ENGINEERING RESEARCH INSTITUTE THE UNIVERSITY OF MICHIGAN ANN ARB OR Progress Report No. 9 COMBINED USE OF HEAT AND RADIATION TREATMENT FOR STERILIZATION OF FOODS Period: 30 September 1956 to 30 November 1956 Lloyd L. Kempe Official Investigator T. T. Graikoski Collaborator Project 2391 QUARTERMASTER RESEARCH AND DEVELOPMENT COMMAND NATICK, MASSACHUSETTS CONTRACT NO. DA-19-129-qm-388, PROJECT NO. 7-84-01-002 December 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 Collaborator: J. T. Graikoski Project No. 7-84-01-002 Contract No. DA-19-129-qm-388 File No. S-510 Report No. 9 (Progress) Period 30 September 1956 to 30 November 1956 Initiation Date: 7 June 1955 Title of Contract: Combined Use of Heat and Radiation Treatment for Sterilization of Foods SUMMARY Tests oftthe combined irradiation-heat processing of canned ground beef inoculated with approximately 10,000 PA-3679 spores per can are in progress. Present data indicate the following for these conditions: 1. Approximately 1.0 megarep preirradiation is required before a significant effect of irradiation is noted on the Fo value required for subsequent heat sterilization of the canned meat. 2. between 8.4 was Following irradiation with 1.7 megarep of gamma radiation, an Fo 1.1 and 3.2 is required for sterilization; an Fo between 6.5 and required with heat processing alone. These studies are being extended to include slightly higher preirradiation levels. 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 THIS INFORMATION IS NOT TO BE PUBLISHED WITHOUT WRITTEN PERMISSION FROM HQ, QM R AND D COMMAND, NATICK, MASS. 1

EFFECT OF PREIRRADIATION OF CANNED GROUND BEEF INOCULATED WITH PA-3679 SPORES ON THE F0 SUBSEQUENTLY REQUIRED FOR STERILIZATION Data have been presented in previous reports to show that the Fo required to sterilize canned ground beef inoculated with C. botulinum 213B spores can be considerably reduced when heat processing follows irradiation of the canned meat with gamma rays. This study is now being extended to include packs inoculated with PA-3679 spores. Up to the present time, all the work has been carried out on previously heat-sterilized meat. The techniques in use are similar to those previously described in the annual report for this project dated September, 1956, except that PA-3679 spores are presently being used. The previous work was done with C. botulinum 213B spores. Briefly these techniques involve preparation of inoculated packs of lean ground beef in No. 1 picnic tin cans which are closed while hot in order to develop vacua. The cans are then irradiated in the large cobalt-60 gamma-radiation source in the Fission Products Laboratory, at The University of Michigan. They are next processed in a steam autoclave and finally incubated at 29~C. Sufficient F0 values are used to permit bracketing that F0 value which is required to sterilize all four cans. Sterility is determined by observation of swelling during incubation. One variation, namely the use of a 250~F processing temperature instead of 230~F, was tried for Runs PA-2 through PA-9. This was revised because of operating difficulties and also because 2300F is more commonly used industrially. Runs following PA-9 were conducted with the processing temperature of 230~F, the temperature previously used with C. botulinum 213B spore packs. The PA-3679 spores are grown, prepared, counted, and used according to techniques described in previously published work from this laboratory.1 Data from the early runs are included in Table I. The steam temperatures for the combined heat and irradiation processing experiments reported in this table was 250~F. This temperature was lowered to 230~F for runs following PA-9. Two runs PA-4 and PA-7, included in this table are useful in establishing the level of irradiation required to sterilize canned ground beef when irradiation is used alone. "Gamma-ray sterilization of canned meat previously inoculated with anaerobic bacterial spores," by Kempe et al., Applied Microbiology, 2, 330 (1954).

Table II and the accompanying figure show that preirradiation reduces the severity of the heat processing required to sterilize canned ground beef inoculated with approximately 10,000 PA-3679 spores per can. The present data indicate that preirradiation with approximately 1.0 megarep is required before a significant reduction in the Fo value, subsequently needed for sterilization, occurs under these conditions. Also, preirradiation with 1.7 megarep reduced the Fo value, subsequently required, to a value between 1.1 and 3.2. This compares with an Fo value between 6.5 and 8.4 that is needed to produce sterility when no preirradiation is used. These studies are being extended to include slightly higher preirradiation levels before work is begun on inoculated packs of raw meat. 3

10 w 0.5' 0.4 z 0.3'o 02 0.I.08.06.05.04.03.02 Irradiation alone 2.2- 3.0 megarep.01 I I.01 1.0 2.0 3.0 MEGAREP Fo required to sterilize ground beef packed in No. 1 picnic tin cans, inoculated with approximately 10,000 PA-3679 spores per can and irradiated with gamma rays from cobalt-60 before heat processing at 230~F.

TABLE I - Fo Value Required to Sterilize Ground Beef in No. 1 Picnic Tin Cans, Previously Inoculated with Approximately 10,000 PA-3679 Spores per Can and Then Processed at 250~F. Run No. PA-2-Can Size Product Inoculum Preirradiation Processing Temperature Incubation Temperature - No. 1 Picnic (211 x 400) - Ground Beef - 10,000 PA-3679 spores per can - none - 270~F - 29 C Fo Can No. Days to Gas Formation MaTr —%4ve, iiI n,-11 A % ^41I -- Ala aTT-I I\oiULLLnC.)cui.La U UL onL'roU 1J I JI NI 2 NI 3 NI 4 Inoculated Controls Can 1, 2.5 Can 2, 2.5 26 27 28 29 3 5 3 3 1 3 4 7 5 4 5 Can 1, 4.1 Can 2, 14.1 Can 3, 3.7 (disregard) 2 5 6 8 6 Can 1, Can 2, Can 3, 1.1 1.1 1.3 9 10 11 12 4 4 4 4 Can Can Can 1, 3.7 2, 3.7 3, 2.8 (disregard) 13 14 15 16 4 5 5 Can 1, 6.2 Can 2, 3.3 Can 1, 8.8 Can 2, 8.8 18 19 20 21 4 22 23 24 25 Conclusion: Ground beef, packed in No. 1 picnic tin cans, and inoculated with 10,000 PA-3679 spores per can, required an Fo value between 3.7 and 8.8 for sterilization when processed at 250~F.

TABLE I (Continued) Run No. PA-3-Can Size Product Inoculum Pre irradiation Processing Temperature Incubation Temperature - No. 1 Picnic (211 x 400) - Ground Beef - 600,000 PA-3679 spores per can - none - 250~F (except cans 1, 2, 3, and 4.- TCC at 230~F) Fo Can No. Days to Gas Formation Noninoculated Controls NI 1 NI 2 Inoculated Controls I 1 5 12 +* Can 1, 2.3 1 4 Can 2, 2.3 2 4 Can 3, 2.5 3 4 4 4 Can 1, 6.1 5 4 Can 2, 6.1 66 Can 5, 3.8 (disregard) 7 4 8 5 Can 1, 9.4 9 9 Can 2, 8.5 10 8 Can 3, 8.7 11 6 12 9 Can 1, 12.4 13 9 Can 2, 12.4 14 15 Can 3, 11.6 (disregard) 15 13 16 16 *Date not recorded. Conclusion: With 600,000 PA-3679 spores ture of 250~F, the Fo value required for exceeds 12.4. per can and a processing temperasterilization of the canned meat

TABLE I (Continued) Run No. PA-4 —Can Size - No. 1 Picnic (211 x 400) Product - Ground Beef Inoculum - 200 PA-3679 spores per can Preirradiation - as indicated Processing Temperature - not heat processed Incubation Temperature - 29~C Irradiation Dosage, megarep Can No. Days to Gas Formation Noninoculated Controls See Run 5 Inoculated Controls See Run 5 4.100 1 2 3 4 35700 5 6 7 8 1.600 9 10 11 6 12 2.100 13 14 15 16 Conclusion: Under the above conditions canned ground beef was sterilized with between 1.600 and 2.100 megarep of gamma irradiation.

TABLE I (Continued) Run No. PA-5-Can Size Product Inoculum Preirradiation Processing Temperature Incubation Temperature - No. 1 Picnic (211 x 400) - Ground Beef - 200 PA-3679 spores per can - none - 250~F - 290C Fo Can No. Days to Gas Formation Noninoculated Controls NI 1 NI 2 NI 3 NI 4 Inoculated C ontrols Can 1, 2.7 Can 2, 2.7 Can 1, 3.6 Can 2, 6.8 Can 3, 8.3 INC 1 INC 2 INC 3 INC 4 3 3 3 3 5 1 2 3 4 5 6 8 9 Can 1, Can 2, Can 5, 12.3 12.5 15.3 i (disregard) (disregard) 10 11 12 15 Can Can Can Can Can Can 1, 9.0 2, 8.0 3, 8.1 14 15 16 17 1, 4.9 2, 2.7 3, 6.8 18 19 20 21 Can 1, 4.4 Can 2, 4.4 23 24 25 26 Can 1, 14.1 Can 2, 12.6 7 22 27 28 an Fo value le-ss than 2.7 was Conclusion: Under the above conditions needed to sterilize ground beef.

TABLE I (Continued) Run No. PA-6-Can Size Product Inoculum Preirraidation Processing Temperature Incubation Temperature - No. 1 Picnic (211 x 400) - Ground Beef - 54,000 spores per can - 1.200 megarep - 250~F - 29 C Fo Can No. Days to Gas Formation Noninoculated Controls NI 1 NI 2 Inoculated Controls 17 4 18 4 19 4 20 4 Can 1, 3.1 1 Can 2, 3.1 2 10 Can 3, 2.7 (disregard) 3 4 Can 1, 3.8 5 8 Can 2, 3.5 6 Can 3, 4.1 7 8 Can 1, 1.0 9 5 Can 2, 1.2 10 7 Can 3, 1.7 11 9 12 7 Can 1, 8.6 13 - Can 2, 6.2 14 Can 3, 5.5 15 16 Conclusion: Under these conditions sterilization obtained by irradiation with 1.2 megarep of gamma value between 3.5 and 8.6. of canned ground beef is rays followed by an Fo

TABLE I (Continued) Run No. PA-7-Can Size Product Inoculum Irradiation Processing Temperature Incubation Temperature - No. 1 Picnic (211 x 400) - Ground Beef - 13,500 PA-3679 spores per can - as indicated - not processed - 29 C Megarep Can No. Days to Gas Formation Noninoculated Controls See Run 8 Inoculated Controls 35 36 37 38 3 3 3 3 5 1.000 1 2 3 4 4 4 4 4 2.200 13 14 15 16 6 4 1.900 31 32 33 34 4 4 4 4 3.000 9 10 11 12 3.900 5 6 7 8 Conclusion: Irradiation-sterilization dosage for canned ground beef under these conditions lies between 2.2 and 3.0 megarep.

TABLE I (Continued) Run No. PA-8-Can Size Product Inoculum Preirradiation Processing Temperature Incubation Temperature - No. 1 Picnic (211 x 400) - Ground Beef - 13,500 PA-3679 spores per can -none - 250~F - 29~C Fo Can No. Days to Gas Formation Noninoculated Controls NI 1 NI 2 NI 5 NI 4 - Inoculated Controls Can 1, 3.1 Can 2, 2.5 Can 1, 9.6 Can 2, 9.6 Can 1, 5.6 Can 2, 5.6 Can 1, 3.8 Can 2, 2.9 55 36 57 58 4 4 4 4 15 6 1 2 3 5 4 5 6 7 8 9 10 11 12 15 14 15 16 9 6 Can 1, Can 2, Can 3, 7.9 10.7 12.8 17 18 19 20 Can 1, 7.0 Can 2, 7.0 21 22 25 24 Can 1, 11.6 25 Can 2, 9.2 26 27 28 Conclusion: Under these conditions canned ground beef was sterilized with heat processing at 250~F, having an Fo value between 2.5 and 5.6.

TABLE I (Concluded) Run No. PA-9-Can Size Product Inoculum Preirradiation Processing Temperature Incubation Temperature - No. 1 Picnic (211 x 400) - Ground Beef - 16,000 PA-3679 spores per can - 1.000 megarep - 250~F - 29 C Fo Can No. Days to Gas Formation Inoculated Controls 21 3 25 3 24 3 Irradiation Controls, 17 3 1.340 megarep 18 3 19 3 20 3 Can 1, 2.3 1 4 Can 2, 2.3 2 5 Can 3, 0.6 (disregard) 3 4 4 5 Can 1, 3.8 5 6 Can 2, 3.8 6 Can 3, 4.2 7 8 7 Can 1, 0.5 9 5 Can 2, 0.5 10 4 11 4 12 4 Can 1, 6.5 13 Can 2, 9.0 14 Can 3, 11.7 15 16 Conclusion: Under these conditions canned ground beef was sterilized by irradiation with 1.500 megarep of gamma irradiation followed by heat processing to an Fo value of between 6.3 and 11.7.

TABLE II - Fo Value Required to Sterilize Ground Beef in No. 1 Picnic Tin Cans, Previously Inoculated with Approximately 10,000 PA-3679 Spores per Can and Then Processed at 230~F. Run No. PA-lO-Can Size Product Inoculum Preirradiation Processing Temperature Incubation Temperature - No. 1 Picnic (211 x 400) - Ground Beef - 10,000 PA-3679 spores per can - none - 250~F - 290C Fo Can No. Days to Gas Formation Noninoculated Controls NI 1 NI 2 NI 3 NI 4 Inoculated Controls Can 1, 6.0 Can 2, 6.0 Can 3, 5.2 Can 1, 3.8 Can 2, 3.8 Can 3, 3.8 Can 1, 3.0 Can 2, 3.0 Can 3, 3.0 17 18 19 20 3 3 3 3 1 2 3 4 9 11 8 11 5 6 7 8 5 6 5 5 6 5 5 5 9 10 11 12 Can 1, Can 2, Can 3, 2.0 2.0 2.0 13 14 15 16 value 4 4 4 4 sterilize Conclusion: Under these conditions the Fo canned ground beef exceeds 6.0. required to

TABLE II (Continued) Run No. PA-11-Can Size Product Inoculum Preirradiation Processing Temperature Incubation Temperature - No. 1 Picnic (211 x 400) - Ground Beef - 10,000 PA-3679 spores per can - 1.500 megarep - 230~F - 29~C Fo Can No. Days to Gas Formation Noninoculated Controls NI 1 NI 2 NI 4 NI 4 - Inoculated Controls Can 1, 1.0 Can 2, 0.8 Can 3, 0.4 17 18 19 20 3 3 3 3 1 2 3 4 5 5 5 5 Can 1, Can 2, Can 3, 1.0 0.8 5 6 7 8 6 5 5 6 Can Can Can 1, 2.0 2, 2.0 3, 3.0 9 10 11 12 9 6 9 (disregard) Can 1, 3.9 13 Can 2, 3.9 14 Can 3, 4.9 (disregard) 15 16 Conclusion: Canned ground beef was sterilized under these conditions by 1.500 megarep of preirradiation followed by heat processing with an F0 value between 2.0 and 3.9.

TABLE II (Continued) Run No. PA-12 —Can Size Product Inoculum Pre irradiation Processing Temperature Incubation Temperature - No. 1 Picnic (211 x 400) - Ground Beef - 10,000 PA-3679 spores per can - None - 230~F -29~C Fo Can No. Days to Gas Formation Noninoculated Controls NI 1 NI 2 NI 3 Inoculated Controls INOC 3 Can 1, 2.9 1 4 Can 2, 2.9 2 4 3 3 4 5 Can 1, 5.8 5 14 Can 2, 5.8 6 7 11 8 14 Can 1, 4.3 9 6 Can 2, 5.5 10 7 11 6 12 6 Conclusion: Canned ground beef requires an Fo value of more than 5.8 for sterilization under these conditions.

TABLE II (Continued) Run No. PA-13-Can Size Product Inoculum Preirradiation Processing Temperature Incubation Temperature - No. 1 Picnic (211 x 400) - Ground Beef - 10,000 PA-3679 spores per can - 1.000 megarep - 250~F - 29~C Fo Can No. Days to Gas Formation Can 1, 4.4* 1 Can 2, 4.4 2 7 3 4 Can 1, 0.7 5 4 Can 2, 0.5 6 3 7 4 8 4 Can 1, 2.5 9 7 Can 2, 2.3 10 4 11 5 12 5 Can 1, 5.7 13 Can 2, 5.7 14 15 16 Conclusion: Canned ground beef was sterilized under these conditions by 1.000 megarep of preirradiation followed by heat processing with an Fo value between 4.4 and 5.7. *Same controls as used for Run 12.

TABLE II (Continued) Run No. PA-14-Can Size Product Inoculum Preirradiation Processing Temperature Incubation Temperature - No. 1 Picnic (211 x 400) - Ground Beef - 10,000 PA-3679 spores per can - 1.700 megarep - 230~F - 29 C Fo Can No. Days to Gas Formation Noninoculated Controls NI 1 NI 2 NI 3 NI 4 Inoculated Controls Can 1, 5.6 Can 2, 4.9 Can 1, 6.5 Can 2, 5.6 Can 1, 1.1 Can 2, 1.1 17 18 19 20 3 3 3 3 1 2 3 4 5 6 7 8 9 10 11 12 9 9 Can 1, 5.2 15 Can 2, 5.2 14 Can 3, 5.2 15 16 Conclusion: Canned ground beef was sterilized 1.700 megarep of gamma irradiation followed by value between 1.1 and 3.2. under these conditions by heat processing with an F0 an

TABLE II (Continued) Run No. PA-15-Can Size Product Inoculum Preirradiation Processing Temperature - Incubation Temperature - No. 1 Picnic (211 x 400) Ground Beef 10,000 PA-3679 spores per can None 230oF 29~C Fo Can No. Days to Gas Formation Noninoculated Controls NI 1 NI 2 NI 3 NI 4 Inoculated Controls 17 18 19 20 3 3 3 3 Can 1, Can 2, Can 3, 6.5 7.0 7.0 1 2 3 4 21 17 13 Can 1, 8.4 Can 2, 7.3 5 6 7 8 Can 1, Can 2, Can 3, 6.0 6.0 6.0 9 10 11 12 8 8 7 7 Can 1, 5.0 13 11 Can 2, 5.0 14 11 Can 3, 5.0 15 11 16 8 Conclusion: Under these conditions, canned ground beef was sterilized with an Fo value between 6.5 and 8.4.

TABLE II (Continued) Run No. PA-16-Can Size Product Inoculum Preirradiation Processing Temperature Incubation Temperature - No. 1 Picnic (211 x 400) - Ground Beef - 10,000 PA-5679 spores per can - None - 250~F - 29~C Fo Can No. Days to Gas Formation Noninoculated Controls NI 1 NI 2 NI 3 NI 4 Inoculated Controls 153 3 14 3 15 3 INOC 4 Can 1, 7.1 1 Can 2, 7.1 2 Can 3, 7.1 3 4 Can 1, 8.7 5 Can 2, 8.7 6 7 8 Can 1, 4.1 9 7 Can 2, 4.6 10 7 11 7 12 7 Conclusion: Under these conditions canned ground beef was sterilized with an Fo value between 4.1 and 7.1.

TABLE II (Concluded) Run No. PA-17-Can Size Product Inoculum Preirradiation Processing Temperature Incubation Temperature - No. 1 Picnic (211 x 400) - Ground Beef - 10,000 PA-3679 spores per can - 0.850 megarep - 230~F - 290C Fo Can No. Days to Gas Formation Noninoculated Controls NI 1 NI 2 NI 3 NI 4 Inoculated ControlsCan 1, 10.2 Can 2, 10.2 Can 3, 10.2 Can 1, 5.5 Can 2, 4.9 Can 1, 6.9 Can 2, 6.6 13 14 15 INOC 3 3 3 4 1 2 3 4 5 6 7 8 9 10 11 12 Can 1, 3.7 13 Can 2, 3.1 14 17 15 12 16 12 Conclusion (tentative because of insufficient incubation time at the time of this report): Canned ground beef was sterilized under these conditions by 0.850 megarep irradiation followed by heat processing with an Fo value between 3.1 and 5.5.