This study investigated the screening, production and optimization of extracellular lipase from a fungus isolated from different samples in Egypt. The isolated fungi were screened on tributyrin agar for exogenous lipolytic activity. A promising lipase producing isolate of SH5Penicillium citrinum was selected and identified by the isolate SH5 of Penicillium citrinum that was identified as the highest lipase-producing strain. Molecular identification was carried out by amplifications of ITS-1, 5.8S and ITS-2 regions. The effects of incubation time, inducers, pH, temperature, carbon and nitrogen sources were varied for optimal lipase production using one factor at a time approach. Maximum lipase production was observed on the third day, pH 7.0 and at 30^oC. Olive oil, maltose and ammonium sulphate were the best inducers, carbon and nitrogen sources were respectively for lipase production.