A gas chromatographic-mass spectrometric method for the quantitative determination of S-nitrosoalbumin (SNALB) in human plasma is described. The method is based on selective extraction of SNALB and its 1 5 N-labeled SNALB analog (S 1 5 NALB) used as internal standard on HiTrapBlue Sepharose affinity columns, Hg 2 + -catalysed conversion of the S-nitroso groups to nitrite and [ 1 5 N]nitrite, respectively, followed by their derivatization to the pentafluorobenzyl derivatives and quantification by GC-MS. Mean recovery of SNALB and S 1 5 NALB from plasma was 45%. Mean precision and accuracy within the range 0-10 μM was 95% and 99%, respectively. The limit of quantitation was determined as 100 nM at a precision of 93.8% and an accuracy of 94.8%. Considerable improvement of method sensitivity is possible by eliminating nitrite present in the elution buffer. The limit of detection was 0.2 nM corresponding to 67 amol of S 1 5 NALB. In 0.4-ml aliquots of plasma samples from healthy humans, endogenous SNALB was determined at concentrations of 181+/-150 nM (mean+/-SD, n=23). External addition of SNALB to these plasma samples at 2 μM and 5 μM serving as quality control samples resulted in quantitative recovery of SNALB. Our results show that SNALB occurs in human plasma at concentrations at least one-order of magnitude smaller than those reported in the literature from measurements using chemiluminescence.