Abstract: Objective:To study the optimal extraction process of red ginseng and Rhodiola crenulata(RR)and observe its anti-fatigue effect. Methods:Red ginseng and Rhodiola crenulata were extracted by heating and reflux extraction method. The effects of ethanol concentration,solid-liquid ratio,extraction time and extraction times on the extraction process were observed by orthogonal experiment,and the total ginseng in the extract was observed. The contents of saponin and salidroside was analyzed,from which the optimal extraction process was preferred. Male Kunming mice were randomly divided into the blank control group and the high,medium and low dose groups. The exhaustive swimming experiment was performed to record the exhaustive swimming time of the mice,and the serum urea nitrogen,blood lactate and hepatic glycogen levels were detected. Results:The optimal extraction process of RR was 80% ethanol,the ratio of material to liquid was 1:10 g/mL,the extraction time was 120 min,and the extraction times were 3 times. Taking the high-dose group as an example,compared with the blank control group,RR could significantly extend the exhaustive swimming time of mice by 3.17 times. And the serum urea nitrogen and lactate levels in mice decreased by 39.08% and 31.20%,respectively,and the liver glycogen was significantly increased. The level was 40.90%(p<0.01). Conclusion:The optimal extraction process of RR was obtained,and its anti-fatigue effect was determined,which could provide a basis for the development of health food with anti-fatigue function.