ENGINEERING RESEARCH INSTITUTE THE UNIVERSITY OF MICHIGAN ANN ARBOR Progress Report No 5 COMBINED USE OF HEAT AND RADIATION TREATMENT FOR STERILIZATION OF FOODS Period 7 February 1956 to 7 April 1956 Lloyd Lo Kempe Official Investigator J. To Graikoski Nancy Jo SWilliams Petex F. Bonventre Collaborators Project 2391 QUARTERMASTER RESEARCH AND DEVELOPMENT COMMAND NATICK, MASSACHUSETTS CONTRACT NO. DA-19-129-qm-388, PROJECT NO, 7-84-01-002 April 1956

CONTRACT RESEARCH PROJECT REPORT QUARTERMASTER FOOD AND CONTAINER INSTITUTE FOR THE ARMED FORCES, CHICAGO Hq, QM Research and Development Command, QM Research and Development Center, Natick, Mass. The University of Michigan Engineering Research Institute Ann Arbor, Michigan Official Investigator: Lloyd L. Kempe Collaborators: J. T. Graikoski Nancy J. Williams Peter F. Bonventre Project No. 7-84-01-002 Contract No. DA-19-129-qm-388 File No. S-510 Report No. 5 (Progress) Period 7 February 1956 to 7 April 1956 Initiation Date: 7 June 1955 Title of Contract: Combined Use of Heat and Radiation Treatment for Sterilization of Foods EVALUATION OF CONSECUTIVE IRRADIATION AND HEAT PROCESSING FOR STERILIZING CANNED MEAT The data shown in Table I establish the combined heat and irradiation processing treatments required to sterilize ground beef packed in No. 1 picnic tin cans. When approximately 5,000,000 C. botulinum 213B spores are inoculated into previously heat-sterilized canned beef, the meat can again be sterilized with from 3.42 to 35.96 megarep of gamma radiation from cobalt-60 or by a heat process developing an Fo of approximately 1.0. When heat and radiation processing are used together, preirradiation of the canned meat with less than half the sterilizing dose of gamma rays reduces the subsequent heat processing time required for THIS IS NOT A FINAL REPORT. CONCLUSIONS STATED ARE SUBJECT TO CHANGE ON THE BASIS OF ADDITIONAL EVIDENCE. THIS INFORMATION IS NOT TO BE PUBLISHED WITHOUT WRITTEN PERMISSION FROM HQ, QM R AND D COMMAND, NATICK, MASS. 1

TABLE I Fo Required to Sterilize Ground Beef Packed in No. 1 Picnic Tin Cans, Inoculated with C. botulinum 213B Spores and Irradiated with Gamma Rays from Cobalt-60 Before Heat Processing Megarep Fo Range, Minutes Actual Number of Spores per Can a) 5,000,000 spores per can (approx.) 3.42-3.96 0 5,000,000 0 0.75-1.50 7,200,000 0 Close to 1.0 10700,000 0 0.36-o.93 10,700,000 0.500 0.77-1.06 6,ooo,ooo 0.675 More than 0.58 6,300,000 1.000 0.41-0.80 6,000,000 1.200 0.09-0.29 3,800,000 1.500 0.063-0.27 5,000,000 b) 300 spores per can (approx.) 0 0.14-0.31 125 0 0.15-0.45 400 1.12-2.30 0 *250 0.5 Approx. 0.4 (now incubating) 300 Note: Incubation temperature for runs in Section (a) is 37~C; for those in Section (b), 29~C, except for *, which also is 37~C. sterilization to less than one-fourth the amount required without preirradiation. These data are based on incubation of the processed cans at 37~C. The data reported in Table I(b), except for the irradiation alone, were obtained from canned beef incubated at 29~C. This incubation temperature is to be used for future work. The data in Table I further show that reducing the C. botulinum 213B spore concentration from 5,000,000 to 300 spores per can results in a corresponding reduction in the Fo required from approximately 1.0 to approximately 0.3; similarly, the radiation sterilization dosage drops from the range of 3.42 to 3.96.megarep to 1.12 to 2.30 megarep. 2

It will be noted in Table I that preirradiation with about 0.9 megarep of gamma radiation is required before the combined radiation-heat process shows significant advantage. This has been established for the 5,000,000 spores per can concentration, and it now appears that this probably will be indicated for the 300 spores per can level also. Data from last year's project, obtained when gelatin was used as the suspending medium for C. botulinum 213B spores, produced this same conclusion [see Kempe, Applied Microbiology, 3:350 (1955), Fig. 5]. Table II shows the basic data, summarized in Table I, that have been accumulated during this reporting interval. At present, work is continuing to complete the series of tests designed to establish combined irradiation and heat processing treatments required to sterilize canned ground beef containing approximately 250 C. botulinum 213B spores per can. Also, the vacuum closing machine is being readied for further studies of the combined irradiation and heat processing treatment. In this process the canned meat will be inoculated while raw. It will then be irradiated in the raw condition and finally heat processed. This will differ from the present studies in that the meat is now first sterilized with steam under pressure, then inoculated, and finally heat processed again. The difference will of course arise from any possible effects of raw-meat components on the effectiveness of the preirradiation treatment. EFFECT OF CHEMICALS IN THE SUSPENDING MEDIUM ON THE LETHALITY OF GAMMA RADIATION FOR ANAEROBIC SPORES Work is continuing on this phase of the project but will not be reported on at this time. EFFECT OF TEMPERATURE OF THE SUSPENDING MEDIUM DURING IRRADIATION ON THE LETHALITY OF GAMMA RADIATION FOR ANAEROBIC BACTERIAL SPORES During the past few weeks equipment has been designed, built, and tested to take data at temperatures above 100~C. This temperature range is important but poses several problems of technique. However, these problems appear to have been satisfactorily solved and data will be taken at temperatures above 1000C within the near future. 3

TABLE II Processing Treatments Required to Sterilize Canned Ground Beef Inoculated with C. botulinum 213B Spores Run CB-14 Objective: Establish radiation sterilization level at low spore concentration Can size - No. 1 Picnic Product - Ground Beef Inoculation - 250 C. botulinum 213B Temperature of irradiation - 6~C spores per can Irradiation Dose. I d i D Can No. Gas Formation megarep Inoculated Controls (from Run CB-135) Experimental Cans 1.12 1.12 1.12 1.12 1.12 1.12 1.12 1.12 17 18 19 20 33 34 55 36 37 38 39 4o + + + + + + 4+ + + + + ++ Days to Gas Formation 2.5 2.5 2.5 2.5 3 4 3 5 4 5 4 2.30 2.350 2.50 2.50 29 350 31 52 2.90 2.90 2.90 2.90 3.4o 5.40 3.40 3.40 25 26 27 28 17 18 19 20 4.00 21 4.00 22 4.oo 23 4.00oo 24 Conclusion: The radiation sterilization dosage for canned ground beef containing 250 C. botulinum 215B spores per can lies between 1.12 and 2.30 megarep of-cobalt-60 gamma radiation. Note: This compares favorably with data reported previously for No. 2 cans of canned beef [see Kempe, Graikoski, and Gillis, Applied Microbiology, 2:55330 (1954)]. 4

TABLE II (Continued) Run CB-15 Objective: Establish radiation sterilization level at high spore concentration Can size - No. 1 Picnic Product - Ground Beef Inoculation - 5,000,000 C. botulinum 213B spores per can Irradiation temperature - 7~C Irradiation DoseCan o Gas Foratio.$ Can No. Gas Formation Days to Gas megarep Formation Noninoculated Controls 1 2 3 4 Inoculated Controls 21 + 2 22 + 2 23 + 2 24 + 2 3.42 1 3.42 2 + 4 3.42 3 + 4 3.42 4 + 4 3.96 5 3.96 6 3.96 7 3.96 8 2.52 9 + 4 2.52 10 + 3 2.52 11 + 3 2.52 12 + 3 2.16 13 + 2 2.16 14 + 3 2.16 15 + 3 2.16 16 + 3 2.88 17 + 4 2.88 18 + 3 2.88 19 + 5 2.88 20 + 4 Conclusion: The radiation sterilization dosage for canned ground beef containing 5,000,000 C. botulinum 213B spores per can lies between 35.42 and 3596 megarep of cobalt-60 gamma radiation. Note: This compares favorably with data reported previously for No. 2 cans of canned beef [see Kempe, Graikoski, and Gillis, Applied Microbiology, 2:330 (1954)]. 5

TABLE II (Continued) Run CB-17 Objective: Establish Fo needed, using approximately 250 C. botulinum 213B spores per can Can size - No. 1 Picnic Product - Ground Beef Inoculation - 250 C. botulinum 213B spores per can Irradiation Dose, C No. Days to Gas I Can No. Gas Formation megarep Formation Noninoculated Controls 1 2 5 4 Inoculated Controls 17 3 + 18 3 + 19 3 + 20 3 + Fo = 0.21 1 2 5 - 4 Fo = 0.51 5 6 7 8 Fo = 0.28 9 10 11 - 12 - Fo = 0.537 13 - - 14 - - 15 - 16 Conclusion: than 0.21. Fo for 250 C. botulinum spores per can appears to be less 6

TABLE II (Continued) Run CB-18 Objective: Establish Fo, using approximately 250 C. botulinum 213B spores per can Can size - No. 1 Picnic Product - Ground Beef Inoculation - 125 C. botulinum 213B spores per can Days to Gas *Toxin F0_ Can aNo _ Formation Production Noninoculated Controls 1 4 0/3 2 3 14 0/3 4 Inoculated Controls 17 3 18 3 19 3 3/3 20 3 Can 1 = 0.31 1 7 0/3 Can 2 = 0.26 2 3 4 Can 1 = 0.14 5 4 3/3 6 4 7 4 0/3 8 4 Can 1 = 0.079 9 3 1/3 Can 2 = 0.058 10 3 1/3 11 3 12 3 Can 1 = 0.035 13 3 14 3 15 3 3/3 16 3 Conclusion: Fo of 250 C. botulinum 213B spores in ground beef packed in No. 1 picnic tin cans lies between 0.14 and 0.31. *Toxin production reported as fraction of Number of dead mice Number of mice inoculated Control of toxin: Heated toxin containing sample 0/3 mice. 7

TABLE II (Concluded) Run CB-19 Objective: Establish Fo, using approximately 250 C. botulinum 213B spores per can Can size - No. 1 Picnic Product - Canned Beef Inoculation - 400 C. botulinum 213B spores per can F ~ Can No. Days to Gas *Toxin F___0 Can__ No. __ Formation Production Inoculated Control 19 20 21 22 4 1 4 4 2/3 Noninoculated Control 1 2 3 4 17 under investigation Can 1 = 0.030 Can 2 = 0.045 1 2 3 4 6 6 4 6 Can 1 = 0.084 5 6 7 8 9 10 11 12 5 4 4 5 2/3 1/3 Can 1 = 0.15 8 3/3 Can 1 = 0.26 0.45 13 14 15 16 4 0/3 Conclusion: and 0.45. F0 of 400 C. botulinum 213B spores per can lies between 0.15.0 _.. 8