1.1.2 Microorganisms and hydrophilic particles found in mixture of fuel and associated bottom water or just associated bottom water.
1.1.3 ATP detected by this bioluminescence test can be derived from cellular ATP, extra-cellular ATP, or some combination of both.
1.1.4 Cellular and extra-cellular ATP utilized to perform ATP bioluminescence are captured and concentrated from a fuel system sample into an aqueous test specimen (that is, sub-sample) for testing. For example, for a fuel system sample that does not contain any visible fuel associated bottom water, the aqueous test specimen is the capture solution itself described in 8.2.1.1. For fuel system samples that are a mixture of fuel and associated bottom water (that is, free water), the test specimen is an aliquant of the capture solution and associated bottom water.
1.2 The ATP is measured using a patented bioluminescence enzyme assay, whereby light is generated in amounts proportional to the concentration of ATP in the sample. The light is produced and measured quantitatively using dedicated ATP test pens2 and a dedicated luminometer2 and reported in (instrument specific) Relative Light Units.
1.3 This test method is equally suitable for use in the laboratory or field.
1.4 Although bioluminescence is a reliable and proven technology, this method does not differentiate ATP from bacteria or fungi.
1.5 For water or capture solution samples, the concentration range of ATP detectable by this test method is 1 × 10–11 M to 3 × 10–8 M which is equivalent to 1 × 10–14 moles/mL to 3 × 10–11 moles/mL for water samples or capture solution. Assuming testing on fuel phase is performed on a 500 mL volume of fuel the equivalent concentrations is fuel would be: 6 × 10–11 M to 2 × 10–14 M.
1.6 The values stated in SI units are to be regarded as standard. No other units of measurement are included in this standard.
1.6.1 There is one exception—Relative Light Unit (RLU) as defined in 3.1.19.
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 limi...
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Designation: D7463 − 14a
StandardTest Method for
Adenosine Triphosphate (ATP) Content of Microorganisms
1
in Fuel, Fuel/Water Mixtures, and Fuel Associated Water
This standard is issued under the fixed designation D7463; 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* 1.3 This test method is equally suitable for use in the
laboratory or field.
1.1 This test method provides a protocol for capturing,
1.4 Although bioluminescence is a reliable and proven
concentrating, and testing the adenosine triphosphate (ATP)
technology, this method does not differentiate ATP from
present in a fuel system sub-sample (that is, test specimen)
bacteria or fungi.
associated with:
1.1.1 Microorganisms and hydrophilic particles found in
1.5 Forwaterorcapturesolutionsamples,theconcentration
–11
liquid fuels as described in Table X6.1,or
range ofATP detectable by this test method is1×10 Mto
–8 –14
1.1.2 Microorganisms and hydrophilic particles found in
3×10 M which is equivalent to1×10 moles/mL to 3 ×
–11
mixture of fuel and associated bottom water or just associated
10 moles/mLfor water samples or capture solution.Assum-
bottom water.
ing testing on fuel phase is performed on a 500 mLvolume of
–11
1.1.3 ATP detected by this bioluminescence test can be
fuel the equivalent concentrations is fuel would be:6×10
–14
derived from cellularATP, extra-cellularATP, or some combi- Mto2×10 M.
nation of both.
1.6 The values stated in SI units are to be regarded as
1.1.4 Cellular and extra-cellular ATP utilized to perform
standard. No other units of measurement are included in this
ATP bioluminescence are captured and concentrated from a
standard.
fuel system sample into an aqueous test specimen (that is,
1.6.1 Thereisoneexception—RelativeLightUnit(RLU)as
sub-sample) for testing. For example, for a fuel system sample
defined in 3.1.19.
that does not contain any visible fuel associated bottom water,
1.7 This standard does not purport to address all of the
the aqueous test specimen is the capture solution itself de-
safety concerns, if any, associated with its use. It is the
scribed in 8.2.1.1. For fuel system samples that are a mixture
responsibility of the user of this standard to establish appro-
offuelandassociatedbottomwater(thatis,freewater),thetest
priate safety and health practices and determine the applica-
specimen is an aliquant of the capture solution and associated
bility of regulatory limitations prior to use.
bottom water.
1.2 TheATPis measured using a patented bioluminescence
2. Referenced Documents
enzyme assay, whereby light is generated in amounts propor-
3
2.1 ASTM Standards:
tional to the concentration of ATP in the sample. The light is
D396Specification for Fuel Oils
producedandmeasuredquantitativelyusingdedicatedATPtest
D910Specification for Leaded Aviation Gasolines
2 2
pens and a dedicated luminometer and reported in (instru-
D975Specification for Diesel Fuel Oils
ment specific) Relative Light Units.
D1655Specification for Aviation Turbine Fuels
4
D2069Specification for Marine Fuels (Withdrawn 2003)
D2880Specification for Gas Turbine Fuel Oils
1
This test method is under the jurisdiction of ASTM Committee D02 on
D3699Specification for Kerosine
Petroleum Products, Liquid Fuels, and Lubricants and is the direct responsibility of
D4012TestMethodforAdenosineTriphosphate(ATP)Con-
Subcommittee D02.14 on Stability and Cleanliness of Liquid Fuels.
tent of Microorganisms in Water
CurrenteditionapprovedJune1,2014.PublishedJuly2014.Originallyapproved
in 2008. Last previous edition approved in 2014 as D7463–14. DOI: 10.1520/
D7463-14A.
2
The sole source of supply, repair, recertification, and technical support of the
3
apparatus or test pen known to the committee at this time is Merck KGaA, 64271 For referenced ASTM standards, visit the ASTM website, www.astm.org, or
Darmstadt,Germany(Worldwide)orFuelQualityServices,Inc.,4584CantrellRd., contact ASTM Customer Service at service@astm.org. For Annual Book of ASTM
FloweryBranch,GA30542(USA).Ifyouareawareofalternativesuppliers,please Standards volume information, refer to the standard’s Document Summary page on
provide this information toASTM International Headquarters.Your comments will the ASTM website.
1 4
receive careful consideration at a meeting of the responsible technical committee, The last approved version of this historical standard is referenced on
which you may attend. www.astm.org.
*A Summary of Changes section appears at the end of this standard
Copyright © ASTM International, 100 Barr Harbor Drive, PO Box C700, West Conshohocken, PA 19428-2959. United States
1
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...
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: D7463 − 14 D7463 − 14a
Standard Test Method for
Adenosine Triphosphate (ATP) Content of Microorganisms
1
in Fuel, Fuel/Water Mixtures, and Fuel Associated Water
This standard is issued under the fixed designation D7463; 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 test method provides a protocol for capturing, extractingconcentrating, and quantifyingtesting the adenosine
triphosphate (ATP) content present in a fuel system sub-sample (that is, test specimen) associated with:
1.1.1 Microorganisms and hydrophilic particles found in conventional liquid fuels with kinematic viscosities (at 40°C) of ≤ 8
2 –1
mmliquid fuels · s as described in Table X6.1, or
1.1.2 Microorganisms found in fuel-associated bottom water, andand hydrophilic particles found in mixture of fuel and
associated bottom water or just associated bottom water.
1.1.3 ATP detected by this bioluminescence test can be derived from cellular ATP, extra-cellular ATP, or some combination of
both.
1.1.4 Extracellular (non-cellular) ATP present Cellular and extra-cellular ATP utilized to perform ATP bioluminescence are
captured and concentrated from a fuel system sample into an aqueous test specimen (that is, sub-sample) for testing. For example,
for a fuel system sample that does not contain any visible fuel associated bottom water, the aqueous test specimen is the capture
solution itself described in 8.2.1.1the sample matrix. For fuel system samples that are a mixture of fuel and associated bottom
water (that is, free water), the test specimen is an aliquant of the capture solution and associated bottom water.
1.2 The ATP is measured using a patented bioluminescence enzyme assay, whereby light is generated in amounts proportional
2
to the concentration of ATP in the sample. The light is produced and measured quantitatively using dedicated ATP test pens and
2
a dedicated luminometer and reported in (instrument specific) Relative Light Units.
1.3 This test method is equally suitable for use in the laboratory or field.
1.4 Although bioluminescence is a reliable and proven method for qualifying and quantifying ATP, technology, this method does
not differentiate between ATP from different sources, for example, from different types of microorganism such as ATP from
bacteria or fungi.
–11
1.5 For water or capture solution samples, the concentration range of ATP detectable by this test method is 1 × 10 M to 3
–8 –14 –11
× 10 M which is equivalent to 1 × 10 moles/mL to 3 × 10 moles/mL for water samples or capture solution. Assuming testing
–11 –14
on fuel phase is performed on a 500 mL volume of fuel the equivalent concentrations is fuel would be: 6 × 10 M to 2 × 10
M.
1.6 The values stated in SI units are to be regarded as standard. No other units of measurement are included in this standard.
1.6.1 There is one exception—Relative Light Unit (RLU) as defined in 3.1.19.
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.
1
This test method is under the jurisdiction of ASTM Committee D02 on Petroleum Products, Liquid Fuels, and Lubricants and is the direct responsibility of Subcommittee
D02.14 on Stability and Cleanliness of Liquid Fuels.
Current edition approved May 1, 2014June 1, 2014. Published May 2014July 2014. Originally approved in 2008. Last previous edition approved in 20082014 as
D7463 – 08.D7463 – 14. DOI: 10.1520/D7463-14.10.1520/D7463-14A.
2
The sole source of supply supply, repair, recertification, and technical support of the apparatus or test pen known to the committee at this time is Merck KGaA, 64271
Darmstadt, Germany. Germany (Worldwide) or Fuel Quality Services, Inc., 4584 Cantrell Rd., Flowery Branch, GA 30542 (USA). If you are aware of alternative suppliers,
1
please provide this information to ASTM International Headquarters. Your comments will receive careful consideration at a meeting of the responsible technical committee,
which you may attend.
*A Summary of Changes section appears at the end of this standard
Copyright © ASTM International, 100 Barr Harbor Drive, PO Box C700, West Conshohocken
...
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