Billakanti Jagan M, Fee Conan J
Department of Chemical and Process Engineering, University of Canterbury, Private Bag 4800, Christchurch 8140, New Zealand.
Biotechnol Bioeng. 2009 Aug 15;103(6):1155-63. doi: 10.1002/bit.22344.
Extraction and purification of high-value minor proteins directly from milk without pre-treatment is a challenge for the dairy industry. Pre-treatment of milk before extraction of proteins by conventional packed-bed chromatography is usually necessary to prevent column blockage but it requires several steps that result in significant loss of yield and activity for many minor proteins. In this paper, we demonstrate that it is possible to pass 40-50 column volumes of various milk samples (raw whole milk, homogenized milk, skim milk and acid whey) through a 5 mL cryogel chromatographic column at 550 cm/h without exceeding its pressure limits if the processing temperature is maintained above 35 degrees C. The dynamic binding capacity obtained for the cryogel matrix (2.1 mg/mL) was similar to that of the binding capacity (2.01 mg/mL) at equilibrium with 0.1 mg/mL of lactoferrin in the feed samples. The cryogel column selectively binds lactoferrin and lactoperoxidase with only minor leakage in flowthrough fractions. Lactoferrin was recovered from elution fractions with a yield of over 85% and a purity of more than 90%. These results, together with the ease of manufacture, low cost and versatile surface chemistry of cryogels suggest that they may be a good alternative to packed-bed chromatography for direct capture of proteins from milk.
直接从牛奶中提取和纯化高价值微量蛋白质而无需预处理，这对乳制品行业来说是一项挑战。通过传统填充床色谱法提取蛋白质之前，通常需要对牛奶进行预处理，以防止柱堵塞，但这需要几个步骤，会导致许多微量蛋白质的产量和活性显著损失。在本文中，我们证明，如果处理温度保持在35摄氏度以上，各种牛奶样品（生鲜全脂牛奶、均质牛奶、脱脂牛奶和酸性乳清）以550厘米/小时的流速通过5毫升冷冻凝胶色谱柱40 - 50个柱体积时，不会超过其压力极限。冷冻凝胶基质获得的动态结合容量（2.1毫克/毫升）与进料样品中乳铁蛋白浓度为0.1毫克/毫升时的平衡结合容量（2.01毫克/毫升）相似。冷冻凝胶柱选择性结合乳铁蛋白和乳过氧化物酶，流穿组分中只有少量泄漏。从洗脱组分中回收的乳铁蛋白产率超过85%，纯度超过90%。这些结果，连同冷冻凝胶易于制造、成本低和表面化学性质多样，表明它们可能是替代填充床色谱法直接从牛奶中捕获蛋白质的良好选择。
