摘要
吡咯里西啶生物碱(PAs)是一类植物内源产生的次生代谢产物,具有肝毒性、遗传毒性和致癌性等毒性作用。现有研究覆盖PAs种类有限,且由于同分异构体色谱分离难,大多研究采用加和定量策略。因此,本研究基于超高效液相色谱-三重四极杆质谱法(UHPLC-MS/MS)建立了干茶样本中欧盟法规限量的35种PAs的分析方法,其中有33种目标化合物获得了色谱分离,可以实现单独定量,有2种异构体共流出,进行加和定量。在电喷雾离子源(ESI)正离子模式下电离、多反应监测(MRM)模式下扫描,35种PAs在各自范围内呈现良好的线性关系(r^(2)>0.99),方法的检出限(LOD)和定量限(LOQ)分别为0.2~8.0μg/kg和0.5~25.0μg/kg。在1、2和5倍LOQ加标水平下,茶叶基质中均有89%以上的目标化合物平均回收率为70%~130%,3次平行实验相对标准偏差(RSD)小于20%(蓝蓟定和毛果天芥菜碱RSD小于30%)。应用该方法对21份黑茶和30份红茶样本进行检测,黑茶样本中35种PAs检出率为19.05%,总含量为5.07~15.48μg/kg,红茶样本中无PAs检出。所建立的分析方法可为茶叶样本中PAs的组成、含量及风险评估提供技术支撑。
Pyrrolizidine alkaloids(PAs)are natural toxins widely distributed in plants,which naturally occur in about 3% of the world’s flowering plants.To date,more than 660 PAs and their nitrogen oxides have been identified in over 6000 plants.Some PAs are hepatotoxic,genotoxic and tumorigenic,posing significant health risks to humans.These alkaloids are commonly detected as contaminants in various food products,including tea,grains,milk,honey,as well as plant-derived pharmaceuticals and dietary supplements.Currently,most studies on the quantitative methods for PAs focus on a limited number of PAs and employ an additive quantification strategy,largely due to the challenges associated with chromatographic separation of isomers.These approaches limit the ability to assess exposure and health risk accurately.Herein,a method was established to quantify 35 PAs individually in dried tea samples using ultra-high performance liquid chromatographytriple quadrupole mass spectrometry(UHPLC-MS/MS).The 35 target compounds were divided into two groups.The first group included 30 PAs,while the second group consisted of 5 PAs.These compounds were separated on Waters Acquity BEH C_(18)(150 mm×2.1 mm,1.7μm)and Thermo AcclaimTM C30(150 mm×2.1 mm,3.0μm)chromatographic columns,respectively.Mobile phases were H_(2)O containning 5 mmol/L ammonium formate and 0.13% formic acid(pH=3)and methanolacetonitrile(4∶6,volume raio)containing 0.1%formic acid.The 33 target compounds were separated and 2 isomers co-eluted.Under positive-ion electrospray ionization(ESI)and multiple reaction monitoring(MRM)mode,target compounds were quantified using the external standard method with the matrix-matched standard curve.The results demonstrated that all target compounds showed good linearity(r^(2)>0.99)in their respective mass concentration ranges.The limits of detection(LODs)and quantification(LOQs)of the method were in the range of 0.2-8.0μg/kg and 0.5-25.0μg/kg,respectively.The average recoveries of more than 89% compounds were in the range of 70%-130% at spiked levels of 1,2,and 5 fold-LOQs and the relative standard deviations(RSDs)were less than 20%(RSDs of lasiocarpine and echimidine were less than 30%).Moreover,the quantitative method of the 35 PAs was applied to 21 dark tea and 30 black tea samples from Yunnan and Fujian Province,where PAs were detected in 4 dark samples with total contents ranging from 5.07 to 15.48μg/kg and were not detected in black tea samples.The detection rate of PAs in dark tea samples was higher than that of black tea samples.That might be because most dark tea was processed from fresh leaves picked by machines,and this attribution could be associated with the mixing of weeds containing PAs during tea harvesting.And the detected concentrations of all samples were lower than the maximum levels of tea in European Union regulations,indicating that the tea samples involved in this research are basically safe.In brief,the quantitative method of the 35 PAs facilitates the analysis of the occurrence,composition and potential risks in tea samples.
作者
董璐
张晓林
杨晨
赵旌妍
王学梅
褚莹倩
李雪花
DONG Lu;ZHANG Xiaolin;YANG Chen;ZHAO Jingyan;WANG Xuemei;CHU Yingqian;LI Xuehua(Key Laboratory of Industrial Ecology and Environmental Engineering,School of Environment,Dalian University of Technology,Dalian 116024,China;Technology Center of Dalian Customs District,Dalian 116001,China)
出处
《色谱》
北大核心
2025年第12期1342-1353,共12页
Chinese Journal of Chromatography
基金
辽宁省应用基础研究项目(2023JH2/101700338)
海关总署科技计划项目(2024HK120)
大连海关科技计划项目(2023DK07).
关键词
吡咯里西啶生物碱
超高效液相色谱-三重四极杆质谱法
茶叶
pyrrolizidine alkaloids(PAs)
ultra-high performance liquid chromatography-triple quadrupole mass spectrometry(UHPLC-MS/MS)
tea
