SBIR-STTR Award

A challenge in the production of dairy-derived hydrolysate products is efficiently generating a high- yield and in the case of peptides specific and active material. Enzymes enable the use of mild reaction conditions and processing techniques compared to standard chemical methods. However the use of commercial enzymes and enzyme mixes presents its own challenges. High quality commercial enzyme products are expensive for a single use and some enzymes can produce allergic or toxic responses if left in products. Extensive purification steps after enzyme reactions adds to production expenditures. A potential method to control process costs is to immobilize the enzymes enabling enzyme recovery and reuse. The aim of the project is to demonstrate the capabilities of immobilized enzymes for the processing of whey products compared to their soluble enzyme counterparts. Our patented immobilization technology will be applied to two commercial protease mixes Alcalase and Protamex and to beta- galactosidase (lactase); converting them from one-time use catalysts to reusable materials. Their activity across pH and temperature conditions and capability to retain enzymatic activity overextended use will be determined. Test results from Phase I will validate immobilized enzymes as are placement for current materials through: 1) a similar reactive capacity vs. soluble enzyme 2)extended enzymatic stability under defined reaction conditions 3) retaining enzyme activity following standard clean in place protocols 4) resistance to microbial growth 5) ease of separation from reaction products and 6) minimal to no enzyme leaching into reaction solutions.

A challenge in the production of dairy-derived hydrolysate products is efficiently generating a high-yield and in the case of peptides specific and active material. Enzymes enable the use of mild reaction conditions and processing techniques compared to standard chemical methods. However the use of commercial enzymes and enzyme mixes presents its own challenges. High quality commercial enzyme products are expensive for a single use and some enzymes can produce allergic or toxic responses if left in products. Extensive purification steps after an enzyme-driven reaction adds to production expenditures. A potential solution to control process costs is to immobilize the enzymes enabling enzyme recovery and reuse. The objectives of the Phase II work are to build on successes from the Phase I effort ultimately producing immobilized enzyme products that meet the needs of current whey-based product manufacturers. Two commercial proteases Alcalase® and Protamex® and a -galactosidase will continue to be developed using our patented immobilization technology ImmobiZyme™. In Phase I we demonstrated the enzymes retained their functionality following immobilization and their susceptibility to microbial contamination and cleaning was evaluated. More significantly these proof-of-concept pellets maintained >90% of their starting enzymatic activity following 350 hours reacting with substrates. Phase II efforts will focus on taking these materials from to products through the following objectives: 1) optimizing pellet stability with respect to activity under reaction conditions 2) developing a pellet-compatible cleaning protocol for removal of microbial contaminants between uses 3) producing immobilized enzymes at the pilot-scale 4)performing initial toxicology studies and 5) demonstrating performance under pilot-scale conditions both internally and through the efforts of potential vendors. The primary markets for the technology developed during this Phase II effort will be milk protein hydrolysates (MPH) and lactose free (LF) products. The 2018 MPH market volume was estimated at 84.7 million kg and the reported 2020 U.S. LF milk product market volume is estimated at 804 million kg. This technology also has potential to be utilized for hydrolysis of other protein sources (wheat pea soy and insect) for hydrolysate and bioactive peptide markets. The primary market advantage of ImmobiZyme™ comes from its potential for cost savings. The number of times it can be reused is critical to the economics of this application. Reuse is mainly determined by retained activity and retained pellet on a batch-to-batch basis. Based on a potential 30 cycle lifetime for the ImmobiZyme™ pellets from the Phase I data the retained activity andmass fraction of enzyme on the pellets and current enzyme use rates (typically 1%-5% of the mass of substrate) for these applications; the potential combined market for ImmobiZyme™-