Simultaneous determination of denatured proteins by hydrophobic interaction chromatography

The quantitation of denatured proteins by hydrophobic interaction chromatog raphy (HIC) with a mobile phase containing 8.0 mol l(-1) urea is proposed. The following couples of enzymes, taken as model molecules, have been exami ned: glyceraldehyde 3-phosphate dehydrogenase and aldolase from rabbit skel etal muscle, alcohol dehydrogenase and phosphoglucose isomerase from baker' s yeast. For each denatured protein, reproducibility, linearity range, reco very (> 97%) and determination limit are reported. The feasibility of the s imultaneous determination of such protein couples in synthetic mixtures has been tested, and HIC proved a useful tool for separation of denatured prot eins, in particular those having similar relative molecular mass and/ or ch arge.