Abstract The glycation of γ‐globulin is catalyzed by phosphate buffer, whereas that of human serum albumin (HSA) and ovalbumin is not. The observed rate constant of spontaneous glycation of HSA is twofold larger than ovalbumin and γ‐globulin. When D ‐glucose‐2‐ h is compared with D ‐glucose‐2‐ d , the overall kinetic isotope effect for the buffer‐independent rates is H k 0 / D k 0 = 4.43 ± 0.06 for HSA. The substrate isotope effect for the buffer‐independent term excludes proton abstraction as the rate‐determining step in the Amadori rearrangement by phosphate buffer. Catalysis by phosphate buffer of the glycation of γ‐globulin indicates that phosphate is the abstracting base in the Amadori rearrangement. These results suggest that the phosphate buffer plays a fundamental role in the glycation of γ‐globulin. Copyright © 2004 John Wiley & Sons, Ltd.