葡萄糖氧化酶固定条件的优化

摘要

The immobilization conditions of glucose oxidase (GOD) were studied using cellulose acetate (CA) film as carrier.CA film was prepared by solution casting method using CA as raw materials.Fluorescent indicator was immobilized on CA film by physical embedding method.The GOD was immobilized on the film surface by the crosslinking agent glutaraldehyde.The effect of sodium periodate, ethylenedia-mine, glutaraldehyde, enzyme content given and pH on the activity of immobilized GOD were studied.The optimal immobilization conditions were determined.The film for the optimal immobilization conditions was obtained.The optimal immobilization conditions were determined; concentration of sodium periodate 0.5 mol/L,reaction time 30 min, concentration of ethylenediamine 0.04 mol/L, reaction time 150 min, concentration of glutaraldehyde 1.5％ (v/v).reaction time 120 min,pH value 6.5 and enzyme content given 35 mg/mL,reaction time 21 h and temperature 4 ℃.SEM analysis showed that the quality of film was better and the fluorescent indicator was evenly distributed in the film.By optimizing the immobilization conditions, both light sensitivity and enzymatic activity of complex sensing film was obtained via immobilization of the fluorescent indicator and GOD on cellulose acetate film.%以醋酸纤维素(CA)为原料,采用溶液浇铸法成膜.用物理包埋法将荧光指示剂固定CA膜中.以戊二醛为交联剂,将葡萄糖氧化酶固定于膜表面.研究了高碘酸钠、乙二胺、戊二醛、给酶量、pH值等固定化条件对酶活的影响,确定了优化的固定条件:在室温下与0.5 mol/L的高碘酸钠反应30 min;与0.04 mol/L的乙二胺反应150 min时;与溶度为1.5%(v/v)的戊二醛交联120 min;在4℃条件下,在pH 6.5、给酶量35 mg/mL的酶液中交联21 h.利用SEM分析表明,荧光指示剂均匀分布膜中,成膜质量较好.通过优化固定条件,把荧光指示剂和葡萄糖氧化物酶(GOD)同时固定醋酸纤维素膜上,可以得到同时具有光敏感性和酶催化能力的复合敏感膜.