Background: Dietary supplements containing conjugated linoleic acid (CLA) are widely promoted for weight loss management over the counter. Recently, FDA approved the CLA as Generally Recognized as Safe category so that it can be used in various food and beverages. The combined effect of CLA isomers have been studied extensively in animals and humans, however, the role of individual isomers remains unraveled. Aim: The present investigation addresses the effects of CLA isomers on body composition and body weight as well as safety using female C57Bl/6J aging mice. Methods: Two main CLA isomers and their mixture were fed to 12-months-old female C57Bl/6J mice. Ten percent corn oil (CO) based fat diet supplemented with 0.5% purified cis 9 trans 11 (c9,t11) CLA or trans 10 cis 12 (t10,c12) CLA or their mixture (CLA mix, 50:50) for 6 months. The lean mass, fat mass, glucose, non-esterified fatty acids, and insulin were examined at the end of study. Results: As a result of 6 months dietary intervention, both t10,c12 CLA and CLA mix groups showed increased lean mass and reduced fat mass compared to that of c9,t11 CLA and CO group. However, insulin resistance and liver hypertrophy were observed in t10,c12 CLA and CLA mix groups based on the results of homeostasis model assessment, revised quantitative insulin-sensitivity check index (R-QUICKI), intravenous glucose tolerance test, and liver histology. Liver histology revealed that increased liver weight was due to hypertrophy. Conclusion: In conclusion, the major CLA isomers have a distinct effect on fat mass, glucose, and insulin metabolism. The t10,c12 isomer was found to reduce the fat mass and to increase the lean mass but significantly contributed to increase insulin resistance and liver hypertrophy, whereas c9,t11 isomer prevented the insulin resistance. Between the two major CLA isomers, the t10,c12 was attributed to reduce fat mass whereas, c9,t11 improves the insulin sensitivity. © 2009 Springer-Verlag.