Microbial survival after isoelectric solubilization and precipitation of fish protein

Protein wasted by the disposal of fish processing byproducts may be recovered using isoelectric solubilization and precipitation. The protein is dissolved by extreme pH shifts and recovered via precipitation and centrifugation. Microbial safety throughout this process had not yet been evaluated; therefore, the purpose of this study was to determine if Listeria innocua and Escherichia coli would survive extreme pH shifts during the protein recovery process. Fresh rainbow trout were headed, gutted, and minced and then inoculated with 109 CFU/g E. coli ATCC 25922 or filleted and minced and then inoculated with 109 cfu/g of L. innocua. The fish was homogenized and brought to the target pH of 2.0, 3.0, 11.5 or 12.5 by the addition of concentrated hydrochloric acid or sodium hydroxide to solubilize the muscle proteins. The homogenate was blended at 4°C for 10 min and centrifuged to separate the lipid and insoluble components (bones, skin, insoluble protein, etc.) from the protein solution. The lipid and insoluble components were removed and the protein solution was subjected to a second pH shift (pH 5.5) resulting in protein precipitation. Centrifugation was applied to separate the precipitated proteins from the water. Each constituent (i.e., lipid, insoluble components, protein, and water) was analyzed for bacterial content using non-selective growth media and selective media. The sums of the surviving bacteria in these fractions were compared to the initial inoculum. For L. innocua there were no significant differences in recovery on growth or selective media (P > 0.05); implying both acidic and basic conditions have an all-or-nothing bactericidal effect on the gram-positive species. The greatest overall microbial reduction occurred when the pH was shifted to 2.0: a total of 3-log reduction in microbes. Compared to the initial inoculum level in the trout filets, there was a 4-log reduction of Listeria cells in recovered protein. For E. coli, the greatest total microbial reduction occurred when the pH was shifted to 12.5 (P < 0.05): a 4.4-log reduction of cells on growth media and a 6.0-log reduction of cells on selective media. Compared to the initial inoculum level in the minced trout, there was a 4.7-log reduction of E. coli cells in recovered protein on selective media. There was significant (P < 0.05) injury sustained by cells exposed to alkaline treatment (pH 11.5 and 12.5) in all fractions except the insoluble fraction at pH 11.5. Increasing the exposure time or the pH, or using a weak organic acid in lieu of a strong acid for Listeria, may result in greater bacterial reductions in the recovered protein.