Objective To explore the effect of antiosteoporotic active fraction from Er-Xian Decoction (EXD) on retinoic acid-induced osteoporosis in rats. Methods Sixty female SD rats were randomly divided into normal control group, model group, estradiol valerate (0.2 mg/kg) group and antiosteoporotic active fraction from EXD low-, middle-and high-dose groups (50, 100, 200 mg/kg). Except for the rats of normal group, the others were intragastrically treated with 70 mg/(kg·d) of retinoic acid for 14 days to induce osteoporosis. Then the rats were treated with estradiol valeratse or antiosteoporotic active fraction from EXD (10 mL/kg) for 28 days. The biochemical parameters in serum and urine of rats were detected by ELISA. The bone mineral density and trabecular bone microstructure of distal femur were analyzed with micro-CT. Results The antiosteoporotic active fraction from EXD significantly decreased calcium content in urine and activities of alkaline phosphatase and tartratse-resistant acid phosphatase in the sera of retinoic acid-induced osteoporotic rats (P<0.05, P<0.01). High-dose of antiosteoporotic active fraction from EXD significantly increased bone mineral density of distal femur (P<0.05). And the antiosteoporotic active fraction from EXD improved the trabecular bone microstructure of distal femur in retinoic acid-induced osteoporotic rats. Conclusion Antiosteoporotic active fraction from EXD can reduce bone loss and deterioration of bone microstructure in retinoic acid-induced osteoporotic rats.