The incidence of colorectal cancer in Indonesia reaches 12.8 per 100,000 adult populations with mortality rate reaching 9.5% of all cancer cases. Current chemotherapy widely used has not been satisfactory, quite expensive, and may produce untoward side effect. Therefore, it is necessary to develop an effective and safety alternative agent from herbs to eradicate colorectal cancer. Previous studies has shown that α-mangostin contained in mangosteen peel extracts and fraction (Garcinia mangostana) has high antioxidant content and activities as anti-inflammatory, antibacterial, and antivirus. To get a higher level of α-mangostin content, the fraction used in this study is acquired by ethyl acetate fraction, and so the fraction yields 25% α-mangostin. The preliminary study done by our research team showed that the fraction has high cytotoxicity based on brine shrimp lethality test and high antioxidant activity. These results indicate the potential of ethyl acetate mangosteen fraction as an anticancer agent. In this study, we investigate acute toxicity of ethyl acetate mangosteen fraction on liver and kidney function of mice. Twenty of female BALB/c mice are divided into 4 groups. Three groups are given intra gastric administration of a single dose of 2, 4, and 6g/kg body weight (BW) of ethyl acetate mangosteen fraction and another group as a control. The findings show that the functional marker of kidney - blood urea nitrogen (BUN) and creatinine serum, and liver - serum glutamic oxaloacetic transaminase (SGOT) and serum glutamic pyruvate transaminase (SGPT), do not differ from those of the control at a single dose of 2g/kg BW of ethyl acetate fraction. However, an increase of serum creatinine occurs at the administration of dose 6g/kg BW compared with the control. The results of this study indicate that a single dose of 2 and 4g/kg BW of ethyl acetate fraction of Garcinia mangostana do not cause toxicity on liver and kidney function of mice. To ensure the safety level of the fraction, further histopathological examination is needed.