基于微孔板比色法的微生物胞内油脂快速检测方法的建立及应用  

Establishment and application of a rapid microbial intracellular lipid determination method based on colorimetric microplate format

作　　者：胡元维[1] 任路静[1] 孙冠男[1] 张丽[1] 李娟[1] 黄和[1] 

机构地区：[1]南京工业大学生物与制药工程学院,南京210009

出　　处：《中国油脂》2013年第7期89-94,共6页China Oils and Fats

基　　金：国家科技支撑计划(2011BAD23B03);"863"项目(SS2012AA021704);国家"973"计划项目(2009CB724700)

摘　　要：对磷酸香草醛显色法的反应条件进行优化研究,建立了磷酸香草醛显色法快速检测微生物胞内油脂的方法,并探讨该方法在微生物油脂测定上的应用。经过实验确定最佳比色条件为:将油脂样品蒸干溶剂后加入2 mL 98%硫酸,于90℃水浴20 min,冷却至室温后添加3 mL 0.25 g/L香草醛磷酸溶液(34%的磷酸溶液配制),摇匀室温显色20 min,扣除非油脂成分干扰,在530 nm波长下测定吸光度,经验证该反应体系稳定性良好。在裂殖壶菌和隐甲藻两种产DHA油脂微生物中,通过称重法与磷酸香草醛显色法比较,进一步证实建立的方法分析结果准确可靠,分析过程简便、快速,稳定性好,适合作为常规分析检测方法。To achieve a rapid and accurate determination method for single - cell lipid, sulfo - phospho - vanillin （SPV） reaction conditions were optimized and the application of SPV reaction in determination of microbial intracellular lipid was discussed. The optimal SPV reaction conditions were that oil samples were added with 2 mL of 98% HESO4 keeping in 90 ℃ water bath for 20 rain, after they were cooled to room temperature, 3 mL of vanillin - phosphoric acid solution （ prepared with 34% phosphoric acid solu- tion） was added for color development and absorbance measurement was made at 530 nm after 20 min of color reaction to eliminate the non - oil components interference. Under the optimal conditions, lipid con- tents of Schizochytrium sp. and Crypthecodinium cohnii were detected, and there was no significant differ- ence in lipid determination comparing with macro - gravimetric method. Thus, SPV reaction was proved to be a simple, accurate, rapid method for detecting microbial intracellular lipid which could be used as a routine analytical method for the determination of single - cell lipid.

关 键 词：单细胞油脂 磷酸香草醛反应 称重法 裂殖壶菌 隐甲藻 

分 类 号：TS227[轻工技术与工程—粮食、油脂及植物蛋白工程] O657.3[轻工技术与工程—食品科学与工程]