GRAS Notice (GRN) No. 685 http://www.fda.gov/Food/IngredientsPackagingLabeling/GRAS/NoticeInventory/default.htm ORIGINAL SUBMISSION 1701 Pennsylvania Avenue, NW Attomeys at Law in Suite 700 Chicago Quoties/ Bmdy u.r Washington, District of Columbia 20006-5805 Indianapolis 202.372.9600 Madison Fax 202.372.9599 Milwaukee www.quarles.com Naples Phoenix Scottsdale Tampa Tucson Washington, D.C. Writer's Direct Dial: 202-372-9529 E-Mail: [email protected] December 20,2016 BY Hand Delivery United States Food and Drug Administration Center for Food Safety and Applied Nutrition Office ofFood Additive Safety HFS-200 5001 Campus Drive College Park, MD 20740 Re: GRAS Notification for Lactobacillus plantarum Strain 299v Dear Dr. Anderson: Enclosed is a copy of a GRAS notification submitted on behalfofProbi AB, of Lund, Sweden ("Probi") through its Agent Mark Yacura ofthe law firm Quarles & Brady LLP in accordance with the requirements of21 C.F.R. Part 170, Subpart E. If you have any questions or concerns regarding these minutes, please contact me at (202) 372-9529 or at [email protected]. Sincerely, (b) (6) fR1[E~[EG~[EfQJ DEC 2 1 2016 OFFICE OF FOOD ADDITIVE SAFETY Generally Recognized as Safe (GRAS) Determination for the Use of Lactobacillus plantarum Strain 299v in Conventional Foods Submitted by ProbiAB Lund, Sweden Submitted to United States Food and Drug Administration Center for Food Safety and Applied Nutrition Office of Food Additive Safety HFS-200 5001 Campus Drive College Park, MD 20740 Prepared by ProbiAB and JHeimbach LLC Port Royal, Virginia December 2016 GRAS Determination for 1 JHEJMBACH LLC Lactobacillus plantarum 299v Table of Contents Part 1 - Signed Statements and Certification............................................................................. 1 Part 2 -Identity, Method of Manufacture, Specifications, and Physical or Technical Effect3 2.1. Name ofthe GRAS Organism ................................................................................. 3 2.2. Source ofthe GRAS Organism ................................................................................ 3 2.3. Description ofthe GRAS Organism ............................ :........................................... 3 1.3 .1. Phenotypic Strain Identification ....................................................................... 5 2.3.2. Genotypic Strain Identification ......................................................................... 6 2.3 .3. Plasmids ............................................................................................................ 7 2.3.4. Surface-Associated Proteins ............................................................................. 8 2.4 Production Method .................................................................................................. 10 Part 3 - Dietary Exposure....................................................................•.••..•.•.............................. 13 Part 4 - Self-limiting Use Levels ...•••••..•••.••...............................................................•••.......•...... 14 Part 5 -Experience Based on Common Use in Food ............................................................... 15 Part 6- Narrative ........................................................................................................................ 16 6.1. History of Safe Ingestion ....................................................................................... 16 6.1.1. Lactic Acid Bacteria and Lactobacillus.......................................................... 16 6.1.2. Lactobacillus plantarum ................................................................................. 17 6.1.3. L. plantarum 299v........................................................................................... 18 6.2. Safety-Related Issues ............................................................................................. 18 6.2.1. Antibiotic Resistance ...................................................................................... 18 6.2.1.1. Minimal Inhibitory Concentrations .......................................................... 19 6.2.1.1.1. 2005 MIC Testing ............................................................................. 19 6.2.1.1.2. 2016 MIC Testing ............................................................................. 20 6.2.1.2. Genetic Analysis ...................................................................................... 21 6.2.2. Production ofBacteriocins .............................................................................. 22 6.2.3. Production ofCarboxylic Acids ...................................................................... 23 6.2.3.1. Total Lactic Acid and Acetic Acid .......................................................... 23 6.2.3.1. D-Lactic Acid ........................................................................................... 23 6.2.4. Production of Biogenic Amines ...................................................................... 25 6.2.5. Allergic Potential ............................................................................................ 26 6.2.6. Infectivity ........................................................................................................ 26 6.3. Research Studies ofL. plantarum 299v................................................................. 27 GRAS Determination for 1 JHEIMBACH LLC Lactobacillus plantarum 299v 6.3 .1. In Vitro Studies ............................................................................................... 27 6.3.2. Animal Studies ................................................................................................ 29 6.3.3. Human Studies ................................................................................................ 47 6.3.3 .1. Healthy Adults ......................................................................................... 4 7 6.3.3.2. Healthy Children ...................................................................................... 51 6.3.3.3. Compromised Adults ............................................................................... 52 6.3.3.4. Compromised Children ............................................................................ 60 6.3.3.5. Conclusions from Human Studies ........................................................... 61 6.4. Evaluations by Authoritative Bodies ..................................................................... 77 6.5. Safety Assessment and GRAS Determination ....................................................... 78 6.5.1. Introduction..................................................................................................... 78 6.5.2. Safety Evaluation ............................................................................................ 79 6.5 .3. General Recognition of the Safety of L. plantarum 299v ............................... 80 Part 7- List of Supporting Data and Information ................................................................... 82 GRAS Determination for 2 1HEIMBACH LLC Lactobacillus plantarum 299v List of Tables Table 1. Growth ofL. plantarum Strain 299v on Different Sugars .................................... 6 Table 2. Sizes ofldentified Plasmids .................................................................................. 8 Table 3. Microbiological Analyses for Prime Ampoules ................................................. 11 Table 4. Results of2005 MIC Testing of L. plantarum 299v........................................... 20 Table 5. Resistance Genes Determined Not To Be Present in L. plantarum 299v........... 22 Table 6. Studies of L. plantarum 299v in Animals............................................. 35 Table 7. Studies ofL. plantarum 299v in Humans ............................................. 58 List of Figures Figure 1. L. plantarum 299v Colonies on MRS Agar (Magnification lx)......................... 4 Figure 2. Gram-Stained L. plantarum 299v (Magnification 1OOOx) .................................. 5 Figure 3. Scanning Electron Micrograph ofL. plantarum 299v (Magnification 20,000x). 5 Figure 4. Plasmid Profiles ofL. plantarum 299v and Other Strains .................................. 8 Figure 5. Production Schematic for L. plantarum 299v................................................... 10 Figure 6. Results of2016 MIC Testing ofL. plantarum 299v......................................... 21 List of Attachments Expert Panel Conclusion ...............................................................Appendix A GRAS Determination for 3 JHEIMBACH LLC Lactobacillus plantarum 299v 0Yc(, Generally Recognized as Safe (GRAS) Determination for the Use of Lactobacillus plantarum Strain 299v in Conventional Foods Part 1 - Signed Statements and Certification GRAS Notice Submission Probi AB, ofLund, Sweden ("Probi") submits this GRAS notification through its Agent Mark Y acura, Partner in the law firm Quarles & Brady LLP in accordance with the requirements of21 C.F.R. Part 170, Subpart E. Name and Address ProbiAB Siilvegatan 41 A SE-223 70 Lund, Sweden Name of Notified Substance The probiotic bacterium Lactobacillus plantarum designated 299v. The strain is also known commercially as Plantarum 299v and Lp299v. Intended Conditions of Use L. plantarum 299v is intended to be added as a pro biotic microorganism to conventional foods at concentrations consistent with cGMP needed to provide beneficial health effects. Intended food applications include but are not limited to the following: