Validated Quantitation of Underivatized Amino Acids in Human Blood Samples by Volatile Ion-Pair Reversed-Phase Liquid Chromatography Coupled to Isotope Dilution Tandem Mass Spectrometry
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- 作者：Jun Qu ; Yiming Wang ; Guoan Luo ; Zhuping Wu ; and Chengdui Yang
- 刊名：Analytical Chemistry
- 出版年：2002
- 出版时间：May 1, 2002
- 年：2002
- 卷：74
- 期：9
- 页码：2034 - 2040
- 全文大小：63K
- 年卷期：v.74,no.9(May 1, 2002)
- ISSN：1520-6882
文摘
Quantitation of amino acids in complex matrixes withoutderivatization is advantageous; however, difficulties existin both the separation and the detection of those compounds. A validated method that is based on the use ofvolatile ion-pair liquid chromatography coupled to stableisotope dilution tandem mass spectrometry has beendeveloped for the simple and accurate quantitation ofunderivatized amino acids in biological samples. Sufficientseparation of 22 underivatized amino acids was achievedon a C18 column in 36 min using perfluoroheptanoic acid(PFHA) and trifluoroacetic acid (TFA) as mobile phasemodifiers. The collisionally activated dissociation spectraof the amino acids were investigated and the transitionsof [M + H]+ 
[M + H - 46]+, which are specific to
-amino acids, were used for the detection of most aminoacids and their stable isotopes. The calibration curveswere linear over the range of 0.10-100 
g/mL, and thedetection limits were 0.03-20 pmol on column. Thequantitative results by this method were compared withthose by an established OPA-derivatization HPLC methodin the assay of 8 human serum samples, and betterrecovery and precision data of this method were observed.The method was also applied to the neonatal screeningfor phenylketonuria (PKU) with dry blood spots, and theresults were satisfactory. This is the first time that allproteinogenic amino acids have been quantified directlyfrom biological extracts without any kind of derivatization.The technique shows potential for routine determinationof amino acids and analogous compounds in complexmatrixes.
[M + H - 46]+, which are specific to-amino acids, were used for the detection of most aminoacids and their stable isotopes. The calibration curveswere linear over the range of 0.10-100 g/mL, and thedetection limits were 0.03-20 pmol on column. Thequantitative results by this method were compared withthose by an established OPA-derivatization HPLC methodin the assay of 8 human serum samples, and betterrecovery and precision data of this method were observed.The method was also applied to the neonatal screeningfor phenylketonuria (PKU) with dry blood spots, and theresults were satisfactory. This is the first time that allproteinogenic amino acids have been quantified directlyfrom biological extracts without any kind of derivatization.The technique shows potential for routine determinationof amino acids and analogous compounds in complexmatrixes.