ASTM D4412-84(1997)
(Test Method)Standard Test Methods for Sulfate-Reducing Bacteria in Water and Water-Formed Deposits
Standard Test Methods for Sulfate-Reducing Bacteria in Water and Water-Formed Deposits
SCOPE
1.1 These test methods cover the procedure for the detection and enumeration by the most probable number (MPN) technique of sulfate-reducing bacteria in water or water-formed deposits.
1.2 Two media preparations are provided. Medium A which is prepared with reagent grade water, and Medium B which is prepared using the water to be sampled as the water source. Medium B is offered for those special conditions where sulfate-reducing bacterial strains have adapted to atypical non-fresh water environment.
1.3 For the isolation and enumeration of thermophilic sulfate-reducing bacteria encountered in waters associated with oil and gas production, all broths, dilution blanks, and incubations must be maintained at tempertures of at least 45°C and preferably within 5° at the sample temperature.
1.4 The sensitivity of these test methods can be increased by purging the dilution blanks and tubes of media with nitrogen immediately prior to use.
1.5 The analyst should be aware that adequate collaborative data for precision and bias statements as required by Practice D2777 are not provided. See Section 11 for details.1.6 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.
General Information
Relations
Standards Content (Sample)
Designation: D 4412 – 84 (Reapproved 1997)
AMERICAN SOCIETY FOR TESTING AND MATERIALS
100 Barr Harbor Dr., West Conshohocken, PA 19428
Reprinted from the Annual Book of ASTM Standards. Copyright ASTM
Standard Test Methods for
Sulfate-Reducing Bacteria in Water and Water-Formed
Deposits
This standard is issued under the fixed designation D 4412; 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 (e) indicates an editorial change since the last revision or reapproval.
1. Scope water, Fifteenth Edition
1.1 These test methods cover the procedure for the detection
3. Terminology
and enumeration by the most probable number (MPN) tech-
3.1 Definitions—For definitions of terms used in these test
nique of sulfate-reducing bacteria in water or water-formed
methods, refer to Terminology D 1129.
deposits.
3.2 Definitions of Terms Specific to This Standard: —For a
1.2 Two media preparations are provided. Medium A which
description of the term MPN used in these test methods, refer
is prepared with reagent grade water, and Medium B which is
to literature.
prepared using the water to be sampled as the water source.
Medium B is offered for those special conditions where
4. Summary of Test Methods
sulfate-reducing bacterial strains have adapted to atypical
4.1 Water and water deposit samples and dilutions of these
non-fresh water environment.
samples are dispensed into tubes of Starkey’s medium (A or B)
1.3 For the isolation and enumeration of thermophilic
following five tube MPN procedures. The tubes are sealed with
sulfate-reducing bacteria encountered in waters associated with
liquid paraffin, and incubated at 20°C for 21 days. Positive
oil and gas production, all broths, dilution blanks, and incuba-
reactions are indicated by the deposit of a black precipitate.
tions must be maintained at temperatures of at least 45°C and
preferably within 5°C at the sample temperature.
5. Significance and Use
1.4 The sensitivity of these test methods can be increased by
5.1 Sulfate-reducing bacteria are widely distributed in ma-
purging the dilution blanks and tubes of media with nitrogen
rine and fresh water muds which, in consequence, frequently
immediately prior to use.
are laden with the hydrogen sulfide produced by these organ-
1.5 The analyst should be aware that adequate collaborative
isms during dissimilatory sulfate reduction.
data for precision and bias statements as required by Practice
5.2 It has been reported that Desulfovibrio can form as
D 2777 are not provided. See Section 11 for details.
much as 10 g of sulfide per litre during active multiplication.
1.6 This standard does not purport to address all of the
Sulfate-reducing bacteria can cause the external or internal
safety concerns, if any, associated with its use. It is the
corrosion of water or wastewater pipelines and pipelines for
responsibility of the user of this standard to establish appro-
petroleum and natural gas. The formation of galvanic cells by
priate safety and health practices and determine the applica-
massive growth of sulfate-reducing bacteria under suitable
bility of regulatory limitations prior to use.
conditions makes the corrosion much worse than just the effect
of the hydrogen sulfide on the metal or concrete.
2. Referenced Documents
2.1 ASTM Standards:
6. Apparatus and Materials
D 1129 Terminology Relating to Water
6.1 Anaerobic Incubator, 20°C, if available, or conventional
D 1193 Specification for Reagent Water
20°C incubator.
D 2777 Practice for Determination of Precision and Bias of
6.2 Pipets, sterile, 1 mL and 10 mL,“ calibrated” to deliver.
Applicable Methods of Committee D-19 on Water
6.3 Test Tubes, with close fitting or airtight caps; 16 by 150
D 3370 Practices for Sampling Water from Closed Con-
mm and 20 by 150 mm.
duits
6.4 Test Tube Racks, of sufficient size to contain 16 and
2.2 APHA Standard:
20-mm tubes.
Standard Methods for the Examination of Water and Waste-
1 3
These test methods are under the jurisdiction of ASTM Committee D-19 on Available from American Public Health Association, 1015 18th St. N.W.,
Water and are the direct responsibility of Subcommittee D19.24 on Water Micro- Washington, DC 20036.
biology. Bonde, G. J., “Bacterial Indicators of Water Pollution,” A Study of Quantitative
Current edition approved Oct. 26, 1984. Published February 1985. Estimation, Teknisk Forlag, Copenhagen, 1963.
2 5
Annual Book of ASTM Standards, Vol 11.01. For thermophilic organisms use a 45°C incubator.
D 4412
7. Reagents
p-Aminodimethylaniline dihydrochloride 1.0 g
(C H N ·2HCl)
8 12 2
7.1 Purity of Reagents—Reagent grade chemicals shall be
HCl (6 N) 500 mL
used in all tests. Unless otherwise indicated, it is intended that
Dissolve1gof p-aminodimethylaniline dihydrochloride in
all reagents conform to the specifications of the Committee on
500 mL of 6 N HCl. Store for up to 1 month in an amber
Analytical Reagents of the American Chemical Society, when
airtight container.
such specifications are available.
7.6 Liquid Paraffın—Heavy, sterile, or sterile mineral oil.
7.2 Purity of Water—Unless otherwise indicated, references
7.7 Buffered Dilution Water—Stock Solution
to water shall be understood to mean Reagent Water Type II
7.7.1 Dissolve 34.0 g of KH PO in 500 mL of water, adjust
2 4
conforming to Specification D 1193. In addition, reagent water
pH to 7.2 with 1 N NaOH and dilute to 1 L with distilled water.
used for these test methods must be sterile.
This is called the stock phosphate solution.
7.3 Starkey’s Medium A— (modified):
7.7.2 Dissolve 38 g of MgCL in 1 L of distilled water.
Sodium lactate (C H NaO ) 3.5 g
3 5 3
7.8 Buffered Dilution Water, Working Solution—Add 1.25
Ammonium chloride (NH Cl) 1.0 g
mL of stock buffered dilution water and 5 mL of MgCl
Dipotassium, hydrogen orthophosphate 0.5 g
(K HPO ) solution to 500 mL of water. Bring to 1 L with water. Mix well
2 4
Magnesium sulfate (MgSO ·7H O) 2.0 g
4 2
and dispense as 90 mL dilution blanks in screw-capped bottles.
Sodium sulfate (Na SO ) 0.5 g
2 4
Sterilize by autoclaving at 121°C for 15 min.
Calcium chloride (CaCl ·2H O) 0.1 g
2 2
Thioglycollic ac
...
Questions, Comments and Discussion
Ask us and Technical Secretary will try to provide an answer. You can facilitate discussion about the standard in here.