LC-MS/MS测定中成药中非法添加物N-乙基他达拉非

Detection of N-ethyl Tadalafil Added Illegally into Traditional Chinese Medicines by LC-MS/MS

目的建立LC-MS/MS定性及定量测定中成药中非法添加N-乙基他达拉非。方法采用DAD检测器进行初步定性筛查,用质谱检测器进行定性确证,再用DAD检测器进行定量。初筛及定量选用SHISEIDO C18柱(250 mm×4.6 mm,5μm),以流动相A[磷酸三乙胺溶液(取三乙胺7 m L,用水稀释至1 000 m L,用磷酸调p H至2.8)-甲醇-乙腈(60∶20∶20)]与流动相B[磷酸三乙胺溶液(取三乙胺7 m L,用水稀释至1 000 m L,用磷酸调p H至2.8)-甲醇-乙腈(8∶46∶46)]梯度洗脱,体积流量1 m L·min-1,检测波长230 nm;确证选用Dikma Spursil C18柱(150 mm×2.1 mm,3μm),以甲醇-乙腈-含0.1%冰乙酸的0.02 mol·L-1乙酸铵溶液(30∶25∶45)等度洗脱,体积流量0.2 m L·min-1。结果 N-乙基他达拉非质量浓度在0.003~0.3 mg·m L-1内与峰面积有良好线性关系,平均回收率为97.8%,RSD为0.7%(n=9);通过对30批次壮阳类中成药进行测定,其中2批检测结果为阳性,含量分别为4.8,4.1 mg·g-1。结论该方法快速、准确、灵敏度高,可作为分析检测壮阳类中成药中非法添加N-乙基他达拉非的有效方法。

OBJECTIVE To establish an HPLC-MS/MS method for the determination of N-ethyl tadalafil which was added illegally into traditional Chinese medicines（TCM）. METHODS The experiment steps was as follows： N-ethyl tadalafil was preliminary screened firstly by DAD detector, and then confirmed by mass spectrometer detector and quantitative determined by DAD detector at last. The preliminary screening and qualitative analysis was performed by HPLC-DAD, which was conducted on a SHISEIDO C18 column（250 mm×4.6 mm, 5 μm） with gradient elution at a flow rate of 1 m L·min^-1. The mobile phase A consisted of water containing 0.7% triethylamine（adjusted to p H 2.8）, methanol and acetonitrile（60∶20∶20）; the mobile phase B consisted of water containing 0.7% triethylamine（adjusted to p H 2.8）, methanol and acetonitrile（8∶46∶46）. The sample was detected at 230 nm. The confirming test was performed by LC-MS/MS, which was conducted on a Dikma Spursil C18 column（150 mm×2.1 mm, 3 μm） at a flow rate of 0.2 m L·min^-1 with mobile phase consisted of methanol-acetonitrile-0.02 mol·L^-1 ammounium acetate（contained 0.1% acetic acid）（30∶25∶45）. RESULTS The linear relationship was good at the range of 0.003-0.3 mg·m L^-1, and the average recovery was 97.8%（n=9） with RSD of 0.7%. It was confirmed that N-ethyl tadalafil was added illegally in two batches of sthenic TCM through screening 30 batches of TCM, and the content of N-ethyl tadalafil was 4.8 mg·g^-1 and 4.1 mg·g^-1, respectively. CONCLUSION The method is fast, accurate and highly sensitive and can be used to screen the illegally sthenic TCM spiked with N-ethyl tadalafil.