陶然 刘富菁 王娅玲

摘      要：以水溶性总蛋白及总多糖双指标优化中药乌贼墨酶解法提取工艺并进行了体外抗肿瘤活性研究。以中药乌贼墨总蛋白、总多糖为指标，采用单因素实验，分别考察酶解时间、加酶量和酶解pH值对双指标的影响，在此基础上，应用响应面试验设计法优化中药乌贼墨酶解提取工艺，最后在最佳酶解提取工艺的条件下，采用MTT法对酶解物冻干粉进行体外抗肝癌Bel-7402细胞株的实验研究。结果最佳工艺条件为：酶解时间4.50 h，加酶量为2 500 u，pH为10.6;在此条件下，乌贼墨水溶性总蛋白的得率最高为38.34%，水溶性总多糖的得率最高为12.26%。最佳条件下制得的乌贼墨酶解液对肝癌Bel-7402细胞株有一定的抑制作用，抑制率达到70%～80%。通过响应面法优化乌贼墨酶解工艺条件，使水溶性总蛋白及总多糖得率增加，为乌贼墨活性成分及抗肿瘤研究奠定基础。

关  键  词：乌贼墨;酶解;响应面法;水溶性总蛋白;水溶性总多糖;肝癌Bel-7402细胞株

中图分类号：R282.77       文献标识码： A       文章编号： 1671-0460（2020）05-0855-05

Abstract： The extraction process of squid ink was optimized， and the antitumor activity in vitro was studied. Taking the total protein and total polysaccharide of squid ink as indexes， the effect of hydrolysis time， amount of enzyme and PH value of hydrolysis on the two indexes was investigated by single factor experiment. On this basis， the extraction process of squid ink was optimized by response surface design. Finally， under the optimum conditions of enzyme hydrolysis and extraction， MTT method was used to study the effect of lyophilized powder of enzyme hydrolysate on liver cancer Bel-7402 cell line in vitro. The optimum conditions were determined as follows： the hydrolysis time 4.50 h， the amount of enzyme 2 500 u， and the pH 10.6. Under these conditions， the highest yield of water-soluble total protein was 38.34%， and the highest yield of water-soluble total polysaccharide was 12.26%. Under the optimum conditions， the squid ink hydrolysate could inhibit the liver cancer Bel-7402 cell line to a certain extent， and the inhibition rate was 70%～80%. After the enzymatic hydrolysis conditions of squid ink were optimized by response surface method， the yield of water-soluble total protein and total polysaccharide was increased， which laid a foundation for the study of active components and antitumor activities of squid ink.

Key words：Squid ink; Enzymolysis; Response surface method; Water soluble total protein; Water soluble total polysaccharide; Liver cancer cell line Bel-7402

乌贼墨存在于乌贼的墨囊中，由黑色素和蛋白多糖复合体组成，其蛋白质和糖类的含量分别为6.3%、3.0%。现代研究表明，乌贼墨中的多糖、蛋白质、多肽等活性物质均具有良好的抗肿瘤、抗氧化等作用。酶解技术在中药有效成分提取中具有明显优势，其可在温和的条件下提高中药有效成分的浸出率，保证药物有效成分的药效[1]。同时酶可使蛋白质、糖蛋白等成分水解成小分子活性肽或多糖类物质，从而有效地发挥其药理作用。……