HPLC-DAD法测定脑血栓片中苦杏仁苷、羟基红花黄色素A、芍药苷、阿魏酸、丹酚酸B和丹参酮ⅡA

目的建立测定脑血栓片中苦杏仁苷、羟基红花黄色素A、芍药苷、阿魏酸、丹酚酸B和丹参酮ⅡA的HPLC-DAD法。方法采用HPLC-DAD法,Agilent Poroshell 120 SB-C18色谱柱(100 mm×4.6 mm,2.7μm);流动相:甲醇–0.2%磷酸溶液,梯度洗脱;检测波长分别为210 nm(苦杏仁苷)、403 nm(羟基红花黄色素A)、230 nm(芍药苷)、321 nm(阿魏酸)、286 nm(丹酚酸B)、270 nm(丹参酮ⅡA);体积流量:0.7 m L/min;柱温:30℃;进样量3~5μL。结果苦杏仁苷、羟基红花黄色素A、芍药苷、阿魏酸、丹酚酸B和丹参酮ⅡA 6个成分的线性范围分别为11.90~1 158.90、9.14~91.39、11.70~1 173.50、4.04~1 011.00、3.97~992.20、4.40~551.00 ng;平均回收率分别为96.47%、96.92%、99.96%、97.20%、97.57%、96.50%,RSD值分别为1.3%、1.6%、1.3%、1.7%、1.9%、0.7%。结论所建立的方法可同时测定脑血栓片中苦杏仁苷、羟基红花黄色素A、芍药苷、阿魏酸、丹酚酸B和丹参酮ⅡA。

Effect of matrine on proliferation inhibition and apoptosis induction in primary leukemia cells in vitro

Objective To investigate the effects of matrine on the human primary 1eukemia cells and its possible molecular mechanisms. Methods The primary leukemia cells were isolated from the PBMC of the patients with myeloid leukemia and cultivated under the regular culture medium. After exposed to the matrine with different concentration, the morphological changes of leukemia cells were observed under the light biomicroscopy. The proliferation was determined by CCK-8 assay. Annexin V-FITC/PI affinity assay was used to analyze the apoptosis of leukemia cells induced by matrine. The cell cycles were analyzed by flow cytometry before and after matrine treatment. Results Compared to the vehicle cells, there was a dramatically proliferation suppression was observed in the cells after matrine treatment. The inhibitory rates of primary leukemia cells were 54.98% and 72.96% after treated with matrine for 24 h at the doses of 0.5 and 0.8 mg/mL, respectively (P < 0.01), which has a significant dose-dependent manner. The half maximal inhibitory concentration of matrine (IC₅₀) was 0.5 mg/mL for 24 h treatment. Matrine could induce early cell apoptosis. The percentage of apoptotic cells were 11.8%, 37.6%, and 54.7% in the cells treated with 0.2, 0.5, or 0.8 mg/mL matrine for 24 h, respectively (with spontaneous apoptosis rate 7.50% for untreated cells) (P < 0.05). FCM method showed that cells at the G0/G1 phase decreased and cells at the S phase were declined obviously, suggested a blockage of cell cycles transition at the G1/S key checkpoint for 48 h. Conclusion Matrine manifests a significant inhibitory effects on the proliferation of human primary leukemia cells. Apoptosis induction and the arrest at G1 phase of cell cycle maybe contribute to its roles on the suppression of cell growth.