HPLC法测定各沪产地龙和广地龙中次黄嘌呤、黄嘌呤、尿嘧啶和尿苷的含量

HPLC in determination of hypoxanthine,xanthine,uridine and uracil in Shanghai Pheretima and Pheretima aspergillum(E.Perrier)

目的测定各种沪产地龙与广地龙中次黄嘌呤、黄嘌呤、尿嘧啶、尿苷的含量,建立地龙药材质量评价的方法。方法用0.9%生理盐水超声提取地龙,SORBAX SB-Aq色谱柱（250 mm×4.6 mm,5μm,Aglient Co.,Ltd）,流动相：5mmol/L磷酸二氢钾（pH 2.9）,流速1mL/min,检测波长254nm,柱温30℃,进样量10μL,采用HPLC法,同时测定次黄嘌呤、黄嘌呤、尿嘧啶、尿苷4种核苷类成分的含量。结果次黄嘌呤的线性范围为0.500 0-100.00μg（r=0.999 9）,平均回收率99.37%,RSD=1.36%（n=6）;黄嘌呤的线性范围为0.500 0-100.00μg（r=0.993 1）,平均回收率91.57%,RSD=1.40%（n=6）;尿嘧啶的线性范围为0.500 0-100.00μg（r=0.999 9）,平均回收率95.31%,RSD=1.64%（n=6）;尿苷的线性范围为0.500 0-100.00μg（r=0.999 9）,平均回收率100.21%,RSD=1.98%（n=6）。结论该方法重复性、回收率好,可用于测定地龙与广地龙中次黄嘌呤等4种核苷类成分的含量。

Objective To establish an effective method for the determining hypoxanthine,xanthine,uridine and uracil in earthworm in Shanghai Pheretima and Pheretima aspergillum（E.Perrier）,contributing to quality control of the medicinal material.Methods Hypoxanthine,xanthine,uridine and uracil were extracted from the earthworms with 0.9% NaCl by ultrasonic and determined by HPLC.The chromatographic conditions were：SORBAX SB-Aq column（250 mm×4.6 mm,5μm,Aglient Co.,Ltd）,5 mmol/L KH2PO4（pH 2.9）as the mobile phase with a flow rate of 1 mL/min,the detection wavelength was 254 nm,the column temperature was set at 30℃,and the injection volume was 10μL.Hypoxanthine,xanthine,uridine and uracil were determined by HPLC at the same time.Results The linear range was 0.5-100μg（r=0.999 9）for hypoxanthine,with the average recovery being 99.37%,RSD=1.36%（n=6）.The linear range was 0.5-100μg（r=0.993 1）for xanthine,with the average recovery being 91.57%,RSD=1.40%（n=6）.The linear range was 0.5-100μg（r=0.999 9）for uridine,with the average recovery being 95.31%,RSD=1.64%（n=6）.The linear range was 0.5-100μg（r=0.999 9）for uracil,with the average recovery being 100.21%,RSD=1.98%（n=6）.Conclusion The current method is reproducible and has satisfactory recovery,and it can be used to determine hypoxanthine,xanthine,uridine and uracil in earthworm medicinal material.