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

item Coll, Brian
item Garcia, Rafael
item Marmer, William

Submitted to: PLoS One
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 12/1/2006
Publication Date: 2/1/2007
Citation: Coll, B., Garcia, R.A., Marmer, W.N. 2007. Diffusion of Protease into Meat & Bone Meal for Solubility Improvement and Potential Inactivation of the BSE Prion. PLoS ONE. 2(2):e245.

Interpretive Summary: Agricultural proteins are a very promising feedstock for bio-based products and materials. The technology to use proteins, such as soy protein isolate and whey, to replace petroleum-based products is developing steadily. Meat & bone meal (MBM) protein, however, suffers some important disadvantages which have so far limited its use in these new applications. It has very low solubility compared to many other commodity proteins. Additionally, even though the risk of prion contamination in North American MBM is insignificant, BSE fears might limit consumer acceptance of MBM-based products. This research project explored the potential of special enzymes to overcome both of these barriers. One enzyme in particular, Versazyme, is attractive because it is able to destroy BSE prions and yet it is cheap enough that it is sold as a poultry feed additive. Practical use of Versazyme or a similar product to decontaminate and solubilize MBM protein involves technical challenges. If the enzymes can only attack MBM particles at the surface, then they will be inefficient at digesting MBM. This research showed that the enzyme can penetrate deep into both the meat and the bone particles of MBM. Further, the benefit of treatments to speed enzyme penetration was studied. Simply allowing MBM to rehydrate had the greatest effect; defatting and milling were also beneficial. This research shows that MBM should be a considered as a feedstock for any new application requiring soluble, prion-free protein.

Technical Abstract: 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 prions. 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. 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 4 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. 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.