Abstract: ObjectiveThe technical conditions of extracting nucleosides from Paris polyphylla var. yunnanensis were optimized, which provided technical support for the exploitation and utilization of P. polyphylla var. yunnanensis. MethodWith rhizome of P. polyphylla var. yunnanensis as experimental material and total extracted amount of six nucleosides (cytidine,uridine,guanosine,thymidine,adenosine and 2'-deoxyadenosine) as examining indexes,high performance liquid chromatography(HPLC) method was adopted to determine the extraction amount. First, the effects of ultrasonic extraction and hot water extraction on the extracted amount of nucleosides from P. polyphylla var. yunnanensis were analyzed, then based on single factor experiments, methanol volume fraction, material liquid ratio, extraction time and extraction frequency were selected as experimental factors. Extraction of nucleosides from P. polyphylla var. yunnanensis was optimized by using L9(34) orthogonal experiment to select the optimal technical conditions. ResultThe total extracted amount of six nucleo-sides by ultrasonic extraction was higher than that by hot water extraction. The effects of four experimental factors on total extracted amount of six nucleosides from P. polyphylla var. yunnanensis were ranked as follows:methanol volume fraction>extraction time>extraction frequency>material-liquid ratio. The optimal extraction conditions were as follows: pure water (methanol volume fraction was 0) was used as extraction solvent, material-liquid ratio 1:10(g/mL), extraction time 60 min at 30 ℃ and extracting twice. Under these conditions, the total extracted amount of cytidine, uridine, guanosine, thymi-dine, adenosine and 2'-deoxyadenosine in rhizome of P. polyphylla var. yunnanensis was 1.4216±0.0198 mg/g, which was higher than theoretical value 1.3434 mg/g. ConclusionThe ultrasonic extraction method optimized by orthogonal test is simple and reliable, the extraction rate of ultrasonic extraction is higher than hot water extraction. It is suitable for the ex-traction of nucleosides from P. polyphylla var. yunnanensis.