Reliable and generic liquid chromatography/mass spectrometry quantification of short peptides using a stable‐isotope‐labeled labeling agent

2020 ◽  
Vol 34 (24) ◽  
Author(s):  
Anne J. Kleinnijenhuis ◽  
Bjorn Vergauwen ◽  
Frédérique L. Holthoon ◽  
Maarten Hekman
Keyword(s):  
Mass Spectrometry ◽  
Liquid Chromatography ◽  
Stable Isotope ◽  
Short Peptides ◽  
Liquid Chromatography Mass Spectrometry ◽  
Chromatography Mass Spectrometry

scholarly journals Quantification of Serum and Urinary S-Adenosylmethionine and S-Adenosylhomocysteine by Stable-Isotope-Dilution Liquid Chromatography-Mass Spectrometry

2004 ◽  
Vol 50 (2) ◽  
pp. 365-372 ◽  
Author(s):  
Sally P Stabler ◽  
Robert H Allen
Keyword(s):  
Mass Spectrometry ◽  
Liquid Chromatography ◽  
Stable Isotope ◽  
Isotope Dilution ◽  
Biological Fluids ◽  
Internal Standard ◽  
Fractional Excretion ◽  
Liquid Chromatography Mass Spectrometry ◽  
Stable Isotope Dilution ◽  
Chromatography Mass Spectrometry

Abstract Background: We have developed an assay that uses stable-isotope-dilution liquid chromatography–mass spectrometry to assess S-adenosylmethionine (SAM) and S-adenosylhomocysteine (SAH) in body fluids to investigate the relationship of these metabolites to hyperhomocysteinemia. Methods: Commercially obtained SAM (D3 methyl) and 13C5-SAH uniformly labeled in the adenosyl moiety, which was synthesized using S-adenosylhomocysteine hydrolase, were added to samples followed by perchloric acid protein precipitation, C18 chromatography, and analysis by liquid chromatography–mass spectrometry with quantification by comparison of the areas of internal standard and endogenous peaks. Results: Estimates of intraassay imprecision (CV) were 5% and 17% for SAM and SAH, respectively (n = 10). SAM decreased and SAH increased in serum and plasma samples at both 4 °C and room temperature over 80 h. SAM and SAH were unstable in samples stored longer than 2 years at −20 °C. In 48 volunteers, the estimated reference intervals [from mean (2 SD) of log-transformed data] for serum SAM and SAH were 71–168 and 8–26 nmol/L, respectively. Fractional excretion of SAM was higher than that of SAH, and the urinary SAM:SAH ratio was much higher than the serum or erythrocyte SAM:SAH ratios. Conclusions: Stable-isotope-dilution liquid chromatography–mass spectrometry can be used to quantify SAM and SAH in biological fluids and tissues. Sample handling and storage must be stringently controlled for any epidemiologic or clinical use of such assays.

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