Four methods were adopted, including the mobile nylon bag (MNB) method, modified three-step in vitro (MTS) method, original three-step in vitro (OTS) method, and acid detergent insoluble nitrogen (ADIN) estimating method, to evaluate the intestinal digestibility of rumen undegradable protein (DRUP) of 10 types of concentrates and 7 types of roughages. After correlation analysis to determine the DRUP values using the MNB, MTS, OTS, and ADIN methods, the study aimed to find out appropriate methods to replace the MNB method due to its disadvantages such as high price, long time period, and use of a duodenal T-fistula. Three dairy cows with a permanent ruminal fistula and duodenal T-fistula were used in a single-factor experimental design. The results showed that the determined DRUP values using the MNB method for soybean meal, cottonseed meal, rapeseed meal, sunflower meal, corn germ meal, corn, rice bran, barley, wheat bran, corn fiber feed, Alfalfa (Zhao dong), Alfalfa (Long mu 801), Alfalfa (Long mu 803), grass (North), Grass (Inner Mongolia), corn silage and corn straw were 98.13%, 87.37%, 88.47%, 82.60%, 75.40%, 93.23%, 69.27%, 91.27%, 72.37%, 79.03%, 66.72%, 68.64%, 73.57%, 50.47%, 51.52%, 54.05%, and 43.84%, respectively. The coefficient of determination (R2 = 0.964) of the results between the MTS method and the MNB method was higher than that (R2 = 0.942) between the OTS method and the MNB method. The coefficient of determination of the DRUP values of the concentrates among the in vitro method (including the MTS and OTS methods) and the MNB method was higher than that of the roughage. There was a weak correlation between the determined DRUP values in concentrates obtained from the ADIN method and those from the MNB method, and there was a significant correlation (p<0.01) between the determined DRUP values of the roughage obtained from the MNB method and those obtained from ADIN method. The DRUP values were significantly correlated with the nutritional ingredients of the feeds. The regression equation was DRUP =100.5566+0.4169CP − 0.4344SP − 0.7102NDF − 0.7950EE (R2 = 0.8668, p<0.01; CP, crude protein; SP, soluble protein; NDF, neutral detergent fiber; EE, ether extract). It was concluded that both the MTS method and the OTS may suitable to replace the MNB method for determining the DRUP values and the former method was more effective. Only the ADIN method could be used to predict the values of the roughages but conventional nutritional ingredients were available for all of the samples’ DRUP.