基于超高效液相色谱-串联质谱法和多肽组学分析的鸡内金真伪鉴别策略

Authenticity discrimination for Galli Gigerii Endothelium Corneum based on UPLC-MS/MS and peptidomics

基于多肽组学技术系统分析鸡内金及其混伪品的特征肽并建立检测方法,用于鉴别鸡内金与其混伪品鸭内金、鹅内金、鸽内金。采用超高效液相色谱-四极杆-静电场轨道阱质谱(UPLC-Q-Exactive Obitriap-MS)技术结合多元统计分析对鸡内金及其混伪品多肽成分进行初步分析,通过pNovo软件结合人工确认图谱的方式鉴定多肽成分的结构,并合成鸡内金特征肽对照品进行验证;利用LC-MS/MS,针对鸡内金特征肽LESY对提取方式、提取时间、提取溶剂和溶剂体积等样品前处理方法进行了优化,以m/z 511.24→269.11和511.24→243.13作为检测离子,采用ESI~+模式进行了多反应监测(MRM)定性定量分析,测定得16批鸡内金中鸡内金特征肽LESY的质量分数为55.03~113.36μg·g^(-1);此外,针对常见混伪品多肽RDPVLVSR,以m/z 471.28→785.45和471.28→670.41作为检测离子,建立了鸡内金常见混伪品的LC-MS/MS定性检测方法。该研究建立的鸡内金及其混伪品特征肽检测方法操作简便,可快速、准确区分鸡内金及其混伪品,具有良好的专属性、稳定性及耐用性,可为鸡内金及其他动物类中药的鉴别及质量控制提供参考。

Peptidomics was employed to systematically analyze the characteristic peptides in Galli Gigerii Endothelium Corneum and its adulterants and establish a method for distinguishing Galli Gigerii Endothelium Corneum from its adulterants,including the gizzard membranes from ducks,geese,and pigeons.UPLC-Q-Exactive Orbitrap-MS was combined with multivariate statistical analysis to analyze the peptides in Galli Gigerii Endothelium Corneum and its adulterants.The structures of peptides were identified by pNovo combined with manual recognition of spectra,and synthetic peptide standards were used for validation.LC-MS/MS was used to optimize the sample pre-processing conditions,including the extraction procedure,extraction time,extraction solvents,and solvent volumes,for the characteristic peptide LESY in Galli Gigerii Endothelium Corneum.Multiple reaction monitoring(MRM)in the ESI+mode with m/z 511.24→269.11 and 511.24→243.13 as detection ions was employed for qualitative and quantitative analyses.The established UPLC-MS/MS method demonstrated good specificity,stability,and durability.The content of LESY in 16 batches of Galli Gigerii Endothelium Corneum samples ranged from 55.03 to 113.36μg·g^(-1).Additionally,a qualitative detection method for the common peptide RDPVLVSR in adulterants was established with m/z 471.28→785.45 and 471.28→670.41 as the detection ions.This study established a convenient,rapid,and accurate detection method for the characteristic peptides in Galli Gigerii Endothelium Corneum and its adulterants.The method possesses good specificity,stability,and durability,providing a valuable reference for the identification and quality control of Galli Gigerii Endothelium Corneum and other traditional Chinese medicines derived from animal sources.