新型柱前衍生-高效液相荧光检测法检测水产品中河豚毒素  

Identification of tetrodotoxin in aquatic products by a new method using pre-column fluorimetric-derivatization high-performance liquid chromatography

作　　者：辛少平[1,2] 岑剑伟[1] 李来好[1] 杨贤庆[1] 郝淑贤[1] 魏涯[1] 周婉君[1] 王子怀[1] 

机构地区：[1]中国水产科学研究院南海水产研究所、农业部水产品加工重点实验室、国家水产品加工技术研发中心,广东广州510300 [2]上海海洋大学食品学院,上海201306

出　　处：《中国水产科学》2015年第1期139-148,共10页Journal of Fishery Sciences of China

基　　金：广东省科技计划项目(2012B040302008);中央级公益性科研院所基本科研业务费专项(2012TS21)

摘　　要：在稀碱条件下,河鲀毒素（TTX）与次溴酸钠和尿素于一定温度下反应生成具有荧光性能的物质,该反应与已报道的TTX在强碱下生成C-9碱的碱解不同。本研究拟对该衍生反应所需条件进行详细研究,并尝试依据该反应建立鲀一种新型的河毒素荧光检测方法。荧光扫描光谱显示,产物最大激发波长（EX）为233 nm,最大发射波长（EM）为370 nm。该反应对pH要求比较严格,反应最适pH在11.6~11.9之间。对样品进行衍生检测获得最强荧光信号的条件为：次溴酸钠0.036~0.09 mol/L、尿素10~50 g/L、碳酸钠0.2~0.3 mol/L,温度75℃。样品检测结果显示,此衍生反应具有高度专一性,信号强度与 TTX 含量成正比,在0.01~10μg/mL 质量浓度范围内线性方程为y=109.17x＋0.3965,线性相关系数为0.999。依据该衍生反应建立柱前衍生–高效液相色谱荧光检测法检测水产品中河鲀毒素含量,检测限达到20μg/kg,平行样品检测相对标准偏差为0.44%~7.25%,准确度高。对河鲀、虾虎鱼和织纹螺产品检测表明,该反应特异性强,不易出现假阳性,鲀可用于河毒素检测的确证。Tetrodotoxin （TTX）, one of the most toxic natural neurotoxins found in several kinds of fishery products, can cause TTX poisoning. Therefore, detection methods for routine analysis of TTX should be developed. We found that TTX could be converted into a new compound with fluorescence properties by reaction with sodium bromate （NaBrO） and urea. The reaction was sensitive to the acidity level of the solution and the pH value needed to be strictly controlled between 11.6 and 11.9. The fluorescence scanning spectrum showed the fluorescent compound had maximum excitation wavelength of 233 nm and emission wavelength of 370 nm, and was not the product of alkaline degradation （quina-zoline）. Hence, a new type of fluorescence detection method was developed for identifying TTX in aquatic products. After reaction with NaBrO （0.036–0.09 mol/L） and urea （10–50 g/L） in 0.2–0.3 mol/L sodium carbonate solution （Na2CO3） at 75℃, the TTX derivative solution was neutralized with 15% phosphoric acid and then detected by re-verse-phase high-performance liquid chromatography with a fluorescence detector. The result was quantified by the standard addition method. The detection limit was 20μg/kg and the relative standard deviation was 0.44%–7.25%. The method was specific, accurate, and sensitive, and can be used for identification of TTX determination.

关 键 词：河鲀毒素 柱前衍生 次溴酸钠 尿素 荧光检测 

分 类 号：O657.7[理学—分析化学]