Ketoprofen is one of the most prescribed medications in medical practice and belongs to the class of non-steroidal anti-inflammatory drugs. Due to the therapeutic significant differences of its enantiomers, several studies have been developed with the aim of producing pure enantiomers by biocatalytic process. Therefore, enantioselective properties of lipases can be explored. In this study, the potential application of Lipase of Thermomyces lanuginosus (LTL) immobilized on silica by different methodologies was evaluated for the chiral resolution of ketoprofen. The best esterification condition of R, S-ketoprofen offered 50% conversion to the ketoprofen ethyl ester.