Departamento de Engenharia Química, Universidade Federal do Ceará, Campus do Pici, Bloco 709, Fortaleza, CE, CEP 60455-760, Brazil.
Instituto de Engenharias e Desenvolvimento Sustentável, Universidade da Integração Internacional da Lusofonia Afro-Brasileira, Acarape, CE, CEP 62785-000, Brazil.
Appl Biochem Biotechnol. 2018 Apr;184(4):1263-1285. doi: 10.1007/s12010-017-2622-1. Epub 2017 Oct 10.
Lipase from Rhizomucor miehei (RML) was immobilized onto chitosan support in the presence of some surfactants added at low levels using two different strategies. In the first approach, the enzyme was immobilized in the presence of surfactants on chitosan supports previously functionalized with glutaraldehyde. In the second one, after prior enzyme adsorption on chitosan beads in the presence of surfactants, the complex chitosan beads-enzyme was then cross-linked with glutaraldehyde. The effects of surfactant concentrations on the activities of free and immobilized RML were evaluated. Hexadecyltrimethylammonium bromide (CTAB) promoted an inhibition of enzyme activity while the nonionic surfactant Triton X-100 caused a slight increase in the catalytic activity of the free enzyme and the derivatives produced in both methods of immobilization. The best derivatives were achieved when the lipase was firstly adsorbed on chitosan beads at 4 °C for 1 h, 220 rpm followed by cross-link the complex chitosan beads-enzyme with glutaraldehyde 0.6% v.v at pH 7. The derivatives obtained under these conditions showed high catalytic activity and excellent thermal stability at 60° and 37 °C. The best derivative was also evaluated in the synthesis of two flavor esters namely methyl and ethyl butyrate. At non-optimized conditions, the maximum conversion yield for methyl butyrate was 89%, and for ethyl butyrate, the esterification yield was 92%. The results for both esterifications were similar to those obtained when the commercial enzyme Lipozyme® and free enzyme were used in the same reaction conditions and higher than the one achieved in the absence of the selected surfactant.
米黑根毛霉脂肪酶（RML）在添加少量表面活性剂的存在下被固定在壳聚糖载体上，使用了两种不同的策略。在第一种方法中，酶在戊二醛预先功能化的壳聚糖载体上存在表面活性剂时被固定。在第二种方法中，在存在表面活性剂的情况下，将酶预先吸附在壳聚糖珠上，然后用戊二醛将壳聚糖珠-酶复合物交联。评估了表面活性剂浓度对游离和固定化 RML 活性的影响。十六烷基三甲基溴化铵（CTAB）促进了酶活性的抑制，而非离子表面活性剂 Triton X-100 导致游离酶和两种固定化方法产生的衍生物的催化活性略有增加。当脂肪酶首先在 4°C 下吸附在壳聚糖珠上 1 小时，220rpm 后用 0.6%（v/v）戊二醛在 pH 7 下交联壳聚糖珠-酶复合物时，获得了最佳的衍生物。在这些条件下获得的衍生物具有较高的催化活性和在 60°C 和 37°C 下的优异热稳定性。最佳衍生物还在两种风味酯即丁酸甲酯和丁酸乙酯的合成中进行了评估。在非优化条件下，丁酸甲酯的最大转化率为 89%，丁酸乙酯的酯化产率为 92%。这两种酯化反应的结果与在相同反应条件下使用商业酶 Lipozyme®和游离酶时获得的结果相似，且高于在没有选择的表面活性剂存在的情况下获得的结果。
