The goal of the current work was to assess biosurfactant (BS) produced by marine fungal isolated from the contaminated site of El-max Bay, Egypt, for bioremediation of oil contaminated sea water. In this regard, the four biosurfactants assessment tests oil displacement (OD), emulsification index (E24), hemolysis tests, and surface tension (ST) were used to screen selected marine fungal isolates. By measuring 1.5cm oil displacement, E24 57%, hemolytic activity, and the highest surface tension 37.1mN/ m, Plectosphaerella cucumerina strain HBKB was considered as the most potent isolate for producing a highly active biosurfactant. To assess the factors influencing biosurfactant production from minimal salt medium (MSM), Plackett-Burman design was used to get the maximum biosurfactants yield. Out of the seven variables that were examined, three (NH4NO3, KH2PO4 and yeast extract) were conducted for response surface methodology (RSM). These key effective elements were tested as independent variables and the surface tension (Mn/ m2) as a dependent variable. It was observed that the correlation coefficient (R2) showed a good fitness with the ‘‘Adjustive R-Squared" (with deference less than 0.1), thus the experimental and predicted values of the model agreed. The 3D curves of the quadratic model indicated the interactions between both NH4NO3 (4g/ L) and yeast extract (0.8g/ L) concentrations increased the surface tension level by 32mN/ m2. GC- MS was used to identify ethyl acetate biosurfactant extract. The largest greatest peak indicated the presence of the ionic surfactant fatty acid 1-Docosene (1-Nonadecene). This biosurfactant enhanced the bioremediation technique that is used to treat contaminated water containing petroleum oil residues. Moreover, the unsterilized- unamended supplied by 1-docosene extracted biosurfactant showed a higher oil remediation represented by E24 (55%), OD (2cm), and ST (34.5mN/ m2). Therefore, the breakdown, deterioration, and remediation of contaminated oil from aquatic habitats could be accomplished by P. cucumerina strain HBKB that produce the docosene surfactant.