Diffusion of Protease into Meat & Bone Meal for Solubility Improvement and Potential Inactivation of the BSE Prion

BACKGROUND: Government-imposed feed bans have created a need for new applications for meat & bone meal (MBM). Many potential new applications require MBM protein to be both soluble and free of infectious prion. Treatment with protease is generally effective in reducing insoluble, thermally-denatured proteins to soluble peptides. It has been reported in the literature that certain proteases, including Versazyme™, are able to degrade infectious prions in a system where the prions are readily accessible to proteolytic attack. Prions distributed within MBM, however, may conceivably be protected from proteases. METHODOLOGY/PRINCIPAL FINDINGS: The overall rate of proteolytic MBM digestion depends greatly on whether the protease can penetrate deep within individual particles, or if the protease can only act near the surface of the particle. This research examined the barriers to the diffusion of Versazyme™ into particles of MBM. Confocal microscopy demonstrated differences in the density distributions between the bone and the soft tissue particles of MBM. By tracking the diffusion of fluorescently labeled Versazyme™ through individual particles, it was found that bone particles show full Versazyme™ penetration within 30 minutes, while penetration of soft tissue particles can take up to four hours, depending on the particle's diameter. From the variety of normal proteins comprising MBM, a specific protein was chosen to serve as a prion surrogate based on characteristics including size, solubility, distribution and abundance. This surrogate was used to measure the effect of several factors on Versazyme™ diffusion. CONCLUSIONS/SIGNIFICANCE: Results showed that surrogate distributed in bone particles was more susceptible to degradation than that in soft tissue particles. Three factors controllable by unit operations in an industrial-scale process were also tested. It was found that removing the lipid content and hydrating MBM prior to incubation both significantly increased the rate of surrogate degradation. In a test of particle size, the smallest collected diameter range demonstrated the largest degradation of the prion surrogate, suggesting milling would be beneficial