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The aim of the current study was to compare the effect of different physical processing methods including roasting at 140 °C for 15 (R15) or 30 minutes (R30), gamma ray irradiation (γ-irradiation) at doses of 15 (γ15), 30 (γ30) and 45 (γ45) kGy, and microwaving at 800 W for 2 (MW2), 4 (MW4) and 6 minutes (MW6) of whole cottonseed (WCS) on ruminal degradation. In vitro crude protein (CP) digestibility and gossypol contents were compared as well. In situ experiment was conducted on three permanent rumen-fistulated bulls. Gossypol content was decreased among treatments (p<0.05). The lowest degradation rate of protein in rumen was obtained for γ45 treatment. The rate of degradation of the potentially degradation fraction was decreased for both dry matter (DM) (p=0.002) and CP (p=0.006) with different treatments. The lowest values for effective degradation in all passage rates were obtained with γ45. Both microwaving and γ irradiation showed difference for CP effective degradability parameter. The greatest value of in vitro CP digestibility (p<0.05) was observed for a dose of 45 kGy gamma-irradiated cottonseed compared to untreated WCS. Based on the results, γ ray irradiation with 45 kGy was the most effective processing method in both reducing the gossypol content and escaping the protein through rumen for WCS in ruminant nutrition. [ABSTRACT FROM AUTHOR]