ENGINEERING RESEARCH INSTITUTE THE UNIVERSITY OF MICHIGAN ANN ARBOR Progress Report DETERMINATION OF RADIATION STERILIZATION DOSE FOR CANNED MEAT L. L-., Kempe J.'T. Graikoski ERI Project 2681 DEPARTMENT OF THE ARMY QUARTERMASTER RESEARCH AND DEVELOPMENT COMMAND QUARTERMASTER FOOD AND CONTAINER INSTITUTE CONTRACT NO. DA-19-129-qm-964 CHICAGO, ILLINOIS February 1958

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 Lo Kempe Collaborator: Jo To Graikoski Project No. 7-84-01-002 Contract No. DA-19-129-qm-964 File No. S-510 Report Noo 3 (Progress) Period 1 December 1957 to 31 January 1958 Initiation Date: 1 August 1957 Title of Contract: Determination of Radiation Sterilization Dose for Canned Meat SUMMARY All phases of the project are now under investigation. Conclusions, however, are premature at present because sufficient incubation time has not yet elapsed to establish even tentative results. THIS IS NOT A FINAL REPORT. CONCLUSIONS STATED ARE SUBJECT TO CHANGE ON THE BASIS OF ADDITIONAL EVIDENCEo THIS INFORMATION IS NOT TO BE PUBLISHED WITHOUT WRITTEN PERMISSION FROM HQ, QM R AND D COMMAND, NATICK, MASS. 1

PHASE I EFFECT OF PREIRRADIATION OF CANNED GREEN PEAS ON THE F0 SUBSEQUENTLY REQUIRED FOR STERILIZATION Combined irradiation-heat processing may conceivably be applied to other foods than meat. The benefits to be logically expected would follow from possible improvement of organoleptic qualities. A suitable test food for this purpose should be one that stimulates germination and subsequent growth of putrefactive anaerobic spores that are significant in food spoilage. Green peas are often used as the base of germination and culture media needed for such spores and in addition, canned green peas are a staple item of human consumption. Hence green peas were selected for study. The combined irradiation-heat processing treatments required to sterilize canned green peas inoculated with 300PA3679 spores per can were presented in Progress Report No. 1 and are summarized here for reference in Fig. 1. This study has been continued with a second series of runs using 5,000,000 C. botulinum 213B spores per can. The canning, irradiation, and heat processing treatments used were described in Progress Report No. 1o RESULTS Data from runs PB-1 through PB-17 are detailed in the table and plotted in the figure. Conclusions are not drawn since longer incubation is required before they would be meaningful. 2

TABLE Fo VALUES REQUIRED TO STERILIZE CANNED GREEN PEAS PACKED IN NO. 1 PICNIC TIN CANS, INOCULATED WITH 5,000,000 C. botulinum 213B SPORES PER CAN, AND IRRADIATED WITH GAMMA RAYS FROM COBALT-60 BEFORE HEAT PROCESSING AT 230OF AND INCUBATION AT 85 F Run No. o Can Size: Product: Inoculum: Irradiation: Processing Temperature: Incubation: PB-1 Noo 1 Picnic Green Peas 5,000,000 C. None 230 ~F 850F (211 x 400) botulinum 215B Spores Per Can.........Fo Can No... Days-to-gas *Toxin F Can No. Formation.Toxin Inoculated controls 1 2 2 2 Noninoculated controls Can 1, Can 2, Can 3, 1.03 1003 1.03 Can 1, 0,63 Can 2, 0.63 Can 3, 0.63 Can 1, 0.52 Can 2, 0.52 Can 1, 0.29 Can 2, 0.29 Can 1, 0o26 Can 2, 0.26 Can 1, 1.44 1 2 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 3 3 9 11 10 12 10 10 10 9 8 9 9 9 5 9 6 9 5 5 6 6 24 13 24 13 2/2 2/2 2/2 2/2 21 22 23 24 2/2 2/2 2/2 2/2 2/2 2/2 25 26 27 28 *Toxin determined by intraperitoneal injection of 0.5 ml of juice into mice (2/2 means 2 dead mice out of 2 injected). Conclusion: An Fo in excess of 1.44 is required to sterilize the canned peas. 3

TABLE (Continued) Run No.: Can Size: Product: Inoculum: Irradiation: Processing Temperature: Incubation Temperature: PB -2 No. 1 Picnic Green Peas 5,000,000 C. As Indicated None 85~F (211 x 400) botulinum 213B Spores Per Can Megarad Can No. ys-to-gas Toxin 1.530 13 6 14 9 15 9 16 9 2.045 9 10 8 11 16 0/2 12 3.80 1 - 2 3 4 35600 5 6 7 17 2/2 8 17 2/2 Conclusion: Provided that no error was made in recording the radiation dosage for cans 1 through 8, it appears that more than 3.60 megarad are needed for sterilization. It seems more likely that the dosage is between 3.08 and 3~60 megarad, however. Note: Same controls as used for Run PB-lo

TABLE (Cont'inued) Run No.: Can Size: Product: Inoculum: Irradiation: Processing Temperature: Incubation Temperature: PB-3 No. 1 Picnic (211 x 400) Green Peas 5,000,000 C. botulinum 213B Spores Per Can 0.465 megarad 2350~F 85~F Days-to-gas Fo Can No. Formation Toxin Can 1, 1553 1 Can 2, 1.53 2 Can 35, 1.53 3 4 5 11 0/2 Can 1, 1.03 6 11 0/2 Can 2, 1.03 7 15 2/2 Can 3, 1.03 8 15 2/2 Can 1, 0.70 9 16 Can 2, 0.70 10 20 Can 3, 0.70 11 13 2/2 12 26 Can 1, 0.46 13 9 Can 2, 0.46 14 17 15 13 16 9 Can 1, 0.26 17 9 Can 2, 0.26 18 13 19 13 20 9 Conclusion: Under these conditions, canned green peas were sterilized by 0.465 megarad of gamma radiation followed with an Fo between 1.03 and l55.3. 5

TABLE (Continued) Run No.: Can Size: Product: Inoculum: Irradiation: Processing Temperature: Incubation Temperature: PB-4 No. 1 Picnic Green Peas 5,000,000 Co None 230~F 85~F (211 x 400) botulinum 213B Spores Per Can ~Fo^~ Can No,~ ~Days-to-gas F0 Can 1.8 Formation Can 1, 1.82 1 Can 2, 1.82 2 3 4 Can 1, 1.10 5 12 2/2 6 13 7 10 8 15 Can 1, 1.39 9 15 2/2 Can 2, 1.39 10 11 12 12 2/2 Conclusion: Under these conditions, by Fo values between 1.39 and 1.82. canned green peas were sterilized 6

TABLE (Continued) Run No.: Can Size: Product: Inoculum: Irradiation: Processing Temperature: Incubation Temperature: PB-5 No. 1 Picnic (211 x 400) Green Peas 5,000,000 C. botulinum 213B Spore Per Can 0.930 megarad 230 ~F 85~F Days-to-gas Fo Can No. Toxin Formation Can 1, 1.06 1 Can 2, 1.06 2 Can 3, 1.06 3 4 Can 1, 0.86 5 Can 2, 0.86 6 Can 31 0.86 7 8 Can 1, 0.36 9 17 Can 2, 0.36 10 20 Can 35 0.36 11 19 12 17 Can 1, 0.50 13 Can 2, 0.50 14 Can 35 0.50 15 16 Conclusion: Under these conditions, canned green peas by 0.930 megarad of gamma radiation followed with a Fo 0.50. were sterilized between 03.6 and 7

TABLE (Continued) Run No.: Can Size: Product: Inoculum: Irradiation: Processing Temperature: Incubation Temperature: PB-6 No. 1 Picnic (211 x 400) Green Peas 5,000,000 C. botulinum 213B Spores Per Can As Indicated Not Heat Processed 85~F Megarad Can No. Daystogas Formation 2.330 1 12 2 18 3 12 4 19 2.790 5 6 7 8 o1980 9 12 10 17 11 18 12 10 Conclusion: Under these conditions, canned green peas were sterilized by between 2.33 and 2.79 megarad of gamma radiation. 8

TABLE (Continued) Run N o: Can Size: Product: Inoculum: Irradiation: Processing Temperature: Incubation Temperature: Object: PB-7 No. 1 Picnic (211 x 400) Green Peas None None 230~F 850F To determine the Fo required to sterilize canned green peas. The peas were frozen green peas purchased from The University of Michigan food stores. F Can No. Days-to-gasoxin Fo Can No. Formation Toxn Can 1, 0.03 1 4 Can 2, 0.03 2 5 Can 3, 0.03 3 4 4 5 Can 1, 0.06 5 4 0/2 Can 2, 0.06 6 6 Can 3, 0o06 7 4 8 Noninoculated, 9 3 0/2 unheated control Inoculated 10 5 2/2 unheated control Irradiated control, 11 11 0.465 megarad 12 11 Conclusion: Canned green peas contain sufficient anaerobic bacterial spores in the frozen condition, as received by us, to require an Fo greater than 0.06 or irradiation with more than 0.465 megarad, to produce sterile peas. 9

TABLE (Continued) Run No o Can Size: Product: Inoculum: Irradiation: Processing Temperature: Incubation Temperature: PB-8 No. 1 Picnic (211 x 400) Green Peas 5,000,000 C. botulinum 213B Spores Per Can 1395 Megarad 230~F 85~F Fo Can No. Days-to-gas Formation Can 1, 0.48 9 Can 2, 0.48 10 Can 3, 0.48 11 12 Can 1, 0.33 13 Can 2, 0.33 14 Can 3, 0.33 15 *32 16 Can 1, 0.21 17 25 Can 2, 0.21 18 25 19 30 20 27 Can 1, 0.12 21 27 Can 2, 0.12 22 30 23 25 24 25 Can 1, 0004 25 19 Can 2, 0o04 26 13 27 18 28 13 Conclusion: Under these conditions, canned green peas were sterilized by 1.395 megarad of gamma radiation followed with Fo between 0533 and 0. 48. *Positive for Type B Botulinum toxin. 10

TABLE (Continued) Run No: Can Size: Product: Inoculum: Irradiation: Processing Temperature: Incubation Temperature: Object: PB-9 No. 1-Picnic (211 x 400) Green Peas None None 230 ~F 85~F Same as Run No. PB-7, to determine the Fo required to sterilize noninoculated peas that we are using for our experiments. Fo Can No. Days-to-gas Toxin F0 Can No. Df-raFormation Can 1, 0.027 1 3 Can 2, 0.027 25 3 5 4 5 Can 1, 0.036 5 Can 2, 0.036 6 5 7 3 8 3 Can 1, 0.036 9 5 Can 2, 0.036 10 5 11 11 12 5 0/4 Can 1, 0.054 13 5 4/4 Can 2, 0.054 14 5 *4/4 15 3 1/4 16 *Tests as Botulinum toxin type B with antiserum neutralization. Conclusion: An Fo in excess of 0.054 is needed to sterilize the noninoculated canned green peas that we are using in our experiments. 11

TABLE (Continued) Run No o: Can Size: Product: Inoculum: Irradiation: Processing Temperature: Incubation Temperature: PB-10 Noo 1 Picnic Green Peas 5,000,000 C. As Indicated None 85~F (211 x 400) botulinum 213B Spores Per Can Megarad Can No. Days-to-gas Formation 1.860 1 12 2 11 3 11 0.930 5 10 6 7 7 10 2.790 9 10 22 11 Conclusion: Under these conditions, slightly more than 20790 megarad of gamma radiation are required to sterilize canned green peas. 12

TABLE (Continued) Run No.: Can Sizeo Product: Inoculum: Irradiation: Processing Temperature: Incubation Temperature: PB-11 Noo 1 Picnic Green Peas 5,000,000 C. None 230~F 85~F (211 x 400) botulinum 213B Spores Per Can Fo Can No. Days-to-gas 0 Formation Canl1, 2.03 5 Can 2, 2.03 6 7 8 Can 1, 1.54 1 Can 2, 1054 2 3 4 Conclusion: Under these conditions, less than 1.54 to produce sterility. canned green peas require an F0 13

TABLE (Continued.) Run No.o Can Sizeo Product: Inoculum: Irradiationo Processing Temperature: Incubation Temperature: Object PB-12 No. 1 Picnic (211 x 400) Green Peas None None 2300F 85~F Same as Runs PB-7 and PB-9, to determine the Fo required to sterilize noninoculated peas that we are using for our experiments. -FO~~ Can~ No.~ ~Days-to-gas F0 Can No. Formation Can 1, 0.18 Can 2, 0.18 1 2 3 4 49 19 5 3 12 12 16 16 Can 1, 0.31 Can 2, 0o31 5 6 7 8 Can 1, 0o17 Can 2, 0.17 9 10 11 12 13 14 15 16 15 7 7 46 12 16 Can 1, 0.35 Can 2., 0.35 Can 1, 0.52 Can 2, 0.52 17 18 19 20 Can 1, 0o76 Can 2, 0.76 21 22 23 24 25 26 27 28 Can 1, 1o22 Can 2, 1,22 Conclusion: Under these conditions, canned green peas were sterilized by an Fo between 0O35 and 0052u This represents a rather considerable resistance to heat processing by the noninoculated (but naturally contaminated) peas which we used in these experiments. 14

TABLE (Continued) Run No o Can Size: Product: Inoculum: Irradiation: Processing Temperature: Incubation Temperature: Object: PB-13 Noo 1 Picnic (211 x 400) Green Peas None None 230 ~F 85~F Same as Runs PB-7, PB-9 and PB-12, to determine the Fo required to sterilize the noninoculated peas we are using in our experiments Days-to-gas Fo Can No. Formation Can 1, 0.041 1 3 Can 2, 0.041 2 3 3 3 4 3 Can 1, 0.20 5 Can 2, 0.20 6 8 7 11 8 20 Conclusion: An Fo greater than 0.20 peas "as received." is needed to sterilize the canned 15

TABLE (Continued) Run No o Product: Inoculum: Irradiation: Processing Temperature: Incubation Temperature: PB-14 Green Peas 5,000,000 C. botulinum 213B Spores Per Can 0. 465 Megarad. 250~F 85~F Fo Can No. Days-to-gas Fp, Can NOo F~~~~~~~~~~0 Can No.Formation Can 1, 0o60 1 25 Can 2, 0.60 2 29 Can 3, 0.60 3 18 4 21 Can 1, 0.75 5 Can 2, 0.75 6 Can 39 0.75 7 8 Can 1, 1.10 9 Can 2, lo10 10 11 12 Can 1, 0o50 13 Can 2, 0.50 14 Can 3, 0.50 15 21 16 18 Conclusion~ Under these conditions, canned green peas were sterilized by 0.465 megarad of gamma radiation followed with an Fo between 0.60 and 0.75. 16

Run No o Product: Inoculum: Irradiation: Processing Temperature: Incubation Temperature: TABLE (Continued) PB-15 Green Peas 59000,000 Co botulinum 213B Spores Per Can 0.650 Megarad 230"F 85OF Fo Can No. Days-to-gas _____F0__________ Can No.__ _Formation Can 1, 0.92 Can 2, 0.92 1 2 3 4 Can 1, lo12 Can 2, o112 Can 1, 0.80 Can 2, o080 Controls: Can 1, 0.50 Can 2, 0.50 Inoculated. nonirradiated Can 1, lo10 Can 2, olO Noninoculated, nonirradiated Can 1, 0.10 Can 2, 0.10 Noninoculated, nonirradiated Can 1, 0O39 Can 2, 0.39 Inoculated. nonirradiated Irradiated only (0O650 megarad) Inoculated Noninoculated 5 6 7 8 9 10 11 12 22 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 8 8 8 8 4 4 4 4 8 8 8 8 4 4 1 2 1 1 Conclusions: (a) Under these conditions, canned green peas were sterilized by 0.650 megarad of gamma radiation followed by an Fo between 0.80 and Oo92. (b) The nonirradiated, noninoculated control cans required an Fo between 0o10 and lo10 for sterilizationo (c) An arradiation dose in excess of 0.650 megarad was needed to sterilize the noninoculated canned peas. 17

TABLE (Continued) Run No.: Product: Inoculum: Irradiation: Processing Temperature: Incubation Temperature: PB-17 Green Peas 5,000,000 C. botulinum 213B Spores Per Can 1.96 Megarad 230~F 85 "F Days-to-gas Fo Can No. Formation Noninoculated a -4 control. b Inoculated A 6 control Can 1, 0.33 1 Can 2, 0.33 2 Can 3, 0.33 3 4 Can 1, 0.22 5 Can 2, 0.22 6 Can 39 0.22 7 8 Can 1, 0.11 9 Can 2, 0ll 10 Can 3, 0oll 11 12 Can 1, 0o06 13 - Can 2, 0.06 14 Can 3, 0.06 15 16 Conclusion: Under these conditions, canned green peas were sterilized by followed by an Fo less than 0,06. and after one month's incubation, lo96 megarad of gamma radiation

TABLE (Continued) Run No o Products Inoculum: Irradiation: Processing Temperature: Incubation Temperature: PB-18 Green Peas 5,000,000 C. botulinum 213B Spores Per Can 1.63 Megarad 230~F 85~F Fo Can No. Days-to-gas Formation Controls-same as on Run No. PB-17 Can No. 1, 0.64 1 Can No. 2, 0.64 2 Can No. 3, 0.64 3 4 Can No. 1, 0.30 5 Can No. 2, 0.30 6 Can No. 3, 0.30 7 8 Conclusions: Under these conditions, and canned green peas were sterilized by 1.63 followed by an Fo less than 0.30. after one month's incubation, megarad of gamma radiation 19

TABLE (Continued) Run No.: Product:t Inoculum: Irradiation: Processing Temperature: Incubation Temperature: PB -19 Green Peas 5,000,000 C. botulinum 213B Spores Per Can As Indicated None 85~F Megarad Can. No. Days-to-gas Megarad Can No. om n Formation Inoculated control 17 6 Noninoculated controls NI 4 NI 4 2.79 1 28 2 3 28 4 3.29 5 6 7 8 3.76 9 10 11 12 2.35 153 28 14 25 15 253 16 25 Conclusions: Under these conditions, and after one month's incubation, canned green peas were sterilized by between 2.79 and 3529 megarad of gamma radiationo 20

2. le% 1. 0. 0 OA — 8.6 i I I IsI.==~~1~'= ----- 1 ------- 0. 0. I I It I (,) w. H D z 2 L2 0.1,~o.u c.0 (.0; 0 ~~~~~~~~.04, -::::.01 0 1.0 2.0 3.0 4.0 MEGARAD Fig. 1. Fo required to sterilize green peas packed in No. 1 Picnic tin cans, inoculated with 5,000,000 C. botulinum 213B spores per can, and irradiated with gamma rays from Cobalt-60 before heat processing at 230~F and incubating at 85~F. 21

PHASE II DETERMINATION OF RADIATION STERILIZATION DOSE FOR CANNED MEAT Five experiments have been run and the cans are now incubating. One experiment was conducted with raw ground beef; the other four are at various C. botulinum 213B spore concentrations using cooked ground beef in 202 x 202 cans. However, incubation has only been in progress for a few weeks, so it would be premature to report data for these experimentso Spore concentrations as high as 100l,000 per gram have been included in the work as of this date. PHASE III DETERMINATION OF COMBINED IRRADIATION-HEAT PROCESSING TREATMENTS REQUIRED TO STERILIZE CANNED MEAT PRODUCTS Eight experiments utilizing "Savortite" Pure Pork Luncheon Meat, furnished by Swift and C o, have been run. To date only the controls have developed gas, so it would also be premature to report these datao Since long incubation periods are apparently going to be needed before results are available, the eight runs were made at various combined irradiationheat processing treatments that, in our judgment, would yield information regarding suitable treatmentso The validity of our estimates will be determined only by observations after a few more montrhs of incubationo Therefore evaluation of results may be delayed for some monthso At present we are continuing Phase II of the work, working principally with Co botulinum 213B in cooked meat. When this series is in the incubator, we will work on the raw meat. Our Cobalt-60 source is to be modified soon. This may interrupt some of the irradiation work for a couple of weeks; this should give us time to collect more spores. STATEMENT OF PROGRESS (a) Approximately 40q of the contracted work is complete as of this date. 22

(b) About half of the budgeted $25,000 have been spent as of this date. (c) It is probable that the work can be completed within the budget, provided that a contract for continuation of work in this general area is again sponsored for us by the Quartermaster Corpso However, if such a contract is not forthcoming for next year, we will need to reduce expenses severely the last two months which represent extension of the time period, by the Quartermaster Corps, beyond our originally calculated expenditure schedule. 253

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