β-半乳糖苷酶的固定化及其合成低聚半乳糖研究  

Study on Immobilization ofβ-Galactosidase and Its Synthesis of Galacto-Oligosaccharide

作　　者：李智慧 赖田甜 张敏 郝瑞敏 姚梦珂 赵华[1] 杨贞耐[1] LI Zhihui;LAI Tiantian;ZHANG Min;HAO Ruimin;YAO Mengke;ZHAO Hua;YANG Zhennai(Key Laboratory of Geriatric Nutrition and Health of Ministry of Education,Beijing Technology and Business University,Beijing 100048,China;Key Laboratory of Agro-Products Primary Processing,Academy of Agricultural Planning and Engineering,Ministry of Agriculture and Rural Affairs,Beijing 100125,China)

机构地区：[1]北京工商大学老年营养与健康教育部重点实验室,北京100048 [2]农业农村部规划设计研究院农业农村部产地初加工重点实验室,北京100125

出　　处：《食品科学技术学报》2024年第6期102-112,共11页Journal of Food Science and Technology

基　　金：国家重点研发计划专项(2017YFE0131800)。

摘　　要：选用壳聚糖、海藻酸钠、海藻酸钠/明胶、海藻酸钠/羧甲基纤维素为固定化载体材料,探究β-半乳糖苷酶的固定化方法,并将获得的适宜固定化酶应用于合成低聚半乳糖(galacto-oligosaccharide,GOS)。结果表明,较优固定化载体为壳聚糖,β-半乳糖苷酶的优化固定化条件为壳聚糖质量浓度0.03 g/mL,静置时间2 h,戊二醛体积分数2%,交联时间3 h,吸附时间12 h,固定化温度60℃,加酶量0.4 mg/g(以壳聚糖微球质量计)。与游离酶相比,固定化酶的耐酸、碱、热的稳定性显著提升。进一步利用壳聚糖固定化β-半乳糖苷酶合成GOS,确定了优化反应条件:加酶量为2 U/mL,pH值为6.0,反应温度为50℃,初始乳糖质量浓度为500 g/L,反应时间为14 h。此条件下GOS产率可达到52.61%,高于已报道的GOS产率(25.1%~46.0%),且产物中聚合度大于等于3的GOS含量为37.07%。固定化酶重复使用5次后,产GOS活性仍保留了97.21%,研究获得的壳聚糖固定化β-半乳糖苷酶在GOS合成领域具有良好的应用前景。Chitosan,sodium alginate,sodium alginate/gelatin and sodium alginate/carboxymethyl cellulose were used as immobilization carrier materials to investigate the immobilization methods ofβ-galactosidase,and the synthesis of galacto-oligosaccharides(GOS)by the suitable immobilized enzyme was studied.The results showed that the obtained suitable immobilization carrier was chitosan,and the optimal immobilization conditions forβ-galactosidase were chitosan mass concentration of 0.03 g/mL,standing time of 2 h,glutaraldehyde volume fraction of 2%,crosslinking time of 3 h,adsorption time of 12 h,immobilization temperature of 60℃,enzyme dosage of 0.4 mg/g(based on the mass of chitosan microspheres).Compared with the free enzymes,the stability of immobilized enzymes against acid,alkali and heat was significantly improved.Chitosan immobilizedβ-galactosidase was further used to synthesize GOS,and the optimal reaction conditions were determined as follows:amount enzyme of 2 U/mL,pH of 6.0,reaction temperature of 50℃,initial lactose mass concentration of 500 g/L,and reaction time of 14 h.Under these conditions,the GOS yield reached 52.61%,which was higher than those reported GOS yield(25.1%~46.0%).And the content of GOS with polymerization degree greater than or equal to 3 in the product was 37.07%.After repeated use of the immobilized enzyme for five times,the GOS production activity remained 97.21%.The chitosan-immobilizedβ-galactosidase obtained from the study had a promising application in the field of GOS synthesis.

关 键 词：Β-半乳糖苷酶 低聚半乳糖合成 固定化方法 壳聚糖 交联 

分 类 号：TS245.5[轻工技术与工程—制糖工程]