© 2018 2018 Walter de Gruyter GmbH, Berlin/Boston.The aim of the study was to evaluate the dynamic thiol/disulfide homeostasis (TDH) with a new method in patients with pubertal gynecomastia and also to investigate the relationship between sex hormones. Thiol/disulfide homeostasis, involving native thiol (SH), disulfide (SS) and total thiol (SS + SH), was evaluated between 20 adolescent boys with gynecomastia in mid-puberty and 50 healthy adolescents, who were matched for age, body mass index (BMI) and pubertal stage. The correlations of total serum testosterone (TT) and estradiol (E2) levels with the oxidative parameters were also determined. No significant difference was found between the total thiol, native thiol, disulfide/native thiol and disulfide/total thiol ratios of the patient and control groups. The mean disulfide concentrations, disulfide/native thiol and the disulfide/total thiol ratios were statistically significantly higher in the patient group than in the controls. A positive correlation was found between the E2 and native thiol levels, also there was a negative correlation between the E2 and disulfide levels. TT was negatively correlated with both native thiol and disulfide levels. All these did not statistically differ between the patients with unilateral and bilateral gynecomastia. There was an impaired thiol/disulfide homeostasis in patients with pubertal gynecomastia. According to this result, we can postulate that oxidative stress may be an etiologic factor that contributes to initiation and/or progression of gynecomastia.