The objective of the present study was to evaluate potential adverse effects of diacylglycerol microemulsion (DAGM) in rats/mice. Acute safety evaluation was carried out by giving intragastrically with 20 mL 25% DAG kg-1 body weight of DAGM or water with two groups of mice. Chronic safety evaluation with 40 male and 40 female Sprague-Dawley rats was carried out by setting a control group and 3 different dose groups (n=10 male+10 female) administered with DAGM with 6.7, 10 and 20 mL kg-1 body weight per day. Relevant parameters of liver and kidney function and biochemistry were determined by standard methods at end point. Acute toxicity study revealed the maximum tolerated dose (MTD) of DAGM was 20 mL kg-1 body weight in mice. No death was observed at the dose of 20 mL kg-1 body weight per day. Chronic safety evaluation did not show significant changes on hematological, histopathologic, clinical, and biochemical tests when administered at levels of 6.7, 10 and 20 mL kg-1 body weight per day to rats for 30 d. No significant body weight changes were observed in different dosages in both male and female rats. Our results suggested that acute and chronic adverse effects were not observed in histology and clinical parameters in both mice and rats. The results suggested that DAGM is safe at the experimental levels.