台盼蓝法测定发酵液中ε-聚赖氨酸产量  

Determination ofε-poly-L-lysine production in fermentation broth by trypan blue

作　　者：张存金 潘龙 韩金洋 宋登科 刘莹莹 蔡婷 廖爱美 侯银臣 宇光海[1] 惠明[1] 黄继红[1,4,5] ZHANG Cunjin;PAN Long;HAN Jinyang;SONG Dengke;LIU Yingying;CAI Ting;LIAO Aimei;HOU Yinchen;YU Guanghai;HUI Ming;HUANG Jihong(Henan Provincial Key Laboratory of Biological Processing and Nutritional Function of Wheat,School of Biological Engineering,Henan University of Technology,Zhengzhou 450001,China;Henan Jindan Lactic Acid Technology Co.,Ltd.,Zhoukou 477150,China;School of Food and Bioengineering,Henan University of Animal Husbandry and Economy,Zhengzhou 450046,China;School of Agriculture,Henan University,Kaifeng 475001,China;School of Food and Pharmacy,Xuchang University,Xuchang 461000,China)

机构地区：[1]河南工业大学生物工程学院,小麦生物加工与营养功能河南省重点实验室,河南郑州450001 [2]河南金丹乳酸科技股份有限公司,河南周口477150 [3]河南牧业经济学院食品与生物工程学院,河南郑州450046 [4]河南大学农学院,河南开封475001 [5]许昌学院食品与药学院,河南许昌461000

出　　处：《河南工业大学学报（自然科学版）》2023年第4期92-97,105,共7页Journal of Henan University of Technology：Natural Science Edition

基　　金：河南省博士后科研项目资助(202103120);河南省青年基金项目(232300421266);河南工业大学高层次人才基金项目(2020BS064)。

摘　　要：ε-聚赖氨酸是一种抑菌效果好、耐高温的天然防腐剂,在食品中有良好的应用前景。建立台盼蓝法测定发酵液中ε-聚赖氨酸产量,以台盼蓝试剂为显色剂,考察了显色剂浓度、反应温度、反应时间、pH值等因素对检测结果的影响。结果表明:最优的试验条件为显色剂质量浓度为3 g/L、pH值为6、反应温度为50℃、反应时间为40 min,标准曲线为Y=-0.0128X+3.1488;线性范围为10~100 mg/L,ε-聚赖氨酸工作曲线线性决定系数(R 2)为0.9998,最低检测限(LOD)为2.03 mg/L,最低定量限(LOQ)为6.14 mg/L,不同浓度标准品的测定结果变异系数(CV)均小于2%,检测样品的加标回收率为98.96%~101.53%;测定结果不受发酵液中底物影响,且能够有效地缩短检测时间。台盼蓝法的建立对ε-聚赖氨酸高产菌株的高通量选育具有重要的意义。ε-poly-L-lysine(ε-PL)is a natural preservative with good antibacterial effect and high temperature resistance,which has a good application prospect in food.ε-PL is a similar amino acid polymer formed by L-lysine monomer by synthesis-catalytic condensation ofα-COOH andε-NH 2.This polymer was isolated by Japanese scholars Shima and Sakai in the fermentation broth of Small White Streptomyces Streptomyces albulus 346 in 1977.The free amino group on theε-PL backbone facilitates it to exhibit cationic properties in acidic to weakly alkaline environments.Therefore,ε-PL is able to bind to anions on the cell membrane surface and damages cell membranes of microbes,including Gram-positive and Gram-negative bacteria,fungi and yeast.In 2014,China officially approved its use for the preservative and preservation of starch products,baked foods,meat products,beverages,condiments and other foods.ε-PL has become the third kind of microbial source natural food preservative approved in China.The trypan blue method was established for the determination ofε-PL in the fermentation broth.The side chain ofε-PL is reported to contain large amounts of amino cation,while trypan blue contains a sulfonate anion.According to the precipitation principle ofε-PL and methyl orange,it was found thatε-PL and trypan blue were also able to form a complex and precipitate in the range of pH 4-10.After centrifugation,the supernatant showed a good linear relationship between its absorbance value and theε-PL concentration at 600 nm.Trypan blue reagent was used as the chromatic reagent.Several factors including the concentration of chromatic reagent,reaction temperature,time,and pH,were investigated to optimize the test conditions.The optimal reaction conditions were determined as follows:the concentration of trypan blue was 3 g/L,pH 6.0,reaction temperature was 50℃and reaction time was 40 min,the standard curve was Y=-0.0128X+3.1488.The calibration curve was linear in the concentration range of 10-100 mg/L.The linear correlation coefficient(

关 键 词：Ε-聚赖氨酸 台盼蓝 回收率 选育 

分 类 号：TS207.3[轻工技术与工程—食品科学]