Objective To exploration the feasibility to detect the drug resistance of human colon cancer in mice with magnetic resonance spectrum in vivo. Methods Nude mice with human colon cancer SW480(drug response) and 5 nude mice with SW480/5-Fu(5- Fu resistance) were given 1 H-MRS examinations when the diameters of tumor were over 1.5 centimeters. Then these mouse models were sacrificed. Their tumors were removed and detected the protein expression of PKC, P-gp and MPR1 (Western blot), apoptosis (TUNNEL). The correlation between metabolites detected with 1 H-MRS and tumor drug resistance and the protein expression of PKC, P-gp and MPR1 (Western blot), tumor apoptosis were analyized. Results The the area of cho, lac and Lip of tumors in SW480 group were significantly higher than that in SW480/5-Fu group (P＜0.05). The protein expression of PKC, P-gp and MPR1 in SW480/5-Fu group were significantly higher than that of SW480 group (P＜0.05). In the TUNNEL section, apoptotic cell were observed in two groups, and there is no significant difference between two groups (P＞0.05). The area of cho, lac and Lip of tumors was positively correlated with drug resistance (r, P:0.870,0.001). The the area of cho has a positive correlation with the protein expression of PKC (r, P:0.745,0.013), P-gp (r, P:0.782,0.008). Conclusion The change of metabolites cho, lac and Lip detected with 1 H - MRS has the potential to help detection of colon cancer drug resistance with MR in vivo.