Technical Abstract: A number of clinical and biochemical studies demonstrate that obesity and insulin resistance are associated with increases in oxidative stress and inflammation. Paradoxically, insulin sensitivity can be enhanced by oxidative inactivation of cysteine residues of phosphatases, and inflammation can be reduced by S-glutathionylation with formation of protein-glutathione mixed disulfides (PSSG). Although oxidation of protein-bound thiols (PSH) regulates protein activity and is increased in multiple diseases, it is not known whether there are changes in PSH oxidation species in obesity. In this work, we tested the hypothesis that obesity is associated with decreased levels of proteins containing oxidized protein thiols. We examined tissue levels of protein sulfenic acids (PSOH) and PSSG in liver, visceral adipose tissue, and skeletal muscle derived from glucose intolerant, obese-prone Sprague-Dawley rats. Our data indicate that decreases in PSSG content occurred in liver (44%) and adipose (26%) but not skeletal muscle in obese rats fed a 45% fat-calorie diet versus lean rats fed a 10% fat-calorie diet. PSOH content did not change in the tissue between the two groups. The activity of the enzyme glutaredoxin (GLRX) responsible for reversal of PSSG formation did not change in muscle and liver between the two groups. However, levels of GLRX1 were elevated 70% in the adipose tissue of the obese, 45% fat calorie-fed rats. These are the first data to link changes in S-glutathionylation and GLRX1 to adipose tissue in the obese and demonstrate that redox changes in thiol status occur in adipose tissue as a result of obesity.