{"title":"利用 Z-HILIC 耦合同位素稀释电喷雾离子化三重四极杆液相色谱-质谱法测定 27 种牛血浆氨基酸和代谢物以及去蛋白时间的影响","authors":"A.F. Ortega, M.E. Van Amburgh","doi":"10.3168/jdsc.2023-0449","DOIUrl":null,"url":null,"abstract":"<div><p>The use of zwitterionic-hydrophilic interaction liquid chromatography (Z-HILIC) columns for analysis of underivatized analytes has allowed simpler sample preparation of bovine plasma for sensitive and selective analysis, when coupled with mass spectrometry (MS). The objective of this study was to evaluate and validate this analytical technique to measure AA and metabolites in bovine plasma at 2 deproteinization times. A robust method using Z-HILIC coupled to a triple quadrupole MS (TQMS) was evaluated and validated to quantitatively analyze 19 AA using isotope dilution and 8 AA metabolites qualitatively in bovine deproteinized plasma. The timing of deproteinization was investigated to determine if plasma should be deproteinized upon collection (on-site) or immediately before analysis (in-lab). Analytes were separated using a Z-HILIC column in a 21 min run and analyzed with a TQMS in positive electrospray ionization for identification and quantification. The method was validated for standard curve linearity, limits of detection (LOD) and quantification (LOQ), intra- and interday precision (% coefficient of variation; CV), recovery (%), and freeze-thaw stability (% CV) after 1 mo. Coefficients of determination (R<sup>2</sup>) were over 0.993, and LOD and LOQ were below measured values for all AA. The CV for the intraday and interday precision were below 18%, except for cystine (Cys2) and Orn in-lab. Recoveries on-site and in-lab ranged from 75% to 120% for all analytes except Cys2 in-lab. Most analytes were stable after 1 mo of freezing regardless of deproteinization timing, CV <25%, except for hydroxyproline (Hyp). The concentration of Cys2 was affected by deproteinization in-lab compared with on-site, and even though Glu and Hyp were different between the 2 deproteinization timings, the concentrations between the 2 timings were within the standard deviation.</p></div>","PeriodicalId":94061,"journal":{"name":"JDS communications","volume":null,"pages":null},"PeriodicalIF":0.0000,"publicationDate":"2024-03-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.sciencedirect.com/science/article/pii/S2666910223001266/pdfft?md5=6b2ccc0cbeebfb5ebdec1b133f8fa305&pid=1-s2.0-S2666910223001266-main.pdf","citationCount":"0","resultStr":"{\"title\":\"Determination of 27 bovine plasma amino acids and metabolites using zwitterionic-hydrophilic interaction liquid chromatography coupled with isotope dilution electrospray ionization triple quadrupole liquid chromatography-mass spectrometry and the effect of deproteinization timing\",\"authors\":\"A.F. Ortega, M.E. Van Amburgh\",\"doi\":\"10.3168/jdsc.2023-0449\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"<div><p>The use of zwitterionic-hydrophilic interaction liquid chromatography (Z-HILIC) columns for analysis of underivatized analytes has allowed simpler sample preparation of bovine plasma for sensitive and selective analysis, when coupled with mass spectrometry (MS). The objective of this study was to evaluate and validate this analytical technique to measure AA and metabolites in bovine plasma at 2 deproteinization times. A robust method using Z-HILIC coupled to a triple quadrupole MS (TQMS) was evaluated and validated to quantitatively analyze 19 AA using isotope dilution and 8 AA metabolites qualitatively in bovine deproteinized plasma. The timing of deproteinization was investigated to determine if plasma should be deproteinized upon collection (on-site) or immediately before analysis (in-lab). Analytes were separated using a Z-HILIC column in a 21 min run and analyzed with a TQMS in positive electrospray ionization for identification and quantification. The method was validated for standard curve linearity, limits of detection (LOD) and quantification (LOQ), intra- and interday precision (% coefficient of variation; CV), recovery (%), and freeze-thaw stability (% CV) after 1 mo. Coefficients of determination (R<sup>2</sup>) were over 0.993, and LOD and LOQ were below measured values for all AA. The CV for the intraday and interday precision were below 18%, except for cystine (Cys2) and Orn in-lab. Recoveries on-site and in-lab ranged from 75% to 120% for all analytes except Cys2 in-lab. Most analytes were stable after 1 mo of freezing regardless of deproteinization timing, CV <25%, except for hydroxyproline (Hyp). The concentration of Cys2 was affected by deproteinization in-lab compared with on-site, and even though Glu and Hyp were different between the 2 deproteinization timings, the concentrations between the 2 timings were within the standard deviation.</p></div>\",\"PeriodicalId\":94061,\"journal\":{\"name\":\"JDS communications\",\"volume\":null,\"pages\":null},\"PeriodicalIF\":0.0000,\"publicationDate\":\"2024-03-01\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"https://www.sciencedirect.com/science/article/pii/S2666910223001266/pdfft?md5=6b2ccc0cbeebfb5ebdec1b133f8fa305&pid=1-s2.0-S2666910223001266-main.pdf\",\"citationCount\":\"0\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"JDS communications\",\"FirstCategoryId\":\"1085\",\"ListUrlMain\":\"https://www.sciencedirect.com/science/article/pii/S2666910223001266\",\"RegionNum\":0,\"RegionCategory\":null,\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"\",\"PubModel\":\"\",\"JCR\":\"\",\"JCRName\":\"\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"JDS communications","FirstCategoryId":"1085","ListUrlMain":"https://www.sciencedirect.com/science/article/pii/S2666910223001266","RegionNum":0,"RegionCategory":null,"ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"","JCRName":"","Score":null,"Total":0}
引用次数: 0
摘要
:使用齐聚物-亲水相互作用液相色谱柱(Z-HILIC)分析未充分活化的分析物,可以简化牛血浆的样品制备,与质谱(MS)联用时可进行灵敏、选择性分析。本研究的目的是评估和验证这种分析技术,以测定牛血浆中 AA 和代谢物在 2 个去蛋白时间内的含量。评估并验证了采用同位素稀释法定量分析牛脱蛋白血浆中 19 种 AA 和定性分析 8 种 AA 代谢物的 Z-HILIC 联用三重四极杆质谱(TQMS)稳健方法。对去蛋白的时间进行了研究,以确定血浆应在采集时(现场)还是在分析前(实验室内)立即去蛋白。使用 Z-HILIC 色谱柱在 21 分钟内分离分析物,然后使用 TQMS 正电离分析法进行鉴定和定量。该方法的标准曲线线性、检出限(LOD)和定量限(LOQ)、日内和日间精密度(%CV)、回收率(%)以及一个月后的冻融稳定性(%CV)均得到了验证。所有 AA 的测定系数(R 2 )均超过 0.993,检出限和定量限均低于测量值。
Determination of 27 bovine plasma amino acids and metabolites using zwitterionic-hydrophilic interaction liquid chromatography coupled with isotope dilution electrospray ionization triple quadrupole liquid chromatography-mass spectrometry and the effect of deproteinization timing
The use of zwitterionic-hydrophilic interaction liquid chromatography (Z-HILIC) columns for analysis of underivatized analytes has allowed simpler sample preparation of bovine plasma for sensitive and selective analysis, when coupled with mass spectrometry (MS). The objective of this study was to evaluate and validate this analytical technique to measure AA and metabolites in bovine plasma at 2 deproteinization times. A robust method using Z-HILIC coupled to a triple quadrupole MS (TQMS) was evaluated and validated to quantitatively analyze 19 AA using isotope dilution and 8 AA metabolites qualitatively in bovine deproteinized plasma. The timing of deproteinization was investigated to determine if plasma should be deproteinized upon collection (on-site) or immediately before analysis (in-lab). Analytes were separated using a Z-HILIC column in a 21 min run and analyzed with a TQMS in positive electrospray ionization for identification and quantification. The method was validated for standard curve linearity, limits of detection (LOD) and quantification (LOQ), intra- and interday precision (% coefficient of variation; CV), recovery (%), and freeze-thaw stability (% CV) after 1 mo. Coefficients of determination (R2) were over 0.993, and LOD and LOQ were below measured values for all AA. The CV for the intraday and interday precision were below 18%, except for cystine (Cys2) and Orn in-lab. Recoveries on-site and in-lab ranged from 75% to 120% for all analytes except Cys2 in-lab. Most analytes were stable after 1 mo of freezing regardless of deproteinization timing, CV <25%, except for hydroxyproline (Hyp). The concentration of Cys2 was affected by deproteinization in-lab compared with on-site, and even though Glu and Hyp were different between the 2 deproteinization timings, the concentrations between the 2 timings were within the standard deviation.