SBIR Phase I: Development of an enzymatic method to produce compounds found in human milk at commercial scale

这项小企业创新研究（SBIR）第一阶段项目的更广泛/商业影响是评估一种经济实惠的酶促方法，用于生产通常在人乳中发现的可销售的天然化合物。一旦这一战略得到验证，就可以由国内和工业用户实施。这项技术能够将牛奶中的乳糖转化为非常理想的化合物。这些稀有而有价值的化合物可以帮助促进有益肠道细菌的生长，是人类母乳的组成部分。这些化合物被广泛用于强化婴儿食品，它们在肠道健康中发挥作用，支持平衡的肠道微生物群，有益免疫，改善认知大脑健康。此外，新的研究表明，含有稳定的生物活性天然化合物的负担得起的食品强化技术将有益于婴儿和整个生命阶段的健康肠道。因此，这项技术为旨在开发独特的肠道强化营养解决方案的食品和膳食补充剂制造商开辟了新的商机。拟议中的项目将酶促生成在人乳中发现的化合物，包括n -乙酰氨基葡萄糖（LacNAc）。研究计划包括测试酶β-己糖基转移酶（BHT）生产的可扩展性，该酶将由K. phaffi异种生产。为了验证工业可扩展性、产品生成和100 L工作体积生物处理反应器的产量，生成的BHT将用于催化半乳糖向n -乙酰氨基葡萄糖（GlcNAc）的重复添加。产品，除了半乳糖低聚糖，将包括LacNAc双糖，由顺序的半乳糖基化反应产生。回收的产品将在临床前安全性/毒性研究中进行测试。在这项研究中收集的数据将允许更精确的成本效益分析，其中将包括产品产量，碳平衡，微生物组效益和代谢数据。由于BHT的特异性、一步反应合成、低成本底物、可持续性和整体低环境影响，一种更有效的酶促生物合成方法有望节省成本。该奖项反映了美国国家科学基金会的法定使命，并通过使用基金会的知识价值和更广泛的影响审查标准进行评估，被认为值得支持。

The broader/commercial impact of this Small Business Innovation Research (SBIR) Phase I project is to assess an affordable enzymatic method for producing marketable natural compounds usually found in human milk. Once this strategy is validated, it could be implemented by domestic and industrial users. The technology is capable of transforming the lactose in milk to highly desirable compounds. These rare and valuable compounds can help enhance growth of beneficial gut bacteria and are building blocks of human breast milk. These compounds are widely desirable to fortify infant food, where they play a role in intestinal health, supporting balanced gut microbiota, benefiting immunity, and improving cognitive brain health. Furthermore, new research suggests that affordable technologies for food fortification containing stable bioactive natural compounds will benefit the healthy gut beyond infancy and across life stages. As such, this technology opens new business opportunities for food and dietary supplement manufacturers aiming to develop unique gut-strengthening nutrition solutions. The proposed project will enzymatically generate compounds found in human milk, including N-acetylglucosamine (LacNAc). The research plan consists of testing the scalability of enzyme β-hexosyltransferase (BHT) production, which will be heterologously produced by K. phaffi. To validate industrial scalability, product generation, and yields from 100 L working volume bioprocessing reactors, the BHT generated will be utilized to catalyze the repeated addition of galactose to N-acetylglucosamine (GlcNAc). The products, in addition to galactooligosaccharides, will include LacNAc disaccharides, generated by sequential transgalactosylation reactions. The recovered products will be tested in preclinical safety/toxicity studies. Data collected during this study will allow for a more precise cost-benefit analysis, which will include product yields, carbon balances, microbiome benefits, and metabolic data. Cost savings are expected from a more efficient enzymatic biosynthesis method for producing LacNAc due to BHT specificity, synthesis in one-step reactions, low-cost substrates, sustainability, and overall low environmental impact.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.