Awards issued prior to january 20, 2025, were funded under previous administrations and may not reflect the priorities and policies of the current administration.** challenges in the production of…

Awards issued prior to january 20, 2025, were funded under previous administrations and may not reflect the priorities and policies of the current administration.** challenges in the production of hydrolysate and other milk-derived products center on the efficient generation of a high-yield product. Enzymes enable reaction processes to take place under less harsh conditions than chemical-only methods, increase product specificity, and reduce the difficulty of reaction processing and waste cleanup. The use of enzymes is not without its own challenges. The primary use limitations come from the cost of the enzyme, and that enzymes in their purified form are limited to a single batch reaction. Recovery of enzyme from the reaction mixture for potential reuse in dairy processing is difficult since the substrates, products, and the enzymes all possess similar physical properties. Other cost-drivers are tied to the need to inactivate the enzyme to stop a reaction, and removal of the enzyme from the final product if the enzyme can cause an allergic or toxic response.to enable the reuse of enzymes, studies have investigated ultrafiltration membranes scaledto retain the larger-size enzymes and simultaneously function as the first purification step, orbinding enzymes to the membranes to combine the reaction and separation processes. These membrane-based solutions demonstrated efficiency gains but suffered from increased enzyme deactivation through shear and thermal stress, enzyme leakage into the product, and membrane fouling. As an alternative, enzymes were immobilized on solid supports such as glass, agarose, and silica. Immobilizing enzymes to these supports enabled improved regulation of the reactions, increased enzyme stability, reduced pressure drop issues in the reactors, improved separation of product from enzyme, and reduced contamination. However, the value gained by these improvements was offset by the high price of the enzyme support material. The technology demonstrated in phase i and to be advanced in the proposed phase II sbir utilizes immobizyme, guild biosciences patented immobilization platform where active components, such as enzymes, are chemically bound to inactive biodegradable support materialsconsisting primarily oflow-cost or recycled BIO-waste. These raw materials offer significant cost savings from traditional immobilization supports, enable higher-quality enzyme preparations, and simplify post-reaction recycling/reuse processes, while retaining the advantages of more expensive support materials. The overall aim of the phase II project is to develop immobizyme into a market-ready product for whey and milk hydrolysis. Phase i efforts demonstrated that these pellets retained more than 90% of enzyme activity through 350 hours of continuous reaction with a per batch material loss of less than 5%. Producing cost-effective immobizyme pellets at-scale is essential to the success of the project. IE must remain active and whole through multiple consecutive batch reactions, have a simple and efficient recovery process, not carry microbial contamination to subsequent reactions, and meet regulatory require, ments for contact with materials for human consumption. Our plan is to start optimization based on performance requirements recommended through our relationships with 4 U.S. Producers. Transitioning this proof of concept material to a product will be achieved by: 1) improving the durability of the pellets so the per batch material loss is less than 1%, 2) defining temperature-related performance limitations and optimums, 3) development of an effective pellet cleaning process that maximizes pellet reuse while minimizing batch-to-batch contamination, 4) developing a cost-efficient, uniform, quality, pilot-scale production process capable of kilogram scale batches, 5) demonstrate the performance of the new material internally and with the support of external facilities, and 6) proceeding towards regulatory approval for the immobizyme platform. Results from phase II will support the technical and economic advantages of using immobizyme pellets as a drop-in replacement for current whey ormilk hydrolysis enzymes.