A method for quantifying the level of glutathione (GSH) in yeast cell lysate is described using 1H NMR analysis. For quantification purposes, the 1H resonances corresponding to the Cys βCH2 of GSH were identified as having the fewest overlapping spectral interferences from lysate matrix components using GSH spiked yeast lysate samples. Two methods, standard addition based on peak integration and a spectral subtraction approach, were evaluated for quantifying GSH in lysate samples. The peak integration procedure required baseline estimation and a peak fitting step to correct for background interferences while the spectral subtraction procedure was comparatively straightforward. The level of GSH measured by 1H NMR was in good agreement with the concentration measured by the DTNB-GSSG reductase recycling assay. The proposed NMR method can lead to a reliable quantitation of GSH and may be applicable to a variety of other analytes of interest in complex biological matrices.
Pub Type: Journals
Glutathione, 1H NMR, Standard addition, Spectral subtraction