The phytochemical content and biological activity of three Chlorella strains cultured in low (35 mg L −1 ) or high (700 mg L −1 ) nitrogen (N) and harvested on days 5 and 10 were evaluated. High N resulted in a higher biomass in Chlorella MACC 438 and MACC 452 while MACC 555 produced a higher biomass in low N. MACC 555 (low N/ day 5) had the highest phenolic content, and MACC 438 in low N/day 5 and high N/day 5 accumulated the highest flavonoids and condensed tannins, respectively. Iridoids were most abundant in MACC 452 on low N/day 10. Harvest time had the greatest effect on the phytochemical content with phenolics, flavonoids, and condensed tannins decreasing over time and iridoids increasing in low N and decreasing in high N conditions. Extracts were more active in β-carotene-linoleic acid model compared to 2,2-diphenyl-1-picrylhydrazyl (DPPH) free radical scavenging assay. Most extracts had good antimicrobial activity. Extracts became less potent over time in the antioxidant, acetylcholinesterase inhibitory (AChE), and antimicrobial assays when growing in low N and more potent in the antioxidant and AChE assays when grown in high N. Thus, phytochemical content and biological activities of the three Chlorella strains were affected by N levels, harvest time, and strain.