U. S. Food and Drug Administration
Center for Food Safety and Applied Nutrition
Office of Food Additive Safety
January 27, 2003

Japan Cellfoods Co., Ltd.
Mr. Toru Akazawa, President
SORIO 3 Building, 5th floor
2-1-1, Sakaemachi, Takarazuka
Hyogo Prefecture, 665-0845
Japan

Re: GRAS Notice No. GRN 000114

Dear Dr. Akazawa:

The Food and Drug Administration (FDA) is responding to the notice, dated August 22, 2002, that you submitted in accordance with the agency's proposed regulation, proposed 21 CFR 170.36 (62 FR 18938; April 17, 1997; Substances Generally Recognized as Safe (GRAS); the GRAS proposal). FDA received the notice on August 29, 2002, filed it on September 5, 2002, and designated it as GRAS Notice No. GRN 000114.

The subject of the notice is pectate lyase enzyme preparation from Bacillus subtilis. The notice informs FDA of the view of Japan Cellfoods that the pectate lyase enzyme preparation is GRAS, through scientific procedures, for use in fruit and vegetable purees and concentrates as an enzyme at a level of 0.5-1.0 percent by weight.

Commercial enzyme preparations that are used in food processing typically contain an enzyme component, which catalyzes the chemical reaction that is responsible for its technical effect, as well as substances used as stabilizers, preservatives, or diluents. Enzyme preparations may also contain constituents derived from the production organism and constituents derived from the manufacturing process, e.g., components of the fermentation media or the residues of processing aids. Japan Cellfoods' notice provides information about each of these components in the pectate lyase enzyme preparation from B. subtilis.

Japan Cellfoods describes the activity of the pectate lyase enzyme as catalyzing the random cleavage of the alpha-1,4-glycosidic bonds in pectic acid by transelimination. This results in the formation of delta-4,5-oligogalacturonates with a 4,5-dehydrogalacturonic acid group at the non-reducing end. The pectate lyase enzyme is identified by the following classification numbers: Chemical Abstracts Service Registry No. 9015-75-2 and Enzyme Commission No. 4.2.2. The pectate lyase enzyme has the following synonyms: endopectate lyase (or transeliminase), pectic acid lyase, polygalacturonate lyase, endogalacturonate transeliminase, endo-alpha-1,4-polygalacturonic acid lyase, polygalacturonic acid lyase, and alpha-1,4-D-endopolygalacturonic acid lyase.

In assessing the safety of the enzyme itself, Japan Cellfoods cites the Food Chemicals Codex, 4th Ed. (1996) to support the conclusion that pectate lyase is one of several enzymes present in commercial pectinase products used in food processing. To support that view, Japan Cellfoods also describes the pectinase enzyme preparation derived from Aspergillus niger, which the Enzyme Technical Association (ETA) determined to be GRAS for use in hydrolysis of pectins in fruit-containing products (GRAS Notice No. GRN 000089).

In assessing the safety of the production microorganism, B. subtilis, Japan Cellfoods describes the production strain as a derivative of the well-known strain, B. subtilis strain (IFO 3134), which was obtained from the Institute of Fermentation, Osaka, Japan. IFO 3134 was subjected to a succession of mutagenic events using the chemical mutagen N-methyl-N'-nitro-N-nitrosoguanidine (NTG) and selections to improve pectate lyase production. To support its view that the resulting B. subtilis strain is safe for use in food applications, Japan Cellfoods cites FDA's previous GRAS affirmation of carbohydrase and protease enzyme preparations from B. subtilis for use as direct food ingredients (64 FR 19887-19895; April 23, 1999) and publications describing prior use of B. subtilisz in fermented bean products.

Japan Cellfoods describes the method of manufacture of the pectate lyase enzyme preparation, which is produced by submerged, controlled, pure-culture fermentation of the B. subtilis production strain. After the enzyme is secreted into the fermentation broth, coagulating agents are added, and the broth is clarified by filtration. The enzyme preparation is purified by ultrafiltration, lyophilized, and standardized with dextrin. Japan Cellfoods cites several published articles and FDA's GRAS affirmation of carbohydrase and protease from B. subtilis (64 FR 19887-19895) to support its view that the manufacturing process used for the production of the pectate lyase enzyme preparation is based on generally available and accepted methods.

Japan Cellfoods describes an unpublished acute oral toxicity study in mice using the enzyme preparation. The test article for this study was prepared according to the standard production method, except that stabilization and standardization were omitted. Japan Cellfoods concludes that this study showed no treatment-related toxicity.

Based on the information provided by Japan Cellfoods, as well as other information available to FDA, the agency has no questions at this time regarding Japan Cellfoods' determination that the pectate lyase enzyme preparation is GRAS under the intended conditions of use. The agency has not, however, made its own determination regarding the GRAS status of the subject use of pectate lyase enzyme preparation. As always, it is the continuing responsibility of Japan Cellfoods to ensure that food ingredients that the firm markets are safe, and are otherwise in compliance with all applicable legal and regulatory requirements.

In accordance with proposed 21 CFR 170.36(f), a copy of the text of this letter, as well as a copy of the information in your notice that conforms to the information in proposed 21 CFR 170.36(c)(1), is available for public review and copying on the homepage of the Office of Food Additive Safety (on the Internet at http://www.cfsan.fda.gov/~lrd/foodadd.html).