Abstract: Background and Objective: The growth rate, primary and secondary metabolites are influenced by nitrogen concentration of media. This study aimed to determine the effect of different concentrations of KNO3 as a source of nitrogen on the growth, phytochemical components, cholesterol reduction, antioxidant and antibacterial activities of the green alga, Scenedesmus obliquus (S. obliquus). Materials and Methods: Scenedesmus obliquus was cultured in Kuhl’s medium and KNO3 was added with different concentrations in the medium at different concentration (0.12, 0.75, 1.5, 2.25 and 3 g L–1). The effect of different five concentrations of nitrogen on protein and carbohydrates was determined. Antioxidant activity, total phenolic content (TPC) of alga extracts, cholesterol reduction and antibacterial activity were evaluated. The data analyses were carried out using SPSS software version 16. Results: The results revealed that the best KNO3 concentration for algal growth and carbohydrate content is1.5 g L–1. Meanwhile, the high lipid content was obtained with KNO3 starvation. Medium containing 0.75 g L–1 of KNO3 has the highest effect on the protein production of S. obliquus. The contents of dry alga extracted with chloroform: methanol and the extracted contents were determined by GC/MS chromatogram; the major contents were 5-Hydroxymethylfurfural followed by hexadecanoic acid (palmitic acid), cis-9-octadecenoic acid (oleic acid) and hexadecanoic acid and methyl ester, respectively. Conclusion: Potassium nitrate limitation caused the highest effect on the total phenol content (TPC), increased antioxidant capacity, cholesterol reduction activity and also antibacterial activity against Staphylococcus aureus and Escherichia coli.