Additivity of enzyme hydrolysate gross energy (EHGE) of feedstuffs determined using computer-controlled simulated digestion system was investigated to validate the method for in vitro digestion. Twelve treatments consisted of 4 feedstuffs and 8 diets were used in a single factorial completely randomized arrangement. Treatments 1 to 4 were corn, soybean meal, cottonseed meal and wheat bran diets, respectively. Treatments 5 to 12 were 8 diets formulated with 2, 3 or 4 feedstuffs at different concentration. Each treatment contained 5 replicates with 1 digestion tube per replicate. The dry matter digestibility (DMD) and EHGE of each treatment were determined using computer-controlled simulated digestion system. The calculated DMD and EHGE of treatments 5 to 12 were estimated according to the determined DMD and EHGE of individual ingredients and their concentrations in the diets, respectively. The results showed as follows: the mean of determined DMD was greater than that of calculated values of 8 diets (70.15% and 69.48% for determined and calculated values, respectively; P=0.000 9). However, the regression of determined values on calculated values for DMD revealed that the coefficient of determination was 0.993 3 (P<0.05), and the intercept did not differ from 0 (intercept=1.77, P=0.468 3) and slope did not differ from 1 (slope=0.98, P=0.647 4). The variance and coefficient of variation (CV) of determined values were less than those of calculated values for DMD (P=0.000 2 and P=0.001 0 for variance and CV, respectively). The regression of observed values on calculated values for variance and CV of DMD showed that the coefficients of determination were 0.020 8 (P>0.05) and 0.115 7 (P>0.05), respectively. No significant difference was observed between the determined values and calculated values for EHGE (13.21 and 13.21 MJ/kg for determined and calculated values, respectively; P=0.965 3). The regression of determined values on calculated values for EHGE revealed that the coefficient of determination was 0.991 0 (P<0.05), and the intercept did not differ from 0 (intercept=0.04, P=0.941 9) and slope did not differ from 1 (slope=1.00, P=0.940 0). The differences between observed values and calculated values were not significant for variance (P=0.681 1), however, significant for CV (P=0.003 6). The regression of observed values on calculated values for variance and CV of DMD showed that the coefficients of determination were 0.020 0 (P>0.05) and 0.080 5 (P>0.05), respectively. In conclusion, the values in corn, soybean meal, cottonseed meal and wheat bran are additive for DMD and EHGE in a complete diet, respectively. However, the values for variance and CV are not additive, respectively.