高效液相色谱-三重四极杆质谱同时测定与筛查海洋浮游藻类中的5种色素

Simultaneous determination and screening of five pigments in marine phytoplanktons by high performance liquid chromatography-triple quadrupole mass spectrometry

对海洋藻类色素的高效液相色谱-三重四极杆质谱（HPLC-QqQ-MS）分析方法的有效性进行了验证。该方法采用C16氨基色谱柱为固定相,以甲醇、乙腈和乙酸铵为流动相,在选择反应监测模式下对叶绿素a、叶绿素b、β,β-胡萝卜素、叶黄素和岩藻黄素的含量进行了测定。结果表明,5种色素标准品的线性关系良好,相关系数（r2）均高于0.996,回收率在82.77%-99.83%之间,日内和日间精密度的相对标准偏差（RSD）均小于5%（n=5）;方法的检出限（LOD）在0.02-0.16μg/L之间,定量限（LOQ）在0.06-0.54μg/L之间。利用该方法对11种藻类中5种色素的含量进行分析,比较了赤潮异弯藻、卡罗藻、微小原甲藻、微拟球藻、蛋白核小球藻、颗石藻、定鞭金藻、中肋骨条藻、威氏海链藻和假微型海链藻之间的物种色素分布情况。该方法具有简单、灵敏度高、重复性好、回收率高等优点,适合藻类色素分析,为进行藻类生物量计算提供了分析手段。

A quantitative method based on high performance liquid chromatography coupled with electrospray ionization tandem triple-quadrupole mass spectrometry （HPLC-ESI-QqQ-MS） has been established for five pigments in marine phytoplanktons. The HPLC method used ternary solvent systems and a reversed-phase C16-amide column. In addition, methanol, acetonilrile and aqueous ammonium acetate were used as mobile phases. Five pigments （ chlorophyll a, chlorophyll b, β, β-carotene, lutein and fucoxanthin） were quantified in selective reaction mode. As results, good linear relationships were achieved between the concentrations and the peak areas of the five pigment standards. And their correlation coefficients （ r^2 ） were higher than 0. 996. The recoveries of the pigment standards were between 82.77% and 99.83%. The inter-day and intra-day precisions were lower than 5% （ n = 5 ）. The detection limits of the pigments for this method were between 0.02 and 0. 16 μg/L and the quantification limits were in the range from 0.06 to 0.54 μg/L. According to the above method, eleven algae （Heterosigma akashiwo （NMBRah03-2） . Heterosigma akashiwo （NMBRah03-2-2） , Karlodinium vent fie＇am （ NMBjah047-1 ）, Prorocentrum minimum （ NMBjah042 ）, Nannochlo＇ropsis oceanic （ NMBluh014 ） , Chlo＇reUa pyrenoidosa （ NMBluh015-1 ）, Pleurochrysis sp. （ NMBjih026-1 ） , Prymnesium sp. （ NMBjih029）, Skeletonema costatum （ NMBguh004-1 ） , Thalassiosira weissflogii （NMBguh021） and Thalassiosira pseudonana ） （NMBguh005）） have been investigated for comparing the pigment distributions. The method is sensitive, accurate, reproducible, and useful for the study of alga compositions.