目的 建立同时分析冬虫夏草中15种核苷类成分的高效液相色谱法(HPLC)方法,比较青海玉树州5县12个不同产地样品中该15种成分的组成和含量的差异。方法 采用Zorbax Bonus-RP(4.6 mm×250 mm, 5 μm)色谱柱,甲醇-0.04%冰乙酸水溶液为流动相,梯度洗脱,柱温35 ℃,流速1.0 mL·min^-1,检测波长260 nm为分析条件,检测12个不同产地样品中15种核苷类成分,并对数据进行初步比较分析。结果 样品中15种核苷类成分分离良好,在各自相应的浓度内,相关系数r在0.999 1~0.999 8之间,具有良好的线性;15种核苷类成分平均加标回收率在92.3%～101.7%之间,RSD均小于等于3.1%(n=6)。不同产地冬虫夏草样品中15种核苷类成分在组成和含量上均存在差异。15种核苷类成分总含量分布在7.820~14.351 mg·g^-1内;尿苷,鸟苷、肌苷、胸苷含量高于其余11种核苷类成分,其含量分别在2.244~4.997 mg·g^-1、1.017~2.388 mg·g^-1、0.810~3.105 mg·g^-1、0.377~2.997 mg·g^-1内。在同一样品中,尿苷、鸟苷、肌苷、胸苷4种核苷占15种核苷类成分总量的比例为58.169%~90.724%。除治多县样品为0.095 mg·g^-1外,其余11个不同产地样品中腺苷含量均高于现行中国药典要求的0.01%(0.10 mg·g^-1)的限量标准,并且在同一样品中腺苷和虫草素的含量基本接近。结论 本实验建立的冬虫夏草中15种核苷检测方法简便、准确,重复性好,能够满足冬虫夏草中核苷类成分分析的要求。对青海玉树5县12个不同产地中15种核苷类成分的含量检测和比较数据能够为冬虫夏草质量控制方法的深入研究提供参考。

OBJECTIVE To establish an HPLC method for simultaneous determination of 15 nucleosides in Cordyceps sinensis, and investigate the differences in the composition and contents of these 15 nucleosides in 12 samples from 5 counties of Yushu prefecture,Qinghai province. METHODS The chromatographic analysis were carried out on a Zorbax Bonus-RP(4.6 mm×250 mm,5 μm) column with detection wavelength of 260 nm. The mobile phase was methanol-0.04% acetic acid solution gradiently eluted at 1.0 mL·min^-1, and the column temperature was set at 35 ℃. Then the composition and contents of the 15 nucleosides were compared and analyzed. RESULTS The 15 nucleosides in the samples were well separated. In their corresponding concentration ranges, the calibration curves of the analytes had good linearity with correlation coefficients between 0.999 1-0.999 8. The average recoveries were 92.3%-101.7%, with RSDs not more than 3.1%(n=6). There were differences in the composition and contents of the 15 nucleosides in the samples from different origins. The total contents of the 15 nucleoside were 7.820-14.351 mg·g^-1. And the contents of uridine, guanosine, thymidine, and inosine were higher than those of the other 11 nucleosides, which ranged from 2.244 to 4.997 mg·g^-1, 1.017 to 2.388 mg·g^-1, 0.810 to 3.105 mg·g^-1, and 0.377 to 2.997 mg·g^-1, respectively. In a same sample, the sum content of uridine, guanosine, thymidine and inosine accounted for 58.169%-90.724% of the total 15 nucleosides. For adenosine content, except which in the sample from Zhiduo County was 0.095 mg·g^-1, those in the other 11 samples were higher than the lower limit required by Chinese Pharmacopoeia of 0.01%. And the content of adenosine was close to that of cordycepin in the same sample. CONCLUSION The established HPLC method for determination of 15 nucleosides in Cordyceps sinensis is simple, accurate, and reproducible, which meets the requirements for nucleosides analysis in Cordyceps sinensis. The data on the contents and comparison of the 15 nucleosides provide references for further study of the quality control method of Cordyceps sinensis.