14220 Federal Register / Vol. 72, No. 57 / Monday, March 26, 2007 / Rules and Regulations 

ENVIRONMENTAL PROTECTION 

AGENCY 

40 CFR Parts 136 and 503 

[EPA–HQ–OW–2004–0014; FRL–8228–1] 

RIN 2040–AE68 

Guidelines Establishing Test 

Procedures for the Analysis of 

Pollutants; Analytical Methods for 

Biological Pollutants in Wastewater 

and Sewage Sludge: Final Rule 

AGENCY: Environmental Protection 

Agency (EPA). 

ACTION: Final rule. 

SUMMARY: This rule modifies the EPA’s 

Guidelines that establish approved 

bacterial testing procedures for analysis 

and sampling under the Clean Water 

Act. EPA proposed these changes for 

public comment on August 16, 2005 and 

April 10, 2006. These changes include 

approval for new methods for 

monitoring microbial pollutants in 

wastewater and sewage sludge, 

including EPA methods, vendordeveloped methods and methods 

developed by voluntary consensus 

bodies (VCSB) as well as updated 

versions of currently approved methods. 

The addition of new and updated 

methods to the wastewater regulations 

provides increased flexibility to the 

regulated community and laboratories 

in the selection of analytical methods. 

In addition, EPA has made a technical, 

non-substantive correction. 

DATES: This regulation is effective April 

25, 2007. The incorporation by reference 

of these methods is approved by the 

Director of the Federal Register on April 

25, 2007. For judicial review purposes, 

this final rule is promulgated as of 1 

p.m. (Eastern time) on April 9, 2007 as 

provided at 40 CFR 23.2 and 23.7. 

ADDRESSES: EPA has established a 

docket for this action under Docket ID 

No. EPA–OW–2004–0014. All 

documents in the docket are listed on 

the www.regulations.gov Web site. 

Although listed in the index, some 

information is not publicly available, 

e.g., CBI or other information whose 

disclosure is restricted by statute. 

Certain other material, such as 

copyrighted material, is not placed on 

the Internet and will be publicly 

available only in hard copy form. 

Publicly available docket materials are 

available either electronically through 

www.regulations.gov or in hard copy at 

the HQ Water Docket Center, EPA/DC, 

EPA West, Room B102, 1301 

Constitution Ave., NW., Washington, 

DC. The Public Reading Room is open 

from 8:30 a.m. to 4:30 p.m., Monday 

through Friday, excluding legal 

holidays. The telephone number for the 

Public Reading Room is (202) 566–1744, 

and the telephone number is (202) 566– 

2426. 

Note: The EPA Docket Center suffered 

damage due to flooding during the last week 

of June 2006. The Docket Center is 

continuing to operate. However, during the 

cleanup, there will be temporary changes to 

Docket Center telephone numbers, addresses, 

and hours of operation for people who wish 

to visit the Public Reading Room to view 

documents. Consult EPA’s Federal Register 

notice at 71 FR 38147 (July 5, 2006) or the 

EPA website at http://www.epa.gov/ 

epahome/dockets.htm for current 

information on docket status, locations and 

telephone numbers. 

FOR FURTHER INFORMATION CONTACT: For 

information regarding the changes to 

wastewater regulations, contact Robin 

K. Oshiro, Engineering and Analysis 

Division (4303T), USEPA Office of 

Science and Technology, 1200 

Pennsylvania Ave., NW., Washington, 

DC 20460, 202–566–1075 (e-mail: 

oshiro.robin@epa.gov). 

SUPPLEMENTARY INFORMATION: 

A. Potentially Regulated Entities 

1. Clean Water Act 

EPA Regions, as well as States, 

Territories and Tribes authorized to 

implement the National Pollutant 

Discharge Elimination System (NPDES) 

program, issue permits with conditions 

designed to ensure compliance with the 

technology-based and water qualitybased requirements of the Clean Water 

Act (CWA). These permits may include 

restrictions on the quantity of pollutants 

that may be discharged as well as 

pollutant measurement and reporting 

requirements. If EPA has approved test 

procedures for analysis of a specific 

pollutant, an NPDES permittee (or 

applicant for an NPDES permit) must 

use an approved test procedure (or an 

approved alternate test procedure) for 

the specific pollutant when testing for 

the required waste constituent. 

Similarly, if EPA has established permit 

monitoring requirements, measurements 

taken and reported under an NPDES 

permit must comply with these 

requirements. Therefore, entities with 

NPDES permits will potentially be 

regulated by the actions in this 

rulemaking. Categories and entities that 

may potentially be subject to the 

requirements of today’s rule include: 

Category Examples of potentially regulated entities 

State, Territorial, and Indian Tribal Governments .................... States, Territories, and Tribes authorized to administer the NPDES permitting 

program; States, Territories, and Tribes providing certification under Clean 

Water Act section 401. 

Industry ..................................................................................... Facilities that must conduct monitoring to comply with NPDES permits. 

Municipalities ............................................................................ POTWs that must conduct monitoring to comply with NPDES permits. 

This table is not intended to be 

exhaustive, but rather provides a guide 

for readers regarding entities likely to be 

regulated by this action. This table lists 

types of entities that EPA is now aware 

could potentially be regulated by this 

action. Other types of entities not listed 

in the table could also be regulated. To 

determine whether your facility is 

regulated by this action, you should 

carefully examine the applicability 

language at 40 CFR 122.1 (NPDES 

purpose and scope), 40 CFR 136.1 

(NPDES permits and CWA), 40 CFR 

403.1 (Pretreatment standards purpose 

and applicability). If you have questions 

regarding the applicability of this action 

to a particular entity, consult the 

appropriate person listed in the 

preceding FOR FURTHER INFORMATION

CONTACT section. 

What process governs judicial review of 

this rule? 

Under Section 509(b)(1) of the Clean 

Water Act (CWA), judicial review of 

today’s CWA rule may be obtained by 

filing a petition for review in the United 

States Circuit Court of Appeals within 

120 days from the date of promulgation 

of this rule. For judicial review 

purposes, this final rule is promulgated 

as of 1 p.m. (Eastern time) on April 25, 

2007 as provided at 40 CFR 23.2. The 

requirements of this regulation may also 

not be challenged later in civil or 

criminal proceedings brought by EPA. 

Abbreviations and Acronyms Used in 

the Preamble and Final Rule 

AOAC: Association of Official Analytical 

Chemists International 

ASTM: American Society for Testing and 

Materials International 


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Federal Register / Vol. 72, No. 57 / Monday, March 26, 2007 / Rules and Regulations 14221 

CWA: Clean Water Act 

EPA: Environmental Protection Agency 

VCSB: Voluntary Consensus Standard Body 

Table of Contents 

I. Statutory Authority 

Clean Water Act 

II. Summary of Final Rule 

A. 40 CFR Part 136 

B. 40 CFR Part 503 

III. Changes Between the Proposed Rule and 

the Final Rule 

A. Revision to 40 CFR Part 136, 

Applicability 

B. Revision to 40 CFR Part 136, 

Identification of Test Procedures 

C. Revision to 40 CFR Part 136, Table IA 

Title 

D. Revisions to 40 CFR Part 136, Table II 

and Footnotes 

E. Revisions to 40 CFR Part 503, Sampling 

and Analysis 

IV. Response to Comments 

Lack of Connecting Language Between 40 

CFR Parts 136 and 503 for Sewage 

Sludge Methods 

V. Statutory and Executive Order Reviews 

A. Executive Order 12866: Regulatory 

Planning and Review 

B. Paperwork Reduction Act 

C. Regulatory Flexibility Act 

D. Unfunded Mandates Reform Act 

E. Executive Order 13132: Federalism 

F. Executive Order 13175: Consultation 

and Coordination with Indian Tribal 

Governments 

G. Executive Order 13045: Protection of 

Children from Environmental Health 

Risks and Safety Risks 

H. Executive Order 13211: Actions that 

Significantly Affect Energy Supply, 

Distribution, or Use 

I. National Technology Transfer and 

Advancement Act 

J. Congressional Review Act 

I. Statutory Authority 

Clean Water Act 

EPA is promulgating today’s rule 

pursuant to the authority of sections 

301(a), 304(h), and 501(a) of the Clean 

Water Act (‘‘CWA’’ or the ‘‘Act’’), 33 

U.S.C. 1311(a), 1314(h), 1361(a). Section 

301(a) of the Act prohibits the discharge 

of any pollutant into navigable waters 

unless the discharge complies with a 

National Pollutant Discharge 

Elimination System (NPDES) permit 

issued under section 402 of the Act. 

Section 304(h) of the Act requires the 

Administrator of the EPA to ‘‘* * * 

promulgate guidelines establishing test 

procedures for the analysis of pollutants 

that shall include the factors which 

must be provided in any certification 

pursuant to [section 401 of this Act] or 

permit application pursuant to [section 

402 of this Act].’’ Section 501(a) of the 

Act authorizes the Administrator to 

‘‘* * * prescribe such regulations as are 

necessary to carry out this function 

under [the Act].’’ EPA generally has 

codified its test procedure regulations 

(including analysis and sampling 

requirements) for CWA programs at 40 

CFR Part 136, though some 

requirements are codified in other Parts 

(e.g., 40 CFR Chapter I, Subchapters N 

and O). 

II. Summary of Final Rule 

The following sections describe the 

changes EPA is making in today’s final 

rule. 

A. 40 CFR Part 136 

This rule approves new and revised 

methods for inclusion in 40 CFR Part 

136. These methods include EPA 

methods, vendor methods submitted by 

IDEXX and Hach, and voluntary 

consensus standards. 

The following discussion briefly 

describes the changes to Part 136 

methods approved today. 

1. This rule amends the regulations at 

40 CFR Part 136 to approve five E. coli 

and two enterococci methods for 

monitoring microbial pollutants in 

wastewaters. The E. coli methods 

include EPA Method 1603 (modified 

mTEC), and vendor methods Colilert

and Colilert-18, and mColiBlue24. 

The enterococci methods include EPA 

Method 1600 (mEI), and vendor method 

EnterolertTM. 

2. The rule approves two fecal 

coliform and one Salmonella method for 

monitoring microbial pollutants in 

sewage sludge (biosolids). The fecal 

coliform methods include EPA Methods 

1680 (LT–EC) and 1681 (A–1) and the 

Salmonella Method 1682 (Modified 

MSRV). The methods approved today 

are alternative methods to those 

currently prescribed for measuring fecal 

coliform and salmonella in sewage 

sludge identified in 40 CFR § 503.8(b). 

3. The rule amends the regulations by 

moving the microbial methods approved 

for use in ambient waters from Table IA 

to a new Table IH, and adding Table IH 

to section 136.3(a). 

4. The rule extends the holding time 

for fecal coliforms using EPA Methods 

1680 (LTB–EC) or 1681 (A–1) in sewage 

sludge for Class A composted, Class B 

aerobically or anaerobically digested 

sewage sludge. 

5. The rule amends 40 CFR 136.1 to 

add a new provision that authorizes the 

use of the methods identified at 40 CFR 

503.8(b) and the newly approved Part 

136 methods for fecal coliform and 

Salmonella for permit applications and 

recordkeeping and reporting required 

under EPA’s sewage sludge regulations 

at 40 CFR Part 503. 

B. 40 CFR Part 503 

This rule amends the regulations at 40 

CFR Part 503 by adding a cross 

reference to the 40 CFR Part 136 

methods in section 503.8(b). 

III. Changes Between the Proposed Rule 

and the Final Rule 

Except as noted below, the content of 

the final rule is the same as that of the 

proposed rule. In some instances, EPA 

revised for clarity the language of the 

final rule from that in the proposed rule. 

A. Revision to 40 CFR Part 136, 

Applicability 

Based on comment received on the 

Agency’s proposal of methods for use in 

sewage sludge, EPA has amended the 

applicability provision to clarify that the 

applicable procedures of Part 136 and 

Part 503 must be used for measurements 

for sewage sludge permit applications 

and reporting and recordkeeping 

requirements under Part 503. 

B. Revision to 40 CFR Part 136, 

Identification of Test Procedures 

Section 553 of the Administrative 

Procedure Act, 5 U.S.C. 553(b)(B), 

provides that, when an agency for good 

cause finds that notice and public 

procedure are impracticable, 

unnecessary or contrary to the public 

interest, the agency may issue a rule 

without providing notice and an 

opportunity for public comment. EPA 

has determined that there is good cause 

for making today’s changes to the rule 

final without prior proposal and 

opportunity for comment. Notice and 

opportunity for public comment is not 

necessary with respect to these changes 

because they are not substantive and 

merely correct errors in cross-referenced 

provisions as explained below. 

Section 136.3(a) provides that 

discharge parameter values for which 

reports are required must be determined 

either by the standard analytical test 

procedures described in the tables in 

Part 136 or approved additional or 

alternate test procedures. EPA has 

modified the language of 40 CFR 

136.3(a) to make three corrections. First, 

EPA has changed the citation in the last 

sentence before Table IA from 

‘‘paragraphs (b) or (c) of this section or 

40 CFR 401.13’’ to ‘‘paragraphs (c) of 

this section, 40 CFR 136.5(a)–(d) and 40 

CFR 401.13.’’ Paragraph (b) does not 

describe circumstances in which 

alternate procedures may be approved 

while section 136.5 does. 

Second, EPA has deleted the clause at 

the end of the last sentence which states 

that other test procedures may be used 

‘‘* * * when such other test procedures 

have been previously approved by the 

Regional Administrator of the Region in 

which the discharge will occur, and 

providing the Director of the State in which 


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14222 Federal Register / Vol. 72, No. 57 / Monday, March 26, 2007 / Rules and Regulations 

the discharge will occur does not object to 

the use of such alternate test procedure 

* * *.’’

Only two of the cited provisions require 

approval by the Regional Administrator 

or Director of a State. 40 CFR 401.13 

does not because it pertains to variances 

of guidelines of national applicability. 

The cross-referenced provisions 

authorize the use of additional or 

alternate test procedures in described 

circumstances. Thus, section 136.3(c) 

authorizes approval by the Regional 

Administrator (or Director of an 

approved State NPDES Program) for 

analysis of additional pollutants or 

parameters required to be reported for a 

particular discharge. Section 136.5(a)–

(d) authorizes approval by the Regional 

Administrator of alternate procedures 

for use within a particular EPA Region. 

40 CFR section 401.13 authorizes the 

use of analytical procedures that are 

specifically defined in 40 CFR Parts 

402–699. This last category of analytical 

procedures that are promulgated for 

specific effluent limitations guidelines 

and pretreatment standards and not 

codified in Part 136 do not require the 

approval of the Director of a State as the 

current language erroneously implies. 

Third, EPA removed an erroneous 

reference that was listed as a source for 

the methods listed in section 136.3. 

EPA has modified the regulation to 

provide the correct citation and delete 

the inaccurate and misleading language. 

None of the changes EPA is 

promulgating today are themselves 

substantive but rather, as noted, only 

either correct an error in citing to the 

other applicable provisions of these 

regulations or correct inaccuracies. The 

substantive provisions in question were 

previously subject to notice and 

comment. Thus, notice and public 

procedures are unnecessary. EPA finds 

that this constitutes good cause under 5 

U.S.C. 553(b)(B). 

C. Revision to 40 CFR Part 136, Table 

IA Title 

The rule revises the title to Table IA 

from ‘‘List of Approved Biological 

Methods’’ to ‘‘List of Approved 

Biological Methods for Wastewater and 

Sewage Sludge.’’ Today’s action 

updating Table IA at § 136.3 more 

clearly defines the removal of approved 

microbiological methods for ambient 

waters from this table. Such methods 

have been moved to a new table, Table 

IH. 

D. Revisions to 40 CFR Part 136, Table 

II and Footnotes 

The rule revises Table II (Required 

Containers, Preservation Techniques, 

and Holding Times), and the footnotes 

to Table II at 40 CFR 136.3(e). Today’s 

action updating Table II at § 136.3(e) 

more clearly defines the holding time 

for bacterial testing as 6 hours holding 

time with 2 hours to process samples. 

E. Revision to 40 CFR Part 503, 

Sampling and Analysis 

Based on comments received on the 

Agency’s proposal of methods for use in 

sewage sludge, EPA is including a cross 

reference to 40 CFR Part 136 in 40 CFR 

503.8(b) which prescribes the methods 

that must be used for sampling and 

analysis of sewage sludge. 

IV. Response to Comments 

EPA received 39 comments regarding 

methods included in this final rule from 

the August 16, 2005 proposal (70 FR 

48256), and 9 comments on the April 

10, 2006 Notice of Data Availability 

(NODA) (71 FR 18329). Commentors 

represented a number of different 

interests, including analytical 

laboratories, water utilities, instrument 

manufacturers, State and local 

governments, trade associations, 

scientists, and private citizens. The 

public docket for this rule includes the 

Agency’s response to all comments. The 

majority of the comments were with 

regard to method inclusion, method use, 

and quality control requirements. The 

following is a summary of our response 

to comments about the lack of 

connecting language between 40 CFR 

Parts 136 and 503 for sewage sludge 

methods. 

EPA proposed to approve methods in 

40 CFR Part 136 for sewage sludge but 

did not include an appropriate cross 

reference in 40 CFR Part 503 to Part 136 

so as to allow the use of appropriate 40 

CFR 136.3 methods as alternative 

methods to those listed in 40 CFR 503.8. 

Based on comments to the proposal, 

EPA has amended the language in 40 

CFR 503.8(b). In addition, as discussed 

above, EPA has also amended the 

language in 40 CFR 136.1 regarding the 

applicability of the methods in this 

section to 40 CFR Part 503. 

V. Statutory and Executive Order 

Reviews 

A. Executive Order 12866: Regulatory 

Planning and Review 

This action is not a ‘‘significant 

regulatory action’’ under the terms of 

Executive Order (EO) 12866 (58 FR 

51735, October 4, 1993) and is therefore 

not subject to review under the EO. 

B. Paperwork Reduction Act 

This action does not impose an 

information collection burden under the 

provisions of the Paperwork Reduction 

Act, 44 U.S.C. 3501 et seq. This rule 

does not impose any information 

collection, reporting, or recordkeeping 

requirements. This rule merely adds 

new and updated versions of testing 

procedures, withdraws some older 

testing procedures, and establishes new 

sample collection, preservation, and 

holding time requirements. 

Burden means the total time, effort, or 

financial resources expended by persons 

to generate, maintain, retain, or disclose 

or provide information to or for a 

Federal agency. This includes the time 

needed to review instructions; develop, 

acquire, install, and utilize technology 

and systems for the purpose of 

collecting, validating, and verifying 

information, processing and 

maintaining information, and disclosing 

and providing information; adjust the 

existing ways to comply with any 

previously applicable instructions and 

requirements; train personnel to be able 

to respond to a collection of 

information; search data sources; 

complete and review the collection of 

information; and transmit or otherwise 

disclose the information. 

An Agency may not conduct or 

sponsor, and a person is not required to 

respond to a collection of information 

unless it displays a currently valid OMB 

control number. The OMB control 

numbers for EPA’s regulations in 40 

CFR are listed in 40 CFR Part 9. 

C. Regulatory Flexibility Act 

The RFA generally requires an agency 

to prepare a regulatory flexibility 

analysis of any rule subject to notice 

and comment rulemaking requirements 

under the Administrative Procedure Act 

or any other statute unless the agency 

certifies that the rule will not have a 

significant economic impact on a 

substantial number of small entities. 

Small entities include small businesses, 

small organizations, and small 

governmental jurisdictions. 

For purposes of assessing the impacts 

of this rule on small entities for methods 

under the Clean Water Act, small entity 

is defined as: (1) A small business as 

defined by the Small Business 

Administration’s (SBA) regulations at 13 

CFR 121.201; (2) a small governmental 

jurisdiction that is a government of a 

city, county, town, school district or 

special district with a population less 

than 50,000; and (3) a small 

organization that is any not-for-profit 

enterprise which is independently 

owned and operated and is not 

dominant in its field. 

After considering the economic 

impacts of today’s final rule on small 

entities, I certify that this action will not 

have a significant economic impact on 

a substantial number of small entities. 


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Federal Register / Vol. 72, No. 57 / Monday, March 26, 2007 / Rules and Regulations 14223 

This final rule will not impose any 

requirements on small entities. This 

action approves new and updated 

versions of testing procedures, 

withdraws some older testing 

procedures, and approves new sample 

collection, preservation, and holding 

time requirements. Generally, these 

changes will have a positive impact on 

small entities by increasing method 

flexibility, thereby allowing entities to 

reduce costs by choosing more costeffective methods. In some cases, 

analytical costs may increase slightly 

due to the additional QC requirements 

included in the methods that are being 

approved. However, most laboratories 

that analyze samples for EPA 

compliance monitoring have already 

instituted QC requirements as part of 

their laboratory practices. 

D. Unfunded Mandates Reform Act 

Title II of the Unfunded Mandates 

Reform Act of 1995 (UMRA), Public 

Law 104–4, establishes requirements for 

Federal agencies to assess the effects of 

their regulatory actions on State, Tribal, 

and local governments and the private 

sector. Under section 202 of the UMRA, 

EPA generally must prepare a written 

statement, including a cost-benefit 

analysis, for proposed and final rules 

with ‘‘Federal mandates’’ that may 

result in expenditures to State, local, 

and Tribal governments, in the 

aggregate, or to the private sector, of 

$100 million or more in any one year. 

Before promulgating an EPA rule for 

which a written statement is needed, 

section 205 of the UMRA generally 

requires EPA to identify and consider a 

reasonable number of regulatory 

alternatives and adopt the least costly, 

most cost-effective or least burdensome 

alternative that achieves the objectives 

of the rule. The provisions of section 

205 do not apply when they are 

inconsistent with applicable law. 

Moreover, section 205 allows EPA to 

adopt an alternative other than the least 

costly, most cost-effective or least 

burdensome alternative if the 

Administrator publishes with the final 

rule an explanation of why that 

alternative was not adopted. 

Before EPA establishes any regulatory 

requirements that may significantly or 

uniquely affect small governments, 

including Tribal governments, it must 

have developed under section 203 of the 

UMRA a small government agency plan. 

The plan must provide for the 

notification of potentially affected small 

governments, enabling officials of 

affected small governments to have 

meaningful and timely input in the 

development of EPA regulatory 

proposals with significant Federal 

intergovernmental mandates, and 

informing, educating, and advising 

small governments on compliance with 

the regulatory requirements. 

This rule contains no Federal 

mandates (under the regulatory 

provisions of Title II of UMRA) for 

State, local, or Tribal governments or 

the private sector. The rule imposes no 

enforceable duty on any State, local, or 

Tribal governments or the private sector. 

In fact, this rule should (on the whole) 

save money for governments and the 

private sector by increasing method 

flexibility, and allowing these entities to 

reduce monitoring costs by taking 

advantage of innovations. Thus, today’s 

rule is not subject to the requirements 

of Sections 202 and 205 of the UMRA. 

EPA has determined that this rule 

contains no regulatory requirements that 

might significantly or uniquely affect 

small governments. Generally, this 

action will have a positive impact by 

increasing method flexibility, thereby 

allowing method users to reduce costs 

by choosing more cost effective 

methods. In some cases, analytical costs 

may increase slightly due to changes in 

methods, but these increases are neither 

significant nor unique to small 

governments. This rule merely approves 

new and updated versions of testing 

procedures, withdraws some older 

testing procedures, and approves new 

sample collection, preservation, and 

holding time requirements. Thus, 

today’s rule is not subject to the 

requirements of Section 203 of UMRA. 

E. Executive Order 13132: Federalism 

Executive Order 13132, entitled 

‘‘Federalism’’ (64 FR 43255, August 10, 

1999), requires EPA to develop an 

accountable process to ensure 

‘‘meaningful and timely input by State 

and local officials in the development of 

regulatory policies that have federalism 

implications.’’ ‘‘Policies that have 

federalism implications’’ is defined in 

the Executive Order to include 

regulations that have ‘‘substantial direct 

effects on the States, on the relationship 

between the national government and 

the States, or on the distribution of 

power and responsibilities among the 

various levels of government.’’

This final rule does not have 

federalism implications. It will not have 

substantial direct effects on the States, 

on the relationship between the national 

government and the States, or on the 

distribution of power and 

responsibilities among the various 

levels of government, as specified in 

Executive Order 13132. This rule merely 

approves new and updated versions of 

testing procedures, withdraws some 

older testing procedures, and approves 

new sample collection, preservation, 

and holding time requirements. The 

costs to State and local governments 

will be minimal (in fact, governments 

may see a cost savings), and the rule 

does not preempt State law. Thus, 

Executive Order 13132 does not apply 

to this rule. 

In the spirit of Executive Order 13132, 

and consistent with EPA policy to 

promote communications between EPA 

and State and local governments, EPA 

specifically solicited comment on the 

proposed rule from State and local 

officials. 

F. Executive Order 13175: Consultation 

and Coordination With Indian Tribal 

Governments 

Executive Order 13175, entitled 

‘‘Consultation and Coordination with 

Indian Tribal Governments’’ (65 FR 

67249, November 9, 2000), requires EPA 

to develop an accountable process to 

ensure ‘‘meaningful and timely input by 

tribal officials in the development of 

regulatory policies that have tribal 

implications.’’

This final rule does not have tribal 

implications, as specified in Executive 

Order 13175. It will not have substantial 

direct effects on Tribal governments, on 

the relationship between the Federal 

government and Indian tribes, or on the 

distribution of power and 

responsibilities between the Federal 

government and Indian tribes. This rule 

merely approves new and updated 

versions of testing procedures, 

withdraws some older testing 

procedures, and approves new sample 

collection, preservation, and holding 

time requirements. The costs to Tribal 

governments will be minimal (in fact, 

governments may see a cost savings), 

and the rule does not preempt State law. 

Thus, Executive Order 13175 does not 

apply to this rule. 

G. Executive Order 13045: Protection of 

Children From Environmental Health 

Risks and Safety Risks 

Executive Order 13045: ‘‘Protection of 

Children from Environmental Health 

Risks and Safety Risks’’ (62 FR 19885, 

April 23, 1997) applies to any rule that: 

(1) Is determined to be ‘‘economically 

significant’’ as defined under Executive 

Order 12866, and (2) concerns an 

environmental health or safety risk that 

EPA has reason to believe may have a 

disproportionate effect on children. If 

the regulatory action meets both criteria, 

the Agency must evaluate the 

environmental health or safety effects of 

the planned rule on children, and 

explain why the planned regulation is 

preferable to other potentially effective 

and reasonably feasible alternatives 


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14224 Federal Register / Vol. 72, No. 57 / Monday, March 26, 2007 / Rules and Regulations 

considered by the Agency. This final 

rule is not subject to the Executive 

Order 13045 because it is not 

economically significant as defined in 

Executive Order 12866. Further it does 

not concern an environmental health or 

safety risk that EPA has reason to 

believe may have a disproportionate 

effect on children. This action approves 

new and updated versions of testing 

procedures, withdraws some older 

testing procedures, and approves new 

sample collection, preservation, and 

holding time requirements. 

H. Executive Order 13211: Actions That 

Significantly Affect Energy Supply, 

Distribution, or Use 

This rule is not subject to Executive 

Order 13211, ‘‘Actions Concerning 

Regulations That Significantly Affect 

Energy Supply, Distribution, or Use’’ (66 

FR 28355 (May 22, 2001)) because it is 

not a significant regulatory action under 

Executive Order 12866. 

I. National Technology Transfer and 

Advancement Act 

As noted in the proposed rule, 

Section 12(d) of the National 

Technology Transfer and Advancement 

Act of 1995, (NTTAA), Public Law 104–

113, section 12(d) (15 U.S.C. 272 note), 

directs EPA to use voluntary consensus 

standards in its regulatory activities 

unless to do so would be inconsistent 

with applicable law or otherwise 

impractical. Voluntary consensus 

standards are technical standards (e.g., 

material specifications, test methods, 

sampling procedures, and business 

practices) that are developed or adopted 

by voluntary consensus standard bodies. 

The NTTAA directs EPA to provide 

Congress, through the OMB, 

explanations when the Agency decides 

not to use available and applicable 

voluntary consensus standards. This 

rulemaking involves technical 

standards. EPA has decided to use E. 

coli, enterococci and fecal coliform 

methods published in Standard 

Methods and ASTM International. 

The E. coli methods from Standard 

Methods are method 9223B (Standard 

Methods 18th, 19th and 20th Editions) 

and method 9223 B–97 (Standard 

Methods Online Edition), as well as 

AOAC method 991.15. The enterococci 

method from ASTM is method D6503–

99. The fecal coliform methods from 

Standard Methods are methods 9221 C 

E (Standard Methods 18th, 19th and 

20th Editions) and method 9221 C E–99 

(Standard Methods Online Edition). 

Standard Methods can be obtained from 

American Public Health Association, 

1015 15th Street, NW., Washington DC 

20005, AOAC methods can be obtained 

from Association of Official Analytical 

Chemists International, 481 North 

Frederick Avenue, Suite 500, 

Gaithersburg, MD 20877–2417, and 

ASTM methods can be obtained from 

ASTM International, 100 Barr Harbor 

Drive, West Conshohocken, PA 19428. 

These newly approved procedures 

reflect improvements in science and 

technology. EPA believes that the 

addition of these methods offer a wider 

variety of options that may be more cost 

effective to conduct compliance 

monitoring of bacterial pollutants. 

J. Congressional Review Act 

The Congressional Review Act, 5 

U.S.C. 801 et seq., as added by the Small 

Business Regulatory Enforcement 

Fairness Act of 1996, generally provides 

that before a rule may take effect, the 

agency promulgating the rule must 

submit a rule report, which includes a 

copy of the rule, to each House of the 

Congress and to the Comptroller General 

of the United States. EPA will submit a 

report containing this rule and other 

required information to the U.S. Senate, 

the U.S. House of Representatives, and 

the Comptroller General of the United 

States prior to publication of the rule in 

the Federal Register. A major rule 

cannot take effect until 60 days after it 

is published in the Federal Register. 

This action is not a ‘‘major rule’’ as 

defined by 5 U.S.C. 804(2). This rule 

will be effective April 25, 2007. 

List of Subjects 

40 CFR Part 136 

Environmental protection, 

Incorporation by reference, Reporting 

and recordkeeping requirements, Water 

pollution control. 

40 CFR Part 503 

Environmental protection, Reporting 

and recordkeeping requirements, Waste 

treatment and disposal, Water pollution 

control. 

Dated: September 28, 2006. 

Stephen L. Johnson, 

Administrator. 

Editorial Note: The Office of the Federal 

Register received this document on March 8, 

2007. 

■ For the reasons set out in the 

preamble, title 40, chapter I of the Code 

of Federal Regulations, is amended as 

follows: 

PART 136—GUIDELINES 

ESTABLISHING TEST PROCEDURES 

FOR THE ANALYSIS OF POLLUTANTS 

■ 1. The authority citation for Part 136 

continues to read as follows: 

Authority: Secs. 301, 304(h), 307, and 

501(a) Pub. L. 95–217, 91 Stat. 1566, et seq. 

(33 U.S.C. 1251, et seq.) (The Federal Water 

Pollution Control Act Amendments of 1972 

as amended by the Clean Water Act of 1977.) 

■ 2. Section 136.1 is revised to read as 

follows: 

§ 136.1 Applicability. 

(a) The procedures prescribed herein 

shall, except as noted in § 136.5, be used 

to perform the measurements indicated 

whenever the waste constituent 

specified is required to be measured for: 

(1) An application submitted to the 

Administrator, or to a State having an 

approved NPDES program for a permit 

under section 402 of the Clean Water 

Act of 1977, as amended (CWA), and/or 

to reports required to be submitted 

under NPDES permits or other requests 

for quantitative or qualitative effluent 

data under parts 122 to 125 of title 40, 

and, 

(2) Reports required to be submitted 

by dischargers under the NPDES 

established by parts 124 and 125 of this 

chapter, and, 

(3) Certifications issued by States 

pursuant to section 401 of the CWA, as 

amended. 

(b) The procedure prescribed herein 

and in part 503 of title 40 shall be used 

to perform the measurements required 

for an application submitted to the 

Administrator or to a State for a sewage 

sludge permit under section 405(f) of 

the Clean Water Act and for 

recordkeeping and reporting 

requirements under part 503 of title 40. 

■ 3. Section 136.3 is amended as 

follows: 

■ a. By revising paragraph (a) 

introductory text and Table IA. 

■ b. In paragraph (a) by adding Table IH 

after the notes of Table IG. 

■ c. In paragraph (b) by revising the 

introductory text and by revising 

references 2, 6, 10, 11, 34, 38, 39, and 

52 through 62; and by adding references 

70 through 72. 

■ d. By revising paragraph (e). 

§ 136.3 Identification of test procedures. 

(a) Parameters or pollutants, for which 

methods are approved, are listed 

together with test procedure 

descriptions and references in Tables 

IA, IB, IC, ID, IE, IF, IG, and IH. In the 

event of a conflict between the reporting 

requirements of 40 CFR Parts 122 and 

125 and any reporting requirements 

associated with the methods listed in 

these tables, the provisions of 40 CFR 

Parts 122 and 125 are controlling and 

will determine a permittee’s reporting 

requirements. The full text of the 

referenced test procedures are 

incorporated by reference into Tables 


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Federal Register / Vol. 72, No. 57 / Monday, March 26, 2007 / Rules and Regulations 14225 

IA, IB, IC, ID, IE, IF, IG, and IH. The 

incorporation by reference of these 

documents, as specified in paragraph (b) 

of this section, was approved by the 

Director of the Federal Register in 

accordance with 5 U.S.C. 552(a) and 1 

CFR Part 51. Copies of the documents 

may be obtained from the sources listed 

in paragraph (b) of this section. 

Documents may be inspected at EPA’s 

Water Docket, EPA West, 1301 

Constitution Avenue, NW., Room B102, 

Washington, DC (Telephone: 202–566–

2426); or at the National Archives and 

Records Administration (NARA). For 

information on the availability of this 

material at NARA, call 202–741–6030, 

or go to: http://www.archives.gov/ 

federal_register/ 

code_of_federal_regulations/ 

ibr_locations.html. These test 

procedures are incorporated as they 

exist on the day of approval and a notice 

of any change in these test procedures 

will be published in the Federal 

Register. The discharge parameter 

values for which reports are required 

must be determined by one of the 

standard analytical test procedures 

incorporated by reference and described 

in Tables IA, IB, IC, ID, IE, IF, IG, and 

IH or by any alternate test procedure 

which has been approved by the 

Administrator under the provisions of 

paragraph (d) of this section and 

§§ 136.4 and 136.5. Under certain 

circumstances paragraph (c) of this 

section, § 136.5(a) through (d) or 40 CFR 

401.13, other additional or alternate test 

procedures may be used. 

TABLE IA.—LIST OF APPROVED BIOLOGICAL METHODS FOR WASTEWATER AND SEWAGE SLUDGE

Parameter and units Method 1 EPA 

Standard methods 18th, 19th, 

20th ed. 

Standard methods online AOAC, ASTM, 

USGS Other 

Bacteria: 

1. Coliform (fecal), 

number per 100 

mL or number per 

gram dry weight.

Most Probable Number 

(MPN),5 tube 3 dilution, or 

p. 132 3 ............

1680 12,14 .........

1681 12,19 .........

9221 C E ......... 9221 C E–99.

Membrane filter (MF) 2, 

single step.

p. 124 3 ............ 9222 D ............ 9222 D–97 ......... B–0050–85 5.

2. Coliform (fecal) in 

presence of chlorine, number per 

100 mL.

MPN, 5 tube, 3 dilution, 

or 

p. 132 3 ............ 9221 C E ......... 9221 C E–99.

MF 2, single step .......... p. 124 3 ............ 9222 D ............ 9222 D–97.

3. Coliform (total), 

number per 100 

mL.

MPN, 5 tube, 3 dilution, 

or 

p. 114 3 ............ 9221 B ............. 9221 B–99.

MF 2, single step or two 

step.

p. 108 3 ............ 9222 B ............. 9222 B–97 ......... B–0025–8 5.

4. Coliform (total), in 

presence of chlorine, number per 

100 mL.

MPN, 5 tube, 3 dilution, 

or 

p. 114 3 ............ 9221 B ............. 9221 B–99.

MF 2 with enrichment ... p. 111 3 ............ 9222 (B+B.5c) 9222 

(B+B.5c)¥97.

5. E. coli, number 

per 100 mL 20.

MPN 7,9,15 multiple 

tube/multiple well.

......................... 9223 B13 ......... 9223 B–97 13 ..... 991.15 11 ......... Colilert13,17

Colilert1813,16,17

MF 2,6,7,8,9 single step .. 1603 21 ............ ......................... ............................ ......................... mColiBlue2418

6. Fecal streptococci, 

number per 100 

mL.

MPN, 5 tube 3 dilution, p. 139 3 ............ 9230 B ............. 9230 B–93.

MF 2, or ........................ p. 136 3 ............ 9230 C ............ 9230 C–93 ......... B–0055–85 5.

Plate count ................... p. 143 3.

7. Enterococci, number per 100 mL 20.MPN 7,9, multiple tube/ 

multiple well.

......................... ......................... ............................ D6503–99 10 .... Enterolert 13,23

MF 2,6,7,8,9 single step .. 1600 24.

8. Salmonella, number per gram dry 

weight 12.

MPN multiple tube ....... 1682 22.

Aquatic Toxicity: 

9. Toxicity, acute, 

fresh water organisms, LC 50, percent effluent.

Ceriodaphnia dubia 

acute.

2002.0 25.

Daphnia puplex and 

Daphnia magna 

acute.

2021.0 25.

Fathead Minnow, 

Pimephales 

promelas, and 

Bannerfin shiner, 

Cyprinella leedsi, 

acute.

2000.0 25.


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TABLE IA.—LIST OF APPROVED BIOLOGICAL METHODS FOR WASTEWATER AND SEWAGE SLUDGE—Continued 

Parameter and units Method 1 EPA 

Standard methods 18th, 19th, 

20th ed. 

Standard methods online AOAC, ASTM, 

USGS Other 

Rainbow Trout, 

Oncorhynchus 

mykiss, and brook 

trout, Salvelinus 

fontinalis, acute.

2019.0 25.

10. Toxicity, acute, 

estuarine and marine organisms of 

the Atlantic Ocean 

and Gulf of Mexico, LC50, percent 

effluent.

Mysid, Mysidopsis 

bahia, acute.

2007.0 25.

Sheepshead Minnow, 

Cyprinodon 

variegatus, acute.

2004.0 25.

Silverside, Menidia 

beryllina, Menidia 

menidia, and Menidia 

peninsulae, acute.

2006.0 25.

11. Toxicity, chronic, 

fresh water organisms, NOEC or 

IC25, percent effluent.

Fathead minnow, 

Pimephales 

promelas, larval survival and growth.

1000.0 26.

Fathead minnow, 

Pimephales 

promelas, embryolarval survival and 

teratogenicity.

1001.0 26.

Daphnia, Ceriodaphnia 

dubia, survival and 

reproduction.

1002.0 26.

Green alga, 

Selenastrum 

capricornutum, 

growth.

1003.0 26.

12. Toxicity, chronic, 

estuarine and marine organisms of 

the Atlantic Ocean 

and Gulf of Mexico, NOEC or IC25, 

percent effluent.

Sheepshead minnow, 

Cyprinodon 

variegatus, larval survival and growth.

1004.0 27.

Sheepshed minnow, 

Cyprinodon 

variegatus, embryolarval survival and 

teratogenicity.

1005.0 27.

Inland silverside, 

Menidia beryllina, larval survival and 

growth.

1006.0 27.

Mysid, Mysidopsis 

bahia, survival, 

growth, and fecundity.

1007.0 27.

Sea urchin, Arbacia 

punctulata, fertilization.

1008.0 27.

1The method must be specified when results are reported. 

2A 0.45 μm membrane filter (MF) or other pore size certified by the manufacturer to fully retain organisms to be cultivated and to be free of 

extractables which could interfere with their growth. 3 USEPA. 1978. Microbiological Methods for Monitoring the Environment, Water, and Wastes. Environmental Monitoring and Support Laboratory, U.S. Environmental Protection Agency, Cincinnati, OH, EPA/600/8–78/017. 4 [Reserved]. 

5 USGS. 1989. U.S. Geological Survey Techniques of Water-Resource Investigations, Book 5, Laboratory Analysis, Chapter A4, Methods for 

Collection and Analysis of Aquatic Biological and Microbiological Samples, U.S. Geological Survey, U.S. Department of the Interior, Reston, VA. 6Because the MF technique usually yields low and variable recovery from chlorinated wastewaters, the Most Probable Number method will be 

required to resolve any controversies. 7Tests must be conducted to provide organism enumeration (density). Select the appropriate configuration of tubes/filtrations and dilutions/volumes to account for the quality, character, consistency, and anticipated organism density of the water sample. 


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8When the MF method has been used previously to test waters with high turbidity, large numbers of noncoliform bacteria, or samples that may 

contain organisms stressed by chlorine, a parallel test should be conducted with a multiple-tube technique to demonstrate applicability and comparability of results. 9To assess the comparability of results obtained with individual methods, it is suggested that side-by-side tests be conducted across seasons 

of the year with the water samples routinely tested in accordance with the most current Standard Methods for the Examination of Water and 

Wastewater or EPA alternate test procedure (ATP) guidelines. 10ASTM. 2000, 1999, 1996. Annual Book of ASTM Standards—Water and Environmental Technology. Section 11.02. ASTM International. 100 

Barr Harbor Drive, West Conshohocken, PA 19428. 11AOAC. 1995. Official Methods of Analysis of AOAC International, 16th Edition, Volume I, Chapter 17. Association of Official Analytical 

Chemists International. 481 North Frederick Avenue, Suite 500, Gaithersburg, MD 20877–2417. 12 Recommended for enumeration of target organism in sewage sludge. 13These tests are collectively known as defined enzyme substrate tests, where, for example, a substrate is used to detect the enzyme b-glucuronidase produced by E. coli. 14 USEPA. July 2006. Method 1680: Fecal Coliforms in Sewage Sludge (Biosolids) by Multiple-Tube Fermentation Using Lauryl-Tryptose Broth 

(LTB) and EC Medium. US Environmental Protection Agency, Office of Water, Washington, DC EPA–821–R–06–012. 15Samples shall be enumerated by the multiple-tube or multiple-well procedure. Using multiple-tube procedures, employ an appropriate tube 

and dilution configuration of the sample as needed and report the Most Probable Number (MPN). Samples tested with Colilert may be enumerated with the multiple-well procedures, Quanti-Tray Quanti-Tray 2000, and the MPN calculated from the table provided by the manufacturer. 16 Colilert-18 is an optimized formulation of the Colilert for the determination of total coliforms and E. coli that provides results within 18 h of 

incubation at 35 °C rather than the 24 h required for the Colilert test and is recommended for marine water samples. 17 Descriptions of the Colilert, Colilert-18, Quanti-Tray, and Quanti-Tray/2000 may be obtained from IDEXX Laboratories, Inc., 1 IDEXX 

Drive, Westbrook, ME 04092. 18A description of the mColiBlue24 test, Total Coliforms and E. coli, is available from Hach Company, 100 Dayton Ave., Ames, IA 50010. 19 USEPA. July 2006. Method 1681: Fecal Coliforms in Sewage Sludge (Biosolids) by Multiple-Tube Fermentation using A–1 Medium. U.S. Environmental Protection Agency, Office of Water, Washington, DC EPA–821–R–06–013. 20 Recommended for enumeration of target organism in wastewater effluent. 21 USEPA. July 2006. Method 1603: Escherichia coli (E. coli) in Water by Membrane Filtration Using Modified membrane-Thermotolerant Escherichia coli Agar (modified mTEC). U.S. Environmental Protection Agency, Office of Water, Washington, DC EPA–821–R–06–011. 22 USEPA. July 2006. Method 1682: Salmonella in Sewage Sludge (Biosolids) by Modified Semisolid Rappaport-Vassiliadis (MSRV) Medium. 

U.S. Environmental Protection Agency, Office of Water, Washington, DC EPA–821–R–06–014. 23A description of the Enterolert test may be obtained from IDEXX Laboratories, Inc., 1 IDEXX Drive, Westbrook, ME 04092. 24 USEPA. July 2006. Method 1600: Enterococci in Water by Membrane Filtration Using membrane-Enterococcus Indoxyl-b-D-Glucoside Agar 

(mEI). U.S. Environmental Protection Agency, Office of Water, Washington, DC EPA–821–R–06–009. 25 USEPA. October 2002. Methods for Measuring the Acute Toxicity of Effluents and Receiving Waters to Freshwater and Marine Organisms. 

Fifth Edition. U.S. Environmental Protection Agency, Office of Water, Washington, DC EPA/821/R–02/012. 26 USEPA. October 2002. Short-term Methods for Estimating the Chronic Toxicity of Effluents and Receiving Waters to Freshwater Organisms. 

Fourth Edition, U.S. Environmental Protection Agency, Office of Water, Washington, DC EPA/821/R–02/013. 27 USEPA. October 2002. Short-term Methods for Estimating the Chronic Toxicity of Effluents and Receiving Waters to Marine and Estuarine 

Organisms. Third Edition. U.S. Environmental Protection Agency, Office of Water, Washington, DC EPA/821/R–02/014. 

* * * * * 

TABLE IH.—LIST OF APPROVED MICROBIOLOGICAL METHODS FOR AMBIENT WATER

Parameter and units Method 1 EPA 

Standard 

methods 18th, 

19th, 20th Ed. 

Standard methods online AOAC, ASTM, 

USGS Other 

Bacteria: 

1. E. coli, number 

per 100 mL.

MPN 6,8,14 multiple tube, .......................... 9221 B.1/9221 

F 11,13.

9221 B.1–99/ 

9221 F 11,13.

Multiple tube/multiple 

well, 

.......................... 9223 B12 ......... 9223 B–97 12 ... 991.15 10 .......... Colilert 12,16

Colilert18 12,15,16. 

MF 2,5,6,7,8 two step, or 1103.1 19 .......... 9222 B/9222 

G 18, 9213 D.

9222 B–97/ 

9222 G 18.

D5392–93 9.

Single step .................... 1603 20, 1604 21 .......................... .......................... .......................... mColiBlue24 17. 

2. Enterococci, number per 100 mL.MPN 6,8 multiple tube, .......................... 9230 B ............. 9230 B–93.

Multiple tube/multiple 

well.

.......................... .......................... .......................... D6503–99 9 ...... Enterolert 12,22. 

MF 2,5,6,7,8 two step ...... 1106.1 23 .......... 9230 C ............. 9230 C–93 ....... D5259–92 9.

Single step, or .............. 1600 24.

Plate count ................... p. 143 3.

Protozoa: 

3. Cryptosporidium .. Filtration/IMS/FA ........... 1622 25,1623 26.

4. Giardia ................. Filtration/IMS/FA ........... 1623 26.

1The method must be specified when results are reported. 

2A 0.45 μm membrane filter (MF) or other pore size certified by the manufacturer to fully retain organisms to be cultivated and to be free of 

extractables which could interfere with their growth. 3 USEPA. 1978. Microbiological Methods for Monitoring the Environment, Water, and Wastes. Environmental Monitoring and Support Laboratory, U.S. Environmental Protection Agency, Cincinnati, OH. EPA/600/8–78/017. 4 [Reserved] 

5Because the MF technique usually yields low and variable recovery from chlorinated wastewaters, the Most Probable Number method will be 

required to resolve any controversies. 6Tests must be conducted to provide organism enumeration (density). Select the appropriate configuration of tubes/filtrations and dilutions/volumes to account for the quality, character, consistency, and anticipated organism density of the water sample. 


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14228 Federal Register / Vol. 72, No. 57 / Monday, March 26, 2007 / Rules and Regulations 

7When the MF method has not been used previously to test waters with high turbidity, large number of noncoliform bacteria, or samples that 

may contain organisms stressed by chlorine, a parallel test should be conducted with a multiple-tube technique to demonstrate applicability and 

comparability of results. 8To assess the comparability of results obtained with individual methods, it is suggested that side-by-side tests be conducted across seasons 

of the year with the water samples routinely tested in accordance with the most current Standard Methods for the Examination of Water and 

Wastewater or EPA alternate test procedure (ATP) guidelines. 9ASTM. 2000, 1999, 1996. Annual Book of ASTM Standards—Water and Environmental Technology. Section 11.02. ASTM International. 100 

Barr Harbor Drive, West Conshohocken, PA 19428. 10AOAC. 1995. Official Methods of Analysis of AOAC International, 16th Edition, Volume I, Chapter 17. Association of Official Analytical 

Chemists International. 481 North Frederick Avenue, Suite 500, Gaithersburg, MD 20877–2417. 11The multiple-tube fermentation test is used in 9221B.1. Lactose broth may be used in lieu of lauryl tryptose broth (LTB), if at least 25 parallel 

tests are conducted between this broth and LTB using the water samples normally tested, and this comparison demonstrates that the false-positive rate and false-negative rate for total coliform using lactose broth is less than 10 percent. No requirement exists to run the completed phase 

on 10 percent of all total coliform-positive tubes on a seasonal basis. 12These tests are collectively known as defined enzyme substrate tests, where, for example, a substrate is used to detect the enzyme b-glucuronidase produced by E. coli. 13After prior enrichment in a presumptive medium for total coliform using 9221B.1, all presumptive tubes or bottles showing any amount of 

gas, growth or acidity within 48 h ± 3 h of incubation shall be submitted to 9221F. Commercially available EC–MUG media or EC media supplemented in the laboratory with 50 μg/mL of MUG may be used. 14Samples shall be enumerated by the multiple-tube or multiple-well procedure. Using multiple-tube procedures, employ an appropriate tube 

and dilution configuration of the sample as needed and report the Most Probable Number (MPN). Samples tested with Colilert may be enumerated with the multiple-well procedures, Quanti-Tray or Quanti-Tray 2000, and the MPN calculated from the table provided by the manufacturer. 15 Colilert-18 is an optimized formulation of the Colilert for the determination of total coliforms and E. coli that provides results within 18 h of 

incubation at 35 °C rather than the 24 h required for the Colilert test and is recommended for marine water samples. 16 Descriptions of the Colilert, Colilert-18, Quanti-Tray, and Quanti-Tray/2000 may be obtained from IDEXX Laboratories, Inc., 1 IDEXX 

Drive, Westbrook, ME 04092. 17A description of the mColiBlue24 test, Total Coliforms and E. coli, is available from Hach Company, 100 Dayton Ave., Ames, IA 50010. 18Subject total coliform positive samples determined by 9222B or other membrane filter procedure to 9222G using NA–MUG media. 19 USEPA. July 2006. Method 1103.1: Escherichia coli (E. coli) in Water by Membrane Filtration Using membrane-Thermotolerant Escherichia 

coli Agar (mTEC). U.S. Environmental Protection Agency, Office of Water, Washington, DC EPA–821–R–06–010. 20 USEPA. July 2006. Method 1603: Escherichia coli (E. coli) in Water by Membrane Filtration Using Modified membrane-Thermotolerant Escherichia coli Agar (Modified mTEC). U.S. Environmental Protection Agency, Office of Water, Washington, DC EPA–821–R–06–011. 21Preparation and use of MI agar with a standard membrane filter procedure is set forth in the article, Brenner et al. 1993. ‘‘New Medium for 

the Simultaneous Detection of Total Coliform and Escherichia coli in Water.’’ Appl. Environ. Microbiol. 59:3534–3544 and in USEPA. September 

2002.: Method 1604: Total Coliforms and Escherichia coli (E. coli) in Water by Membrane Filtration by Using a Simultaneous Detection Technique (MI Medium). U.S. Environmental Protection Agency, Office of Water, Washington, DC EPA 821–R–02–024. 22A description of the Enterolert test may be obtained from IDEXX Laboratories, Inc., 1 IDEXX Drive, Westbrook, ME 04092. 23 USEPA. July 2006. Method 1106.1: Enterococci in Water by Membrane Filtration Using membrane-Enterococcus-Esculin Iron Agar (mE–

EIA). U.S. Environmental Protection Agency, Office of Water, Washington, DC EPA–821–R–06–008. 24 USEPA. July 2006. Method 1600: Enterococci in Water by Membrane Filtration Using membrane-Enterococcus Indoxyl-b-D-Glucoside Agar 

(mEI). U.S. Environmental Protection Agency, Office of Water, Washington, DC EPA–821–R–06–009. 25 Method 1622 uses filtration, concentration, immunomagnetic separation of oocysts from captured material, immunofluorescence assay to determine concentrations, and confirmation through vital dye staining and differential interference contrast microscopy for the detection of 

Cryptosporidium. USEPA. 2001. Method 1622: Cryptosporidium in Water by Filtration/IMS/FA. U.S. Environmental Protection Agency, Office of 

Water, Washington, DC EPA–821–R–01–026. 26 Method 1623 uses filtration, concentration, immunomagnetic separation of oocysts and cysts from captured material, immunofluorescence 

assay to determine concentrations, and confirmation through vital dye staining and differential interference contrast microscopy for the simultaneous detection of Cryptosporidium and Giardia oocysts and cysts. USEPA. 2001. Method 1623. Cryptosporidium and Giardia in Water by Filtration/IMS/FA. U.S. Environmental Protection Agency, Office of Water, Washington, DC EPA–821–R–01–025. 

(b) The full texts of the methods from 

the following references which are cited 

in Tables IA, IB, IC, ID, IE, IF, IG and 

IH are incorporated by reference into 

this regulation and may be obtained 

from the source identified. All costs 

cited are subject to change and must be 

verified from the indicated source. The 

full texts of all the test procedures cited 

are available for inspection at the 

National Archives and Records 

Administration (NARA). For 

information on the availability of this 

material at NARA, call 202–741–6030, 

or go to: http://www.archives.gov/ 

federal_register/ 

code_of_federal_regulations/ 

ibr_locations.html. 

References, Sources, Costs, and Table 

Citations 

* * * * * 

(2) USEPA. 1978. Microbiological 

Methods for Monitoring the 

Environment, Water, and Wastes. 

Environmental Monitoring and Support 

Laboratory, U.S. Environmental 

Protection Agency, Cincinnati, Ohio. 

EPA/600/8–78/017. Available at http:// 

www.epa.gov/clariton/srch.htm or from: 

National Technical Information Service, 

5285 Port Royal Road, Springfield, 

Virginia 22161, Pub. No. PB–290329/ 

A.S. Table IA, Note 3; Table IH, Note 3. 

* * * * * 

(6) American Public Health 

Association. 1992, 1995, and 1998. 

Standard Methods for the Examination 

of Water and Wastewater. 18th, 19th, 

and 20th Edition (respectively). 

Available from: American Public Health 

Association, 1015 15th Street, NW., 

Washington, DC 20005. Standard 

Methods Online is available through the 

Standard Methods Web site (http:// 

www.standardmethods.org). Tables IA, 

IB, IC, ID, IE, and IH. 

* * * * * 

(10) ASTM International. Annual 

Book of ASTM Standards, Water, and 

Environmental Technology, Section 11, 

Volumes 11.01 and 11.02, 1994, 1996, 

1999, Volume 11.02, 2000, and 

individual standards published after 

2000. Available from: ASTM 

International, 100 Barr Harbor Drive, 

P.O. Box C700, West Conshohocken, PA 

19428–2959, or http://www.astm.org. 

Tables IA, IB, IC, ID, IE, and IH. 

* * * * * 

(11) USGS. 1989. U.S. Geological 

Survey Techniques of Water-Resources 

Investigations, Book 5, Laboratory 

Analysis, Chapter A4, Methods for 

Collection and Analysis of Aquatic 

Biological and Microbiological Samples, 

U.S. Geological Survey, U.S. 

Department of the Interior, Reston, 

Virginia. Available from USGS Books 

and Open-File Reports Section, Federal 

Center, Box 25425, Denver, Colorado 

80225. Table IA, Note 5; Table IH. 

* * * * * 

(34) USEPA. October 2002. Methods 

for Measuring the Acute Toxicity of 

Effluents and Receiving Waters to 

Freshwater and Marine Organisms. Fifth 

Edition. U.S. Environmental Protection 

Agency, Office of Water, Washington, 

DC EPA 821–R–02–012. Available at 

http://www.epa.gov/epahome/index/ 


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sources.htm or from National Technical 

Information Service, 5285 Port Royal 

Road, Springfield, Virginia 22161, Pub. 

No. PB2002–108488. Table IA, Note 25. 

* * * * * 

(38) USEPA. October 2002. ShortTerm Methods for Measuring the 

Chronic Toxicity of Effluents and 

Receiving Waters to Freshwater 

Organisms. Fourth Edition. U.S. 

Environmental Protection Agency, 

Office of Water, Washington, DC EPA 

821–R–02–013. Available at http:// 

www.epa.gov/epahome/index/ 

sources.htm or from National Technical 

Information Service, 5285 Port Royal 

Road, Springfield, Virginia 22161, Pub. 

No. PB2002–108489. Table IA, Note 26. 

(39) USEPA. October 2002. ShortTerm Methods for Measuring the 

Chronic Toxicity of Effluents and 

Receiving Waters to Marine and 

Estuarine Organisms. Third Edition. 

U.S. Environmental Protection Agency, 

Office of Water, Washington, DC EPA 

821–R–02–014. Available at http:// 

www.epa.gov/epahome/index/ 

sources.htm or from National Technical 

Information Service, 5285 Port Royal 

Road, Springfield, Virginia 22161, Pub. 

No. PB2002–108490. Table IA, Note 27. 

* * * * * 

(52) IDEXX Laboratories, Inc. 2002. 

Description of Colilert, Colilert-18, 

Quanti-Tray, Quanti-Tray/2000, 

Enterolert methods are available from 

IDEXX Laboratories, Inc., One Idexx 

Drive, Westbrook, Maine 04092. Table 

IA, Notes 17 and 23; Table IH, Notes 16 

and 22. 

(53) Hach Company, Inc. Revision 2, 

1999. Description of m-ColiBlue24 

Method, Total Coliforms and E. coli, is 

available from Hach Company, 100 

Dayton Ave, Ames IA 50010. Table IA, 

Note 18; Table IH, Note 17. 

(54) USEPA. July 2006. Method 

1103.1: Escherichia coli (E. coli) in 

Water by Membrane Filtration Using 

membrane-Thermotolerant Escherichia 

coli Agar (mTEC). U.S. Environmental 

Protection Agency, Office of Water, 

Washington DC EPA–621–R–06–010. 

Available at http://www.epa.gov/ 

waterscience/methods/. Table IH, Note 

19. 

(55) USEPA. July 2006. Method 

1106.1: Enterococci in Water by 

Membrane Filtration Using membraneEnterococcus-Esculin Iron Agar (mE–

EIA). U.S. Environmental Protection 

Agency, Office of Water, Washington DC 

EPA–621–R–06–008. Available at 

http://www.epa.gov/waterscience/ 

methods/. Table IH, Note 23 

(56) USEPA. July 2006. Method 1603: 

Escherichia coli (E. coli) in Water by 

Membrane Filtration Using Modified 

membrane-Thermotolerant Escherichia 

coli Agar (Modified mTEC). U.S. 

Environmental Protection Agency, 

Office of Water, Washington DC EPA–

821–R–06–011. Available at http:// 

www.epa.gov/waterscience/methods/. 

Table IH, Note 19; Table IH, Note 20. 

(57) Brenner et al. 1993. New Medium 

for the Simultaneous Detection of Total 

Coliforms and Escherichia coli in Water. 

Appl. Environ. Microbiol. 59:3534–

3544. Available from the American 

Society for Microbiology, 1752 N Street 

NW., Washington DC 20036. Table IH, 

Note 21. 

(58) USEPA. September 2002. Method 

1604: Total Coliforms and Escherichia 

coli (E. coli) in Water by Membrane 

Filtration Using a Simultaneous 

Detection Technique (MI Medium). U.S. 

Environmental Protection Agency, 

Office of Water, Washington DC EPA–

821–R–02–024. Available at http:// 

www.epa.gov/waterscience/methods/. 

Table IH, Note 20. 

(59) USEPA. July 2006. Method 1600: 

Enterococci in Water by Membrane 

Filtration Using membraneEnterococcus Indoxyl-b-D-Glucoside 

Agar (mEI). U.S. Environmental 

Protection Agency, Office of Water, 

Washington DC EPA–821–R–06–009. 

Available at http://www.epa.gov/ 

waterscience/methods/. Table IA, Note 

24; Table IH, Note 24. 

(60) USEPA. April 2001. Method 

1622: Cryptosporidium in Water by 

Filtration/IMS/FA. U.S. Environmental 

Protection Agency, Office of Water, 

Washington DC EPA–821–R–01–026. 

Available at http://www.epa.gov/ 

waterscience/methods/. Table IH, Note 

25. 

(61) USEPA. April 2001. Method 

1623: Cryptosporidium and Giardia in 

Water by Filtration/IMS/FA. U.S. 

Environmental Protection Agency, 

Office of Water, Washington DC. EPA–

821–R–01–025. Available at http:// 

www.epa.gov/waterscience/methods/. 

Table IH, Note 26. 

(62) AOAC. 1995. Official Methods of 

Analysis of AOAC International, 16th 

Edition, Volume I, Chapter 17. AOAC 

International, 481 North Frederick 

Avenue, Suite 500, Gaithersburg, 

Maryland 20877–2417. Table IA, Note 

11; Table IH. 

* * * * * 

(70) USEPA. July 2006. Method 1680: 

Fecal Coliforms in Sewage Sludge 

(Biosolids) by Multiple-Tube 

Fermentation using Lauryl Tryptose 

Broth (LTB) and EC Medium. U.S. 

Environmental Protection Agency, 

Office of Water, Washington DC. EPA 

821–R–06–012. Available at http:// 

www.epa.gov/waterscience/methods/. 

(71) USEPA. July 2006. Method 1681: 

Fecal Coliforms in Sewage Sludge 

(Biosolids) by Multiple-Tube 

Fermentation using A–1 Medium. U.S. 

Environmental Protection Agency, 

Office of Water, Washington DC. EPA 

821–R–06–013. Available at http:// 

www.epa.gov/waterscience/methods/. 

(72) USEPA. July 2006. Method 1682: 

Salmonella in Sewage Sludge 

(Biosolids) by Modified Semisolid 

Rappaport-Vassiliadis (MSRV) Medium. 

U.S. Environmental Protection Agency, 

Office of Water, Washington DC. EPA 

821–R–06–014. Available at http:// 

www.epa.gov/waterscience/methods/. 

* * * * * 

(e) Sample preservation procedures, 

container materials, and maximum 

allowable holding times for parameters 

are cited in Tables IA, IB, IC, ID, IE, IF, 

IG and IH are prescribed in Table II. 

Information in the table takes 

precedence over information in specific 

methods or elsewhere. Any person may 

apply for a variance from the prescribed 

preservation techniques, container 

materials, and maximum holding times 

applicable to samples taken from a 

specific discharge. Applications for 

variances may be made by letters to the 

Regional Administrator in the Region in 

which the discharge will occur. 

Sufficient data should be provided to 

assure such variance does not adversely 

affect the integrity of the sample. Such 

data will be forwarded by the Regional 

Administrator, to the Alternate Test 

Procedure Program Coordinator, 

Washington, DC, for technical review 

and recommendations for action on the 

variance application. Upon receipt of 

the recommendations from the Alternate 

Test Procedure Program Coordinator, 

the Regional Administrator may grant a 

variance applicable to the specific 

discharge to the applicant. A decision to 

approve or deny a variance will be made 

within 90 days of receipt of the 

application by the Regional 

Administrator. 


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14230 Federal Register / Vol. 72, No. 57 / Monday, March 26, 2007 / Rules and Regulations 

TABLE II.—REQUIRED CONTAINERS, PRESERVATION TECHNIQUES, AND HOLDING TIMES

Parameter No./name Container 1 Preservation 2,3 Maximum holding time 4

Table IA—Bacterial Tests: 

1–5. Coliform, total, fecal, and E. coli .............. PA, G ............................. Cool, <10 °C, 0.0008% Na2S2O3 5 .... 6 hours.22,23

6. Fecal streptococci ........................................ PA, G ............................. Cool, <10 °C, 0.0008% Na2S2O3 5 .... 6 hours.22

7. Enterococci ................................................... PA, G ............................. Cool, <10 °C, 0.0008% Na2S2O3 5 .... 6 hours.22

8. Salmonella .................................................... PA, G ............................. Cool, <10 °C, 0.0008% Na2S2O3 5 .... 6 hours.22

Table IA—Aquatic Toxicity Tests: 

9–11. Toxicity, acute and chronic .................... P, FP, G ........................ Cool, ≤6 °C 16 ..................................... 36 hours. 

Table lB—Inorganic Tests: 

1. Acidity ........................................................... P, FP, G ........................ Cool, ≤6 °C 18 ..................................... 14 days. 

2. Alkalinity ....................................................... P, FP, G ........................ Cool, ≤6 °C 18 ..................................... 14 days. 

4. Ammonia ...................................................... P, FP, G ........................ Cool, ≤6 °C 18, H2SO4 to pH<2 ......... 28 days. 

9. Biochemical oxygen demand ....................... P, FP, G ........................ Cool, ≤6 °C 18 ..................................... 48 hours. 

10. Boron .......................................................... P, FP, or Quartz ............ HNO3 to pH<2 ................................... 6 months. 

11. Bromide ...................................................... P, FP, G ........................ None required .................................... 28 days. 

14. Biochemical oxygen demand, carbonaceous.P, FP G ......................... Cool, ≤6 °C 18 ..................................... 48 hours. 

15. Chemical oxygen demand .......................... P, FP, G ........................ Cool, ≤6 °C 18, H2SO4 to pH<2 ......... 28 days. 

16. Chloride ...................................................... P, FP, G ........................ None required .................................... 28 days. 

17. Chlorine, total residual ............................... P, G ............................... None required .................................... Analyze within 15 minutes. 

21. Color ........................................................... P, FP, G ........................ Cool, ≤6 °C 18 ..................................... 48 hours. 

23–24. Cyanide, total or available (or CATC) .. P, FP, G ........................ Cool, ≤6 °C 18, NaOH to pH>12 6, reducing agent 5.14 days. 

25. Fluoride ....................................................... P .................................... None required .................................... 28 days. 

27. Hardness .................................................... P, FP, G ........................ HNO3 or H2SO4 to pH<2 .................. 6 months. 

28. Hydrogen ion (pH) ...................................... P, FP, G ........................ None required .................................... Analyze within 15 minutes. 

31, 43. Kjeldahl and organic N ......................... P, FP, G ........................ Cool, ≤6 °C 18, H2SO4 to pH<2 ......... 28 days. 

Table IB—Metals: 7

18. Chromium VI .............................................. P, FP, G ........................ Cool, ≤6 °C 18, pH = 9.3–9.7 20 .......... 28 days. 

35. Mercury (CVAA) ......................................... P, FP, G ........................ HNO3 to pH<2 ................................... 28 days. 

35. Mercury (CVAFS) ....................................... FP, G; and FP-lined 

cap 17.

5 mL/L 12N HCl or 5 mL/L BrCl 17 .... 90 days.17

3, 5–8, 12, 13, 19, 20, 22, 26, 29, 30, 32–34, 

36, 37, 45, 47, 51, 52, 58–60, 62, 63, 70–

72, 74, 75.

P, FP, G ........................ HNO3 to pH<2, or at least 24 hours 

prior to analysis 19.

6 months. 

Metals, except boron, chromium VI, and mercury.

38. Nitrate ......................................................... P, FP, G ........................ Cool, ≤6 °C 18 ..................................... 48 hours. 

39. Nitrate-nitrite ............................................... P, FP, G ........................ Cool, ≤6 °C 18, H2SO4 to pH<2 ......... 28 days. 

40. Nitrite .......................................................... P, FP, G ........................ Cool, ≤6 °C 18 ..................................... 48 hours. 

41. Oil and grease ............................................ G .................................... Cool to ≤6 °C 18, HCl or H2SO4 to 

pH<2.

28 days. 

42. Organic Carbon .......................................... P, FP, G ........................ Cool to ≤6 °C 18, HCl, H2SO4, or 

H3PO4 to pH<2.

28 days. 

44. Orthophosphate .......................................... P, FP, G ........................ Cool, ≤6 °C 18 ..................................... Filter within 15 minutes; 

Analyze within 48 

hours. 

46. Oxygen, Dissolved Probe ........................... G, Bottle and top ........... None required .................................... Analyze within 15 minutes. 

47. Winkler ........................................................ G, Bottle and top ........... Fix on site and store in dark ............. 8 hours. 

48. Phenols ....................................................... G .................................... Cool, ≤6 °C 18, H2SO4 to pH<2 ......... 28 days. 

49. Phosphorous (elemental) ........................... G .................................... Cool, ≤6 °C 18 ..................................... 48 hours. 

50. Phosphorous, total ..................................... P, FP, G ........................ Cool, ≤6 °C 18, H2SO4 to pH<2 ......... 28 days. 

53. Residue, total ............................................. P, FP, G ........................ Cool, ≤6 °C 18 ..................................... 7 days. 

54. Residue, Filterable ..................................... P, FP, G ........................ Cool, ≤6 °C 18 ..................................... 7 days. 

55. Residue, Nonfilterable (TSS) ..................... P, FP, G ........................ Cool, ≤6 °C 18 ..................................... 7 days. 

56. Residue, Settleable .................................... P, FP, G ........................ Cool, ≤6 °C 18 ..................................... 48 hours. 

57. Residue, Volatile ........................................ P, FP, G ........................ Cool, ≤6 °C 18 ..................................... 7 days. 

61. Silica ........................................................... P or Quartz .................... Cool, ≤6 °C 18 ..................................... 28 days. 

64. Specific conductance ................................. P, FP, G ........................ Cool, ≤6 °C 18 ..................................... 28 days. 

65. Sulfate ........................................................ P, FP, G ........................ Cool, ≤6 °C 18 ..................................... 28 days. 

66. Sulfide ......................................................... P, FP, G ........................ Cool, ≤6 °C 18, add zinc acetate plus 

sodium hydroxide to pH>9.

7 days. 

67. Sulfite .......................................................... P, FP, G ........................ None required .................................... Analyze within 15 minutes. 

68. Surfactants ................................................. P, FP, G ........................ Cool, ≤6 °C 18 ..................................... 48 hours. 

69. Temperature ............................................... P, FP, G ........................ None required .................................... Analyze. 

73. Turbidity ...................................................... P, FP, G ........................ Cool, ≤6 °C 18 ..................................... 48 hours. 

Table lC—Organic Tests 8


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Federal Register / Vol. 72, No. 57 / Monday, March 26, 2007 / Rules and Regulations 14231 

TABLE II.—REQUIRED CONTAINERS, PRESERVATION TECHNIQUES, AND HOLDING TIMES—Continued 

Parameter No./name Container 1 Preservation 2,3 Maximum holding time 4

13, 18–20, 22, 24–28, 34–37, 39–43, 45–47, 

56, 76, 104, 105, 108–111, 113. Purgeable 

Halocarbons.

G, FP-lined septum ....... Cool, ≤6 °C 18, 0.008% Na2S2O3 5 .... 14 days. 

6, 57, 106. Purgeable aromatic hydrocarbons G, FP-lined septum ....... Cool, ≤6 °C 18, 0.008% Na2S2O3 5, 

HCl to pH 2 9.

14 days.9

3, 4. Acrolein and acrylonitrile .......................... G, FP-lined septum ....... Cool, ≤6 °C 18, 0.008% Na2S2O3 5, 

pH to 4–5 10.

14 days.10

23, 30, 44, 49, 53, 77, 80, 81, 98, 100, 112. 

Phenols 11.

G, FP-lined cap ............. Cool, ≤6 °C 18, 0.008% Na2S2O3 5 .... 7 days until extraction, 

40 days after extraction. 

7, 38. Benzidines 11, 12 ..................................... G, FP-lined cap ............. Cool, ≤6 °C 18, 0.008% Na2S2O3 5 .... 7 days until extraction.13

14, 17, 48, 50–52. Phthalate esters 11 ............. G, FP-lined cap ............. Cool, ≤6 °C 18 ..................................... 7 days until extraction, 

40 days after extraction. 

82–84. Nitrosamines 11, 14 ................................ G, FP-lined cap ............. Cool, ≤6 °C 18, store in dark, 0.008% 

Na2S2O3 5.

7 days until extraction, 

40 days after extraction. 

88–94. PCBs 11 ................................................. G, FP-lined cap ............. Cool, ≤6 °C 18 ..................................... 1 year until extraction, 1 

year after extraction. 

54, 55, 75, 79. Nitroaromatics and 

isophorone 11.

G, FP-lined cap ............. Cool, ≤6 °C 18, store in dark, 0.008% 

Na2S2O3 5.

7 days until extraction, 

40 days after extraction. 

1, 2, 5, 8–12, 32, 33, 58, 59, 74, 78, 99, 101. 

Polynuclear aromatic hydrocarbons 11.

G, FP-lined cap ............. Cool, ≤6 °C 18, store in dark, 0.008% 

Na2S2O3 5.

7 days until extraction, 

40 days after extraction. 

15, 16, 21, 31, 87. Haloethers 11 ...................... G, FP-lined cap ............. Cool, ≤6 °C 18, 0.008% Na2S2O3 5 .... 7 days until extraction, 

40 days after extraction. 

29, 35–37, 63–65, 107. Chlorinated hydrocarbons 11.G, FP-lined cap ............. Cool, ≤6 °C 18 ..................................... 7 days until extraction, 

40 days after extraction. 

60–62, 66–72, 85, 86, 95–97, 102, 103. 

CDDs/CDFs 11.

Aqueous Samples: Field and Lab Preservation G .................................... Cool, ≤6 °C 18, 0.008% Na2S2O3 5, 

pH<9.

1 year. 

Solids and Mixed-Phase Samples: Field Preservation.G .................................... Cool, ≤6 °C 18 ..................................... 7 days. 

Tissue Samples: Field Preservation ................. G .................................... Cool, ≤6 °C 18 ..................................... 24 hours. 

Solids, Mixed-Phase, and Tissue Samples: 

Lab Preservation.

G .................................... Freeze, ≤¥10 °C ............................... 1 year. 

Table lD—Pesticides Tests: 

1–70. Pesticides 11 ........................................... G, FP-lined cap ............. Cool, ≤6 °C 18, pH 5–9 15 ................... 7 days until extraction, 

40 days after extraction. 

Table IE—Radiological Tests: 

1–5. Alpha, beta, and radium ........................... P, FP, G ........................ HNO3 to pH<2 ................................... 6 months. 

Table IH—Bacterial Tests: 

1. E. coli ............................................................ PA, G ............................. Cool, <10 °C, 0.0008% Na2S2O3 5 .... 6 hours.22

2. Enterococci ................................................... PA, G ............................. Cool, <10 °C, 0.0008% Na2S2O3 5 .... 6 hours.22

Table IH—Protozoan Tests: 

8. Cryptosporidium ........................................... LDPE; field filtration ....... 0–8 °C ................................................ 96 hours.21

9. Giardia .......................................................... LDPE; field filtration ....... 0–8 °C ................................................ 96 hours.21

1 ‘‘P’’ is polyethylene; ‘‘FP’’ is fluoropolymer (polytetrafluoroethylene (PTFE; Teflon), or other fluoropolymer, unless stated otherwise in this 

Table II; ‘‘G’’ is glass; ‘‘PA’’ is any plastic that is made of a sterlizable material (polypropylene or other autoclavable plastic); ‘‘LDPE’’ is low density polyethylene. 2Except where noted in this Table II and the method for the parameter, preserve each grab sample within 15 minutes of collection. For a composite sample collected with an automated sampler (e.g., using a 24-hour composite sampler; see 40 CFR 122.21(g)(7)(i) or 40 CFR Part 403, 

Appendix E), refrigerate the sample at ≤6 °C during collection unless specified otherwise in this Table II or in the method(s). For a composite 

sample to be split into separate aliquots for preservation and/or analysis, maintain the sample at ≤6 °C, unless specified otherwise in this Table II 

or in the method(s), until collection, splitting, and preservation is completed. Add the preservative to the sample container prior to sample collection when the preservative will not compromise the integrity of a grab sample, a composite sample, or an aliquot split from a composite sample; 

otherwise, preserve the grab sample, composite sample, or aliquot split from a composite sample within 15 minutes of collection. If a composite 

measurement is required but a composite sample would compromise sample integrity, individual grab samples must be collected at prescribed 

time intervals (e.g., 4 samples over the course of a day, at 6-hour intervals). Grab samples must be analyzed separately and the concentrations 

averaged. Alternatively, grab samples may be collected in the field and composited in the laboratory if the compositing procedure produces results equivalent to results produced by arithmetic averaging of the results of analysis of individual grab samples. For examples of laboratory 

compositing procedures, see EPA Method 1664A (oil and grease) and the procedures at 40 CFR 141.34(f)(14)(iv) and (v) (volatile organics). 


cprice-sewell on PROD1PC66 with RULES2
14232 Federal Register / Vol. 72, No. 57 / Monday, March 26, 2007 / Rules and Regulations 

3When any sample is to be shipped by common carrier or sent via the U.S. Postal Service, it must comply with the Department of Transportation Hazardous Materials Regulations (49 CFR Part 172). The person offering such material for transportation is responsible for ensuring such 

compliance. For the preservation requirements of Table II, the Office of Hazardous Materials, Materials Transportation Bureau, Department of 

Transportation has determined that the Hazardous Materials Regulations do not apply to the following materials: Hydrochloric acid (HCl) in water 

solutions at concentrations of 0.04% by weight or less (pH about 1.96 or greater); Nitric acid (HNO3) in water solutions at concentrations of 

0.15% by weight or less (pH about 1.62 or greater); Sulfuric acid (H2SO4) in water solutions at concentrations of 0.35% by weight or less (pH 

about 1.15 or greater); and Sodium hydroxide (NaOH) in water solutions at concentrations of 0.080% by weight or less (pH about 12.30 or less). 4Samples should be analyzed as soon as possible after collection. The times listed are the maximum times that samples may be held before 

the start of analysis and still be considered valid (e.g., samples analyzed for fecal coliforms may be held up to 6 hours prior to commencing analysis). Samples may be held for longer periods only if the permittee or monitoring laboratory has data on file to show that, for the specific types of 

samples under study, the analytes are stable for the longer time, and has received a variance from the Regional Administrator under § 136.3(e). 

For a grab sample, the holding time begins at the time of collection. For a composite sample collected with an automated sampler (e.g., using a 

24-hour composite sampler; see 40 CFR 122.21(g)(7)(i) or 40 CFR Part 403, Appendix E), the holding time begins at the time of the end of collection of the composite sample. For a set of grab samples composited in the field or laboratory, the holding time begins at the time of collection 

of the last grab sample in the set. Some samples may not be stable for the maximum time period given in the table. A permittee or monitoring 

laboratory is obligated to hold the sample for a shorter time if it knows that a shorter time is necessary to maintain sample stability. See 

§ 136.3(e) for details. The date and time of collection of an individual grab sample is the date and time at which the sample is collected. For a 

set of grab samples to be composited, and that are all collected on the same calendar date, the date of collection is the date on which the samples are collected. For a set of grab samples to be composited, and that are collected across two calendar dates, the date of collection is the 

dates of the two days; e.g., November 14–15. For a composite sample collected automatically on a given date, the date of collection is the date 

on which the sample is collected. For a composite sample collected automatically, and that is collected across two calendar dates, the date of 

collection is the dates of the two days; e.g., November 14–15. 5Add a reducing agent only if an oxidant (e.g., chlorine) is present. Reducing agents shown to be effective are sodium thiosulfate (Na2S2O3), 

ascorbic acid, sodium arsenite (NaAsO2), or sodium borohydride (NaBH4). However, some of these agents have been shown to produce a positive or negative cyanide bias, depending on other substances in the sample and the analytical method used. Therefore, do not add an excess of 

reducing agent. Methods recommending ascorbic acid (e.g., EPA Method 335.4) specify adding ascorbic acid crystals, 0.1–0.6 g, until a drop of 

sample produces no color on potassium iodide (KI) starch paper, then adding 0.06 g (60 mg) for each liter of sample volume. If NaBH4 or 

NaAsO2 is used, 25 mg/L NaBH4 or 100 mg/L NaAsO2 will reduce more than 50 mg/L of chlorine (see method ‘‘Kelada-01’’ and/or Standard 

Method 4500–CN¥ for more information). After adding reducing agent, test the sample using KI paper, a test strip (e.g. for chlorine, SenSafeTM

Total Chlorine Water Check 480010) moistened with acetate buffer solution (see Standard Method 4500–Cl.C.3e), or a chlorine/oxidant test 

method (e.g., EPA Method 330.4 or 330.5), to make sure all oxidant is removed. If oxidant remains, add more reducing agent. Whatever agent is 

used, it should be tested to assure that cyanide results are not affected adversely. 6Sample collection and preservation: Collect a volume of sample appropriate to the analytical method in a bottle of the material specified. If 

the sample can be analyzed within 48 hours and sulfide is not present, adjust the pH to > 12 with sodium hydroxide solution (e.g., 5% w/v), refrigerate as specified, and analyze within 48 hours. Otherwise, to extend the holding time to 14 days and mitigate interferences, treat the sample 

immediately using any or all of the following techniques, as necessary, followed by adjustment of the sample pH to > 12 and refrigeration as 

specified. There may be interferences that are not mitigated by approved procedures. Any procedure for removal or suppression of an interference may be employed, provided the laboratory demonstrates that it more accurately measures cyanide. Particulate cyanide (e.g., ferric ferrocyanide) or a strong cyanide complex (e.g., cobalt cyanide) are more accurately measured if the laboratory holds the sample at room temperature and pH > 12 for a minimum of 4 hours prior to analysis, and performs UV digestion or dissolution under alkaline (pH=12) conditions, if necessary. 

(1) Sulfur: To remove elemental sulfur (S8), filter the sample immediately. If the filtration time will exceed 15 minutes, use a larger filter or a 

method that requires a smaller sample volume (e.g., EPA Method 335.4 or Lachat Method 01). Adjust the pH of the filtrate to > 12 with NaOH, 

refrigerate the filter and filtrate, and ship or transport to the laboratory. In the laboratory, extract the filter with 100 mL of 5% NaOH solution for a 

minimum of 2 hours. Filter the extract and discard the solids. Combine the 5% NaOH-extracted filtrate with the initial filtrate, lower the pH to approximately 12 with concentrated hydrochloric or sulfuric acid, and analyze the combined filtrate. Because the detection limit for cyanide will be 

increased by dilution by the filtrate from the solids, test the sample with and without the solids procedure if a low detection limit for cyanide is 

necessary. Do not use the solids procedure if a higher cyanide concentration is obtained without it. Alternatively, analyze the filtrates from the 

sample and the solids separately, add the amounts determined (in μg or mg), and divide by the original sample volume to obtain the cyanide 

concentration. 

(2) Sulfide: If the sample contains sulfide as determined by lead acetate paper, or if sulfide is known or suspected to be present, immediately 

conduct one of the volatilization treatments or the precipitation treatment as follows: Volatilization—Headspace expelling. In a fume hood or wellventilated area, transfer 0.75 liter of sample to a 4.4 L collapsible container (e.g., CubitainerTM). Acidify with concentrated hydrochloric acid to pH 

< 2. Cap the container and shake vigorously for 30 seconds. Remove the cap and expel the headspace into the fume hood or open area by collapsing the container without expelling the sample. Refill the headspace by expanding the container. Repeat expelling a total of five headspace 

volumes. Adjust the pH to > 12, refrigerate, and ship or transport to the laboratory. Scaling to a smaller or larger sample volume must maintain 

the air to sample volume ratio. A larger volume of air will result in too great a loss of cyanide (> 10%). Dynamic stripping: In a fume hood or wellventilated area, transfer 0.75 liter of sample to a container of the material specified and acidify with concentrated hydrochloric acid to pH < 2. 

Using a calibrated air sampling pump or flowmeter, purge the acidified sample into the fume hood or open area through a fritted glass aerator at 

a flow rate of 2.25 L/min for 4 minutes. Adjust the pH to > 12, refrigerate, and ship or transport to the laboratory. Scaling to a smaller or larger 

sample volume must maintain the air to sample volume ratio. A larger volume of air will result in too great a loss of cyanide (> 10%). Precipitation: If the sample contains particulate matter that would be removed by filtration, filter the sample prior to treatment to assure that cyanide associated with the particulate matter is included in the measurement. Ship or transport the filter to the laboratory. In the laboratory, extract the filter 

with 100 mL of 5% NaOH solution for a minimum of 2 hours. Filter the extract and discard the solids. Combine the 5% NaOH-extracted filtrate 

with the initial filtrate, lower the pH to approximately 12 with concentrated hydrochloric or sulfuric acid, and analyze the combined filtrate. Because the detection limit for cyanide will be increased by dilution by the filtrate from the solids, test the sample with and without the solids procedure if a low detection limit for cyanide is necessary. Do not use the solids procedure if a higher cyanide concentration is obtained without it. Alternatively, analyze the filtrates from the sample and the solids separately, add the amounts determined (in μg or mg), and divide by the original 

sample volume to obtain the cyanide concentration. For removal of sulfide by precipitation, raise the pH of the sample to > 12 with NaOH solution, then add approximately 1 mg of powdered cadmium chloride for each mL of sample. For example, add approximately 500 mg to a 500-mL 

sample. Cap and shake the container to mix. Allow the precipitate to settle and test the sample with lead acetate paper. If necessary, add cadmium chloride but avoid adding an excess. Finally, filter through 0.45 micron filter. Cool the sample as specified and ship or transport the filtrate 

and filter to the laboratory. In the laboratory, extract the filter with 100 mL of 5% NaOH solution for a minimum of 2 hours. Filter the extract and 

discard the solids. Combine the 5% NaOH-extracted filtrate with the initial filtrate, lower the pH to approximately 12 with concentrated hydrochloric or sulfuric acid, and analyze the combined filtrate. Because the detection limit for cyanide will be increased by dilution by the filtrate from 

the solids, test the sample with and without the solids procedure if a low detection limit for cyanide is necessary. Do not use the solids procedure 

if a higher cyanide concentration is obtained without it. Alternatively, analyze the filtrates from the sample and the solids separately, add the 

amounts determined (in μg or mg), and divide by the original sample volume to obtain the cyanide concentration. If a ligand-exchange method is 

used (e.g., ASTM D6888), it may be necessary to increase the ligand-exchange reagent to offset any excess of cadmium chloride. 

(3) Sulfite, thiosulfate, or thiocyanate: If sulfite, thiosulfate, or thiocyanate is known or suspected to be present, use UV digestion with a glass 

coil (Method Kelada-01) or ligand exchange (Method OIA–1677) to preclude cyanide loss or positive interference. 

(4) Aldehyde: If formaldehyde, acetaldehyde, or another water-soluble aldehyde is known or suspected to be present, treat the sample with 20 

mL of 3.5% ethylenediamine solution per liter of sample. 


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Federal Register / Vol. 72, No. 57 / Monday, March 26, 2007 / Rules and Regulations 14233 

(5) Carbonate: Carbonate interference is evidenced by noticeable effervescence upon acidification in the distillation flask, a reduction in the pH 

of the absorber solution, and incomplete cyanide spike recovery. When significant carbonate is present, adjust the pH to ≥12 using calcium hydroxide instead of sodium hydroxide. Allow the precipitate to settle and decant or filter the sample prior to analysis (also see Standard Method 

4500–CN.B.3.d). 

(6) Chlorine, hypochlorite, or other oxidant: Treat a sample known or suspected to contain chlorine, hypochlorite, or other oxidant as directed 

in footnote 5. 7For dissolved metals, filter grab samples within 15 minutes of collection and before adding preservatives. For a composite sample collected 

with an automated sampler (e.g., using a 24-hour composite sampler; see 40 CFR 122.21(g)(7)(i) or 40 CFR Part 403, Appendix E), filter the 

sample within 15 minutes after completion of collection and before adding preservatives. If it is known or suspected that dissolved sample integrity will be compromised during collection of a composite sample collected automatically over time (e.g., by interchange of a metal between dissolved and suspended forms), collect and filter grab samples to be composited (footnote 2) in place of a composite sample collected automatically. 8Guidance applies to samples to be analyzed by GC, LC, or GC/MS for specific compounds. 

9 If the sample is not adjusted to pH 2, then the sample must be analyzed within seven days of sampling. 

10The pH adjustment is not required if acrolein will not be measured. Samples for acrolein receiving no pH adjustment must be analyzed within 3 days of sampling. 11 When the extractable analytes of concern fall within a single chemical category, the specified preservative and maximum holding times 

should be observed for optimum safeguard of sample integrity (i.e., use all necessary preservatives and hold for the shortest time listed). When 

the analytes of concern fall within two or more chemical categories, the sample may be preserved by cooling to ≤6 °C, reducing residual chlorine 

with 0.008% sodium thiosulfate, storing in the dark, and adjusting the pH to 6–9; samples preserved in this manner may be held for seven days 

before extraction and for forty days after extraction. Exceptions to this optional preservation and holding time procedure are noted in footnote 5 

(regarding the requirement for thiosulfate reduction), and footnotes 12, 13 (regarding the analysis of benzidine). 12 If 1,2-diphenylhydrazine is likely to be present, adjust the pH of the sample to 4.0 ± 0.2 to prevent rearrangement to benzidine. 13Extracts may be stored up to 30 days at < 0 °C. 14For the analysis of diphenylnitrosamine, add 0.008% Na2S2O3 and adjust pH to 7–10 with NaOH within 24 hours of sampling. 15The pH adjustment may be performed upon receipt at the laboratory and may be omitted if the samples are extracted within 72 hours of collection. For the analysis of aldrin, add 0.008% Na2S2O3. 16Sufficient ice should be placed with the samples in the shipping container to ensure that ice is still present when the samples arrive at the 

laboratory. However, even if ice is present when the samples arrive, it is necessary to immediately measure the temperature of the samples and 

confirm that the preservation temperature maximum has not been exceeded. In the isolated cases where it can be documented that this holding 

temperature cannot be met, the permittee can be given the option of on-site testing or can request a variance. The request for a variance should 

include supportive data which show that the toxicity of the effluent samples is not reduced because of the increased holding temperature. 17Samples collected for the determination of trace level mercury (<100 ng/L) using EPA Method 1631 must be collected in tightly-capped 

fluoropolymer or glass bottles and preserved with BrCl or HCl solution within 48 hours of sample collection. The time to preservation may be extended to 28 days if a sample is oxidized in the sample bottle. A sample collected for dissolved trace level mercury should be filtered in the laboratory within 24 hours of the time of collection. However, if circumstances preclude overnight shipment, the sample should be filtered in a designated clean area in the field in accordance with procedures given in Method 1669. If sample integrity will not be maintained by shipment to and 

filtration in the laboratory, the sample must be filtered in a designated clean area in the field within the time period necessary to maintain sample 

integrity. A sample that has been collected for determination of total or dissolved trace level mercury must be analyzed within 90 days of sample 

collection. 18Aqueous samples must be preserved at ≤6 °C, and should not be frozen unless data demonstrating that sample freezing does not adversely 

impact sample integrity is maintained on file and accepted as valid by the regulatory authority. Also, for purposes of NPDES monitoring, the 

specification of ‘‘≤°C’’ is used in place of the ‘‘4 °C’’ and ‘‘< 4 °C’’ sample temperature requirements listed in some methods. It is not necessary 

to measure the sample temperature to three significant figures (1⁄100th of 1 degree); rather, three significant figures are specified so that rounding 

down to 6 °C may not be used to meet the ≤6 °C requirement. The preservation temperature does not apply to samples that are analyzed immediately (less than 15 minutes). 19An aqueous sample may be collected and shipped without acid preservation. However, acid must be added at least 24 hours before analysis to dissolve any metals that adsorb to the container walls. If the sample must be analyzed within 24 hours of collection, add the acid immediately (see footnote 2). Soil and sediment samples do not need to be preserved with acid. The allowances in this footnote supersede the preservation and holding time requirements in the approved metals methods. 20To achieve the 28-day holding time, use the ammonium sulfate buffer solution specified in EPA Method 218.6. The allowance in this footnote supersedes preservation and holding time requirements in the approved hexavalent chromium methods, unless this supersession would 

compromise the measurement, in which case requirements in the method must be followed. 21 Holding time is calculated from time of sample collection to elution for samples shipped to the laboratory in bulk and calculated from the time 

of sample filtration to elution for samples filtered in the field. 22Samples analysis should begin immediately, preferably within 2 hours of collection. The maximum transport time to the laboratory is 6 hours, 

and samples should be processed within 2 hours of receipt at the laboratory. 23For fecal coliform samples for sewage sludge (biosolids) only, the holding time is extended to 24 hours for the following sample types using 

either EPA Method 1680 (LTB–EC) or 1681 (A–1): Class A composted, Class B aerobically digested, and Class B anaerobically digested. 

PART 503—STANDARDS FOR THE 

USE OR DISPOSAL OF SEWAGE 

SLUDGE 

■ 3. The authority citation for Part 503 

continues to read as follows: 

Authority: Secs. 405(d) and (e) of the Clean 

Water Act, as amended by Pub. L. 95–217, 

sec. 54(d), 91 Stat. 1591 (33 U.S.C. 1345(d) 

and (e)); and Pub. L. 100–4, title IV, sec. 

406(a), (b), 101 Stat., 71, 72 (33 U.S.C. 1251 

et seq.). 

■ 4. Section 503.8 is amended by 

revising paragraph (b) introductory text 

to read as follows: 

§ 503.8 Sampling and analysis. 

* * * * * 

(b) Methods. The materials listed 

below are incorporated by reference in 

this part. These incorporations by 

reference were approved by the Director 

of the Federal Register in accordance 

with 5 U.S.C. 552(a) and 1 CFR part 51. 

The materials are incorporated as they 

exist on the date of approval, and notice 

of any change in these materials will be 

published in the Federal Register. They 

are available for inspection at the HQ 

Water Docket Center, EPA/DC, EPA 

West, Room B102, 1301 Constitution 

Ave., NW., Washington, DC, and at the 

National Archives and Records 

Administration (NARA). For 

information on the availability of this 

material at NARA, call 202–741–6030, 

or go to: http://www.archives.gov/ 

federal_register/ 

code_of_federal_regulations/ 

ibr_locations.html. 

Copies may be obtained from the 

standard producer or publisher listed in 

the regulation. The methods in the 

materials listed below (or in 40 CFR Part 

136) shall be used to analyze samples of 

sewage sludge. 

* * * * * 

[FR Doc. 07–1455 Filed 3–23–07; 8:45 am] 

BILLING CODE 6560–50–P 


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