27020 Federal Register / Vol. 66, No. 95 / Wednesday, May 16, 2001 / Rules and Regulations

Federal Register, 800 North Capitol Street,

NW., suite 700, Washington, DC.

Effective Date of This AD

(h) This amendment becomes effective on

May 31, 2001.

Issued in Burlington, Massachusetts, on

May 7, 2001.

Francis A. Favara,

Acting Manager, Engine and Propeller

Directorate, Aircraft Certification Service.

[FR Doc. 01–12006 Filed 5–15–01; 8:45 am]

BILLING CODE 4910–13–U

DEPARTMENT OF HEALTH AND

HUMAN SERVICES

Food and Drug Administration

21 CFR Part 173

[Docket No. 92F–0396]

Secondary Direct Food Additives

Permitted in Food for Human

Consumption; Alpha-Acetolactate

Decarboxylase Enzyme Preparation

AGENCY: Food and Drug Administration,

HHS.

ACTION: Final rule.

SUMMARY: The Food and Drug

Administration (FDA) is amending the

food additive regulations to provide for

the safe use of alpha-acetolactate

decarboxylase (α-ALDC) enzyme

preparation derived from Bacillus

subtilis, modified by recombinant

deoxyribonucleic acid (DNA)

techniques to contain the gene coding

for α-ALDC from B. brevis, for use as a

processing aid to produce alcoholic malt

beverages and distilled liquors. This

action is in response to a petition filed

by Novozymes North America, Inc.

(formerly Novo Nordisk Bioindustrials,

Inc.).

DATES: This rule is effective May 16,

2001. Submit written objections and

requests for a hearing by June 15, 2001.

The Director of the Office of the Federal

Register approves the incorporation by

reference in accordance with 5 U.S.C.

552(a) and 1 CFR part 51 of a certain

publication in § 173.115(b)(3), effective

as of May 16, 2001.

ADDRESSES: Submit written objections to

the Dockets Management Branch (HFA–

305), Food and Drug Administration,

5630 Fishers Lane, rm. 1061, Rockville,

MD 20852.

FOR FURTHER INFORMATION CONTACT:

Rudaina H. Alrefai, Center for Food

Safety and Applied Nutrition (HFS–

206), Food and Drug Administration,

200 C St. SW., Washington, DC 20204,

202–418–3034.

SUPPLEMENTARY INFORMATION: In a notice

published in the Federal Register of

November 30, 1992 (57 FR 56585), FDA

announced that a food additive petition

(FAP 2A4345) had been filed by Novo

Nordisk Bioindustrials, Inc., later

renamed Novozymes North America,

Inc., 77 Perry Chapel Church Rd., P.O.

Box 576, Franklinton, NC 27525. The

petition proposed that the food additive

regulations be amended to provide for

the safe use of α-acetolactate

decarboxylase (ALDC) derived from B.

subtilis modified by recombinant DNA

techniques to contain the gene coding

for ALDC from B. brevis for use as a

processing aid in the brewing and

alcohol industries.

When FDA filed the petition in the

Federal Register of November 30, 1992

(57 FR 56585), it contained an

environmental assessment (EA). The

notice of filing stated ‘‘ * * * if the

agency finds that an environmental

impact statement is not required and

this petition results in a regulation, the

notice of availability of the agency’s

finding of no significant impact and the

evidence supporting that finding will be

published with the regulation * * *.’’ In

the Federal Register of July 29, 1997 (62

FR 40570), FDA published a final rule

on its National Environmental Policy

Act policies and procedures, which

became effective on August 28, 1997. In

a letter dated January 4, 2001, the

petitioner submitted a claim of

categorical exclusion under 21 CFR

25.32(k). The agency has reviewed the

claim of categorical exclusion and has

concluded that it is warranted.

I. Evaluation of Safety of the Petitioned

Use of the Additive

A. Introduction

The use of α-ALDC enzyme

preparation from B. subtilis is to prevent

the formation of diacetyl that causes

unpleasant taste in beer and other

alcoholic beverages. The enzyme αALDC is to be distinguished from the αALDC enzyme preparation, which

contains α-ALDC as the principal active

component in addition to other

components derived from the

production organism and fermentation

media. This document will refer to the

former as ‘‘α-ALDC’’ and the latter as

‘‘α-ALDC enzyme preparation.’’ Diacetyl

is normally formed from α-acetolactate

during fermentation. Alpha-ALDC,

which is the active component of the

petitioned enzyme preparation,

catalyzes the conversion of αacetolactate directly to acetoin, thereby

reducing the time needed for

spontaneous degradation of diacetyl to

acetoin.

B. Host Organism

The host organism, B. subtilis, for

production of α-ALDC is widely

distributed in nature and is commonly

present in foods eaten by both humans

and animals. It also has a history of safe

use as a source of enzymes in food

enzyme manufacturing industry prior to

1958. Thus, B. subtilis is considered to

be a nonpathogenic microorganism.

C. Donor Organism

B. brevis is the microorganism used as

the source of the genetic material for the

α-ALDC enzyme that is the subject of

FAP 2A4345. FDA reviewed the safety

of the DNA that encodes the enzyme αALDC from B. brevis and the enzyme it

produces (discussed below), because

only that DNA is transferred to the host

strain from the donor organism.

D. Production Organism

The petitioner provided information

demonstrating that the plasmid carrying

the gene for α-ALDC is stably integrated

into the chromosome of B. subtilis

production strain. The petitioner

conducted a study to evaluate the

pathogenicity of three B. subtilis strains.

In this study, mice received an

intraperitoneal injection with the B.

subtilis host strain, B. subtilis

production strain, and a B. subtilis

strain capable of producing α-ALDC but

not used as a source of the petitioned

enzyme preparation. FDA reviewed this

study as well as the scientific literature

concerning potential pathogenicity of B.

subtilis and did not identify any

microbiological concern (Refs. 1, 2, and

3).

E. Enzyme Preparation

The α-ALDC enzyme preparation is

manufactured by a submerged pure

culture fermentation of a genetically

engineered strain of B. subtilis carrying

the B. brevis gene that encodes α-ALDC.

The enzyme is secreted to the

fermentation broth and processed by

removing the cellular debris, followed

by concentration and formulation. For

certain applications, the α-ALDC

enzyme preparation is stabilized by

crosslinking with glutaraldehyde

(referred to as d-ALDC).

The petitioner submitted several

toxicological studies that address the

safety of the petitioned α-ALDC and dALDC enzyme preparations. These

include: A teratogenicity study in rats

and genotoxicity studies, including tests

for mutagenic activity in Salmonella

typhimurium and mammalian cells, as

well as tests for chromosome-damaging

activity in human lymphocytes. FDA

has reviewed these studies and

concludes that the petitioned α-ALDC

Federal Register / Vol. 66, No. 95 / Wednesday, May 16, 2001 / Rules and Regulations 27021

enzyme preparation does not raise any

toxicity concerns at the expected level

of consumption or have any mutagenic

potential (Refs. 4 and 5).

F. Source of Impurities

Enzyme preparations used in food are

usually not chemically pure, but contain

cellular and processing material. The

nature and amounts of these impurities

in the finished enzyme preparation

depend on the organism from which the

enzyme preparation is produced (the

production organism), the fermentation

materials and methods used to grow the

production organism, and the materials

and methods used to generate the

finished enzyme preparation. Thus, the

question is whether the production

organism or the manufacturing methods

used to grow the production organism

or to generate the finished enzyme

preparation from recombinant B. subtilis

will introduce impurities that raise

concerns about the safety of the enzyme

preparation.

One issue raised by the use of

recombinant DNA techniques is the

potential transfer of DNA encoding for

extraneous proteins along with the gene

of interest (i.e., α-ALDC), thereby

contaminating the enzyme preparation.

As a matter of current good

manufacturing practice, manufacturers

using recombinant DNA technology

must ensure that they have not

inadvertently cloned extraneous

protein-encoding DNA along with the αALDC gene that may lead to

contamination of the α-ALDC enzyme

preparation. Such assurance can come

from reviewing the details of the cloning

steps, which include the origin and

sequence of the DNA fragments used in

the cloning, and full characterization of

the final genetic constructs via

techniques such as DNA sequencing.

The petition contains information

demonstrating that the petitioner

evaluated the cloning process to ensure

that the final cloning product, i.e., the

DNA with the α-ALDC gene, used in the

development of the recombinant B.

subtilis was accurately constructed. The

petitioner submitted evidence to

demonstrate that it cloned a full-length

copy of the α-ALDC gene from B. brevis

into B. subtilis. The petitioner also

described the multistep process for

constructing the B. subtilis production

strain. These steps involve the use of

several plasmids (intermediate

plasmids) that confer resistance to

chloramphenicol and kanamycin, both

of which are clinically useful

antibiotics. Through various techniques,

these plasmids are eliminated during

the construction of the gene encoding αALDC. The petitioner tested the final

enzyme preparation for the presence of

the production strain or other microbial

activity and for antibacterial activity

and reported none present. Therefore,

the agency concludes that the

production strain is effectively removed

by the enzyme purification procedure.

Furthermore, the α-ALDC enzyme

preparation conforms to the general and

additional requirements for enzyme

preparations in the Food Chemicals

Codex (Ref. 6) and does not contain the

production organism or antimicrobial

activity.

FDA concludes that, when the αALDC enzyme preparation is

manufactured in conformity with

§ 173.115, there is no basis for concern

regarding the possibility that the αALDC enzyme preparation will be

contaminated by the products of

extraneous genetic material inserted

along with the α-ALDC gene in B.

subtilis (Ref. 1). Furthermore, FDA

concludes, having considered the

evidence concerning the production

organism and the processing steps to

derive the α-ALDC enzyme preparation,

that: (1) B. subtilis containing α-ALDC

gene from B. brevis is safe for use as a

source of food-grade α-ALDC enzyme

preparation, (2) impurities resulting

from the use of B. subtilis containing αALDC gene from B. brevis in the

production of α-ALDC enzyme

preparation will not affect the safety of

the α-ALDC enzyme preparation, and (3)

processing aids and their impurities that

are used to make the commercial αALDC enzyme preparation and that may

remain in food processed with this

enzyme preparation present no safety

concerns (Refs. 5 and 7).

II. Conclusion

FDA has evaluated the data in the

petition and other relevant material.

Based on this information, the agency

concludes that: (1) The proposed use of

α-ALDC enzyme preparation from B.

subtilis containing the α-ALDC gene

from B. brevis is safe, (2) the additive

will achieve its intended technical

effect, and (3) the regulations in

§ 173.115 should be amended as set

forth below in this document.

III. Inspection of Documents

In accordance with § 171.1 (21 CFR

171.1(h)), the petition and the

documents that FDA considered and

relied upon in reaching its decision to

approve the petition are available for

inspection at the Center for Food Safety

and Applied Nutrition by appointment

with the information contact person

listed above. As provided in § 171.1(h),

the agency will delete from the

documents any materials that are not

available for public disclosure before

making the documents available for

inspection.

IV. Environmental Impact

The agency has determined under 21

CFR 25.32(k) that this action is of a type

that does not individually or

cumulatively have a significant effect on

the human environment. Therefore,

neither an EA nor an environmental

impact statement is required.

V. Paperwork Reduction Act of 1995

This final rule contains no collection

of information. Therefore, clearance by

the Office of Management and Budget

under the Paperwork Reduction Act of

1995 is not required.

VI. Objections

Any person who will be adversely

affected by this regulation may at any

time file with the Dockets Management

Branch (address above) written

objections by June 15, 2001. Each

objection shall be separately numbered,

and each numbered objection shall

specify with particularity the provisions

of the regulation to which objection is

made and the grounds for the objection.

Each numbered objection on which a

hearing is requested shall specifically so

state. Failure to request a hearing for

any particular objection shall constitute

a waiver of the right to a hearing on that

objection. Each numbered objection for

which a hearing is requested shall

include a detailed description and

analysis of the specific factual

information intended to be presented in

support of the objection in the event

that a hearing is held. Failure to include

such a description and analysis for any

particular objection shall constitute a

waiver of the right to a hearing on the

objection. Three copies of all documents

are to be submitted and are to be

identified with the docket number

found in brackets in the heading of this

document. Any objections received in

response to the regulation may be seen

in the Dockets Management Branch

between 9 a.m. and 4 p.m., Monday

through Friday.

VII. References

The following references have been

placed on display in the Dockets

Management Branch (address above)

and may be seen by interested persons

between 9 a.m. and 4 p.m., Monday

through Friday.

1. Memorandum from W. Koch, FDA, to L.

Kahl, FDA, September 23, 1993.

2. Memorandum from J. Madden, FDA, to

the Biotechnology Policy Branch, FDA,

August 29, 1995.

27022 Federal Register / Vol. 66, No. 95 / Wednesday, May 16, 2001 / Rules and Regulations

3. Memorandum from J. Madden, FDA, to

the Biotechnology Policy Branch, FDA,

October 20, 1995.

4. Memorandum from R. D. Benz, FDA, to

K. C. Raffaele, FDA, July 20, 1995.

5. Memorandum from the Division of

Health Effects Evaluation, FDA, to R. H.

Alrefai, FDA, May 4, 1999.

6. Food Chemicals Codex, 1996, 4th ed.,

National Academy Press, Washington, DC,

pp. 133–134.

7. Memorandum from the Division of

Health Effects Evaluation, FDA, to R. H.

Alrefai, FDA, August 20, 1999.

List of Subjects in 21 CFR Part 173

Food additives, Incorporation by

reference.

Therefore, under the Federal Food,

Drug, and Cosmetic Act and under

authority delegated to the Commissioner

of Food and Drugs, 21 CFR part 173 is

amended as follows:

PART 173—SECONDARY DIRECT

FOOD ADDITIVES PERMITTED IN

FOOD FOR HUMAN CONSUMPTION

1. The authority citation for 21 CFR

part 173 continues to read as follows:

Authority: 21 U.S.C. 321, 342, 348.

2. Section 173.115 is added to subpart

B to read as follows:

§ 173.115 Alpha-acetolactate

decarboxylase (α-ALDC) enzyme

preparation derived from a recombinant

Bacillus subtilis.

The food additive alpha-acetolactate

decarboxylase (α-ALDC) enzyme

preparation, may be safely used in

accordance with the following

conditions:

(a) The food additive is the enzyme

preparation derived from a modified

Bacillus subtilis strain that contains the

gene coding for α-ALDC from Bacillus

brevis.

(b)(1) The manufacturer produces the

additive from a pure culture

fermentation of a strain of Bacillus

subtilis that is nonpathogenic and

nontoxigenic in man or other animals.

(2) The manufacturer may stabilize

the enzyme preparation with

glutaraldehyde or with other suitable

approved food additives or generally

recognized as safe substances.

(3) The enzyme preparation must

meet the general and additional

requirements for enzyme preparations

in the Food Chemicals Codex, 4th ed.,

1996, pp. 133–134, which is

incorporated by reference. The Director

of the Office of the Federal Register

approves this incorporation by reference

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

1 CFR part 51. Copies may be obtained

from the National Academy Press, 2101

Constitution Ave. NW., Washington, DC

20055, or may be examined at the

Center for Food Safety and Applied

Nutrition, 200 C St. SW., rm. 3321,

Washington, DC, or at the Office of the

Federal Register, 800 North Capitol St.

NW., suite 700, Washington, DC.

(c) The additive is used in an amount

not in excess of the minimum required

to produce its intended effect as a

processing aid in the production of

alcoholic malt beverages and distilled

liquors.

Dated: May 4, 2001.

Margaret M. Dotzel,

Associate Commissioner for Policy.

[FR Doc. 01–12225 Filed 5–15–01; 8:45 am]

BILLING CODE 4160–01–F

DEPARTMENT OF HEALTH AND

HUMAN SERVICES

Food and Drug Administration

21 CFR Part 558

New Animal Drugs For Use in Animal

Feeds; Narasin, Nicarbazin, and

Bambermycins

AGENCY: Food and Drug Administration,

HHS.

ACTION: Final rule.

SUMMARY: The Food and Drug

Administration (FDA) is amending the

animal drug regulations to reflect

approval of a new animal drug

application (NADA) filed by Elanco

Animal Health. The NADA provides for

use of approved narasin/nicarbazin and

bambermycins Type A medicated

articles to make three-way combination

Type C medicated feeds used for

prevention of coccidiosis, increased rate

of weight gain, and improved feed

efficiency in broiler chickens.

DATES: This rule is effective May 16,

2001.

FOR FURTHER INFORMATION CONTACT:

Charles J. Andres, Center for Veterinary

Medicine (HFV–128), Food and Drug

Administration, 7500 Standish Pl.,

Rockville, MD 20855, 301–827–1600.

SUPPLEMENTARY INFORMATION: Elanco

Animal Health, A Division of Eli Lilly

& Co., Lilly Corporate Center,

Indianapolis, IN 46285, filed NADA

140-942 that provides for use of

Maxiban (36 grams per pound (g/lb)

each of narasin and nicarbazin) and

Flavomycin (2, 4, or 10 g/lb of

bambermycins activity) Type A

medicated articles to make three-way

combination Type C medicated feeds for

use in broiler chickens. The

combination Type C medicated feeds

contain 27 to 45 g/ton narasin and 27 to

45 g/ton nicarbazin (in a fixed 1:1 ratio)

and 1 to 2 g/ton bambermycins, and are

used for prevention of coccidiosis

caused by Eimeria tenella, E. necatrix,

E. acervulina, E. brunetti, and E. mivati,

and for increased rate of weight gain

and improved feed efficiency. The

NADA is approved as of March 8, 2001,

and the regulations are amended in 21

CFR 558.95 and 558.363 to reflect the

approval. The basis of approval is

discussed in the freedom of information

summary

In accordance with the freedom of

information provisions of 21 CFR part

20 and 514.11(e)(2)(ii), a summary of

safety and effectiveness data and

information submitted to support

approval of this application may be seen

in the Dockets Management Branch

(HFA–305), Food and Drug

Administration, 5630 Fishers Lane, rm.

1061, Rockville, MD 20852, between 9

a.m. and 4 p.m., Monday through

Friday.

The agency has determined under 21

CFR 25.33(a)(2) that this action is of a

type that does not individually or

cumulatively have a significant effect on

the human environment. Therefore,

neither an environmental assessment

nor an environmental impact statement

is required.

This rule does not meet the definition

of ‘‘rule’’ in 5 U.S.C. 804(3)(A) because

it is a rule of ‘‘particular applicability.’’

Therefore, it is not subject to the

congressional review requirements in 5

U.S.C. 801–808.

List of Subjects in 21 CFR Part 558

Animal drugs, Animal feeds.

Therefore, under the Federal Food,

Drug, and Cosmetic Act and under

authority delegated to the Commissioner

of Food and Drugs and redelegated to

the Center for Veterinary Medicine, 21

CFR part 558 is amended as follows:

PART 558—NEW ANIMAL DRUGS FOR

USE IN ANIMAL FEEDS

1. The authority citation for 21 CFR

part 558 continues to read as follows:

Authority: 21 U.S.C. 360b, 371.

§ 558.95 [Amended]

2. Section 558.95 Bambermycins is

amended in paragraph (d)(5)(iv) by

adding ‘‘nicarbazin or’’ following

‘‘with’’, and in paragraph (d)(5)(v) by

removing ‘‘Nicarbazine’’ and adding in

its place ‘‘Nicarbazin’’.

3. Section 558.363 is amended by

adding paragraph (d)(1)(xii) to read as

follows:

§ 558.363 Narasin.

* * * * *