Esterases are known to play essential role inmetabolism, reproductive physiology and behaviour of Drosophila. Esterases are highly polymorphic enzymes in Drosophila, but the polymorphism of these enzymes is not well studied in Drosophila ananassae. Recent studies on esterase polymorphism in D. ananassae revealed that Est-4 locus comprises Est-4 active and Est-4 null alleles depending on enzymatic activity. For the in vivo functional characterization of this locus, homozygous lines of genotypes Est-4 active and Est-4 null were derived from the flies collected from Gangtok, Sikkim, in 2006. Mating propensity, mating pattern, fecundity, fertility and productivity of female, life span and triglycerides level were investigated in the flies bearing either Est-4 active or Est-4 null genotypes. Results showed that mating occurred randomly with nonsignificant difference in mating propensity between Est-4 active and Est-4 null flies. However, a significant difference in fecundity and strong dependency between genotypes and the rate of fertility was found. The median values of progeny produced per female were 24 and 20 for Est-4 active and Est-4 null genotypes, respectively. The life span assay showed a significant difference in the survivorship between the two genotypes. Triglycerides level was higher in Esterase-4 active larval haemolymph as well as in mature flies' homogenate than that of Esterase-4 null. Thus, Esterase-4 locus of D. ananassae has its role in fecundity, fertility and productivity of female, life span control and lipid metabolism.