Of the total carotenoids in respective algal samples, beta-carotene in Spirulina platensis was 69.5%, astaxanthin and its esters in Haematococcus pluvialis was 81.38%, and lutein in Botryococcus braunii was 74.6%. The carotenoids were characterized by mass spectrometry. A time-course study of carotenoids in rats after administration of microalgal biomass showed peak levels in plasma, liver, and eyes at 2, 4, and 6 h, respectively. Beta-carotene accumulation in Spirulina-fed rats was maximum in eye tissues at 6 h. Similarly, levels of astaxanthin and lutein in Haematococcus- and Botryococcus-fed rats were also maximal in eye tissues. Astaxanthin from H. pluvialis showed better bioavailability than beta-carotene and lutein. The antioxidant enzymes, catalase, superoxide dismutase, peroxidase, and TBARS were significantly high in plasma at 2 h and in liver at 4 h, evidently offering protection from free radicals. This study implies that microalgae can be a good source of carotenoids of high bioavailability and nutraceutical value.