Abstract: Sedimentary Fatty Acids (FAs) were investigated in surficial sediment of four sub-branches (canals) of River Nile, Egypt, to distinguish their origin of organic matter. Seventeen stations were investigated for four seasons from spring, 2014 till winter, 2015. Analysis of variance (ANOVA) with post hoc suggested significant seasonal differences in total fatty acids, fatty acid classes and main fatty acids with obvious increase in spring. The dominant Saturated Fatty Acids (SFA) in sediment samples were C16:0 and C18:0. Dominant monounsaturated fatty acids (MUFA) included C16:1ω7 and C17:1ω7 whereas, C18:1ω9 was less important. Fatty acids indicators including microalgal classes, bacteria, sewage and terrestrial plants were used to distinguish their contribution to surficial sediment organic matter. The results indicated that microalgae were major contributor to sediment organic matter, alternatively, land plant organic matter was nearly absent from the sedimentary organic matter in the four canals. Between microalgae, diatoms were prevalence over cyanobacteria and green algae. The seasonal changes in fatty acids composition were driven by the changes in species composition of microalgal communities rather than the direct effect of environmental characteristics.