The acute and chronic toxicity of the β-carotenoid-rich fraction (BCRF) from Dunaliella salina (Chlorophyta) microalgae was evaluated by the supplementation of high doses of BCRF to experimental animals. The HPLC analysis showed that ‎hexane crude extract of D. salina contains 0.813%‎ of β-carotene. Various biomarkers were involved in this study, which includes hematological and biochemical analyses such as complete blood count, liver enzymes, and kidney function. In addition albumin and glucose levels were determined. The histopathological examination was also performed on hepatic, renal, and cardiac architectures to evaluate its safety. An acute toxicity study revealed no toxicological relevance post administration of serial doses (500 mg/kg body weight up to 5000 mg/kg b.wt.).The supplementation of experimental animals (male and female mice and rats) with a dose of 500 mg/kg body weight of βCRF of D. salina daily for three consecutive months did not show any toxicity symptoms in both genders of mice and rats and/or mortality. Histopathological observation exhibited no noticeable change in the liver, kidney and heart architectures of both male and female mice and rats. Therefore, BCRF of D. salina LD50 is up to 5000 mg/kg, and it is safe at the dose of 500 mg/kg b.wt. in chronic toxicity.