目的：建立超高效液相色谱-四极杆飞行时间质谱（UPLC-Q-TOF-MS）同时测定樟芝菌粉中尿嘧啶、胞苷、腺嘌呤、鸟苷、胸苷和腺苷6个核苷类成分及其指纹图谱的分析方法。方法：采用Acquity UPLC HSS T3色谱柱（2.1 mm×100 mm，1.8 μm），以甲醇-5 mmol·L-1醋酸铵溶液（pH 8.5）为流动相，梯度洗脱，流速为0.3 mL·min-1，检测波长为254 nm，柱温为30 ℃。在电喷雾正离子模式下，输入各化合物的分子离子质量数得到相应的提取离子色谱图，以色谱峰面积进行定量分析；以腺苷为参照物，测定其指纹图谱，并作相似度评价。结果：尿嘧啶、胞苷、腺嘌呤、鸟苷、胸苷和腺苷的浓度分别在0.3571~35.71 μg·mL-1（r=0.9993）、0.2032~20.32 μg·mL-1（r=0.9995）、0.0601~6.01 μg·mL-1（r=0.9998）、1.6160~161.60 μg·mL-1（r=0.9996）、0.4809~48.09 μg·mL-1（r=0.9998）、0.5988~59.88 μg·mL-1（r=0.9993）范围内与各自峰面积值呈良好的线性关系；平均回收率（n=6）分别为106.7%、103.3%、98.8%、94.6%、97.1%、102.6%，RSD分别为3.9%、5.3%、5.7%、4.8%、6.4%、4.8%；初步建立了以9个共有峰为特征指纹信息的樟芝菌粉UPLC-MS提取离子流（EIC）指纹图谱。结论：该定量方法快速、准确，所建立的提取离子流指纹图谱专属性强，两者结合可实现对樟芝菌粉内在质量的综合评价和较全面控制。
Objective: To establish an ultra-performance liquid chromatography quadrupole-time-of-flight mass spectrometric(UPLC-Q-TOF-MS)method for the fingerprint analysis and simultaneous determination of six nucleosides(uracil,cytidine,adenine,guanosine,thymidine and adenosine)in Antrodia camphorata powder. Methods: The analysis was performed on an Acquity UPLC HSS T3 chromatographic column with a gradient elution of 5 mmol·L-1 ammonium acetate(pH 8.5)and methanol solution at a flow rate of 0.3 mL·min-1.The detection wavelength was at 254 nm and the column temperature was at 30 ℃.Identification and detection were achieved in the positive electrospray ionization(ESI)mode using Q-TOF-MS.The extracted ion chromatograms of each compound were obtained by entering the molecular mass.The samples were quantified with the chromatographic peak area of each compound.Adenosine was used as a reference standard,the chromatographic fingerprint was determined.The data were analyzed by the fingerprint similarity evaluation to compare the similarity of samples. Results: Good linearity was obtained for uracil,cytidine,adenine,guanosine,thymidine and adenosine in the ranges of 0.3571-35.71 μg·mL-1(r=0.9993),0.2032-20.32 μg·mL-1(r=0.9995),0.0601-6.01 μg·mL-1(r=0.9998),1.6160-161.60 μg·mL-1(r=0.9996),0.4809-48.09 μg·mL-1(r=0.9998)and 0.5988-59.88 μg·mL-1(r=0.9993),respectively.The average recoveries(n=6)of 6 components were 106.7%,103.3%,98.8%,94.6%,97.1%,and 102.6%,respectively;RSDs were 3.9%,5.3%,5.7%,4.8%,6.4%,and 4.8%,respectively.UPLC-MS extracted ion fingerprints with 9 common peaks of Antrodia camphorata powder were established preliminarily. Conclusion: The quantitative method is rapid and accurate,and the established fingerprint has strong specificity.A combination of both can be used for a comprehensive evaluation and full control of the intrinsic quality of Antrodia camphorata powder.
