HPLC-DAD法同时测定益肺清化颗粒的10种成分

Simultaneous determination of ten constituents in Yifei Qinghua Granules by HPLC-DAD

目的建立HPLC-DAD法同时测定益肺清化颗粒中苦杏仁苷、齐墩果酸、熊果酸、仙鹤草酚B、甘草苷、毛蕊异黄酮葡萄糖苷、鲁斯可皂苷元、没食子酸、补骨脂素和紫菀酮的方法。方法采用反相液相色谱法,色谱柱为Waters Symmetry-C_(18)(250 mm×4.6 mm,5.0μm);流动相为KH2PO4缓冲溶液(KH2PO4 1.0 g,加水1 000 mL使溶解,磷酸调节p H值至3.5)-(乙腈-甲醇1∶1),梯度洗脱,体积流量0.8 mL/min;柱温40℃。结果苦杏仁苷、齐墩果酸、熊果酸、仙鹤草酚B、甘草苷、毛蕊异黄酮葡萄糖苷、鲁斯可皂苷元、没食子酸、补骨脂素和紫菀酮10种成分能够达到良好分离;其线性范围分别为0.2～2.0μg/mL(r=0.999 5)、0.4～4.0μg/mL(r=0.999 4)、0.3～3.0μg/mL(r=0.999 6)、0.1～1.0μg/mL(r=0.999 3)、0.5～5.0μg/mL(r=0.999 1)、0.15～1.50μg/mL(r=0.999 2)、0.25～2.50μg/mL(r=0.999 5)、0.6～6.0μg/mL(r=0.999 1)、0.45～4.50μg/mL(r=0.999 3)、0.12～1.20μg/mL(r=0.999 4),平均加样回收率分别为98.7%、98.0%、99.6%、98.0%、99.1%、99.4%、98.8%、101.1%、100.6%、101.7%,RSD分别为0.7%、1.3%、1.4%、1.4%、1.1%、0.8%、0.9%、0.5%、0.5%、0.8%。10批次供试品中苦杏仁苷、齐墩果酸、熊果酸、仙鹤草酚B、甘草苷、毛蕊异黄酮葡萄糖苷、鲁斯可皂苷元、没食子酸、补骨脂素和紫菀酮质量浓度分别为0.104～0.123、0.600～0.621、0.501～0.523、0.100～0.121、0.103～0.122、0.231～0.260、0.043～0.065、0.055～0.069、0.061～0.079、0.031～0.043 mg/g。结论本方法操作简便,结果准确可靠,可用于益肺清化颗粒的质量控制。

Objective To establish an HPLC-DAD method for the simultaneous determination of amygdalin, oleanolic acid, ursolic acid, agrimol B, liquiritin, calycosin-7-glucoside, ruscogenin, gallic acid, psoralen, and shionone in Yifei Qinghua Granules（YQG）. Methods The chromatographic separation was achieved on an Waters Symmetry-C_（18）（250 mm × 4.6 mm, 5.0 μm） column with mobile phase consisted of（1.0 g potassium phosphate monobasic in 1 000 mL water, the p H value was adjusted to 3.5 with phosphate）-（acetonitrile-methanol 1∶1） for gradient elution, at the flow rate of 0.8 mL/min; The column temperature was 40 ℃. Results The linear ranges of amygdalin, oleanolic acid, ursolic acid, agrimol B, liquiritin, calycosin-7-glucoside, ruscogenin, gallic acid, psoralen, and shionone were 0.2—2.0 μg/mL（r = 0.999 4）, 0.4—4.0 μg/mL（r = 0.999 4）, 0.3—3.0 μg/mL（r = 0.999 6）, 0.1—1.0 μg/mL（r = 0.999 3）, 0.5—5.0 μg/mL（r = 0.999 1）, 0.15—1.50 μg/mL（r = 0.999 2）, 0.25—2.50 μg/mL（r = 0.999 5）, 0.6—6.0 μg/mL（r = 0.999 1）, 0.45—4.50 μg/mL（r = 0.999 3）, and 0.12—1.20 μg/mL（r = 0.999 4）, respectively. These ten components were well resolved. Their average recoveries（n = 6） respectively were 98.7%（RSD = 0.7%）, 98.0%（RSD = 1.3%）, 99.6%（RSD = 1.4%）, 98.0%（RSD = 1.4%）, 99.1%（RSD = 1.1%）, 99.4%（RSD = 0.8%）, 98.8%（RSD = 0.9%）, 101.1%（RSD = 0.5%）, 100.6%（RSD = 0.5%）, and 101.7%（RSD = 0.8%）. The content of ten batches of the amygdalin, oleanolic acid, ursolic acid, agrimol B, liquiritin, calycosin-7-glucoside, ruscogenin, gallic acid, psoralen, and shionone was 0.104—0.123, 0.600—0.621, 0.501—0.523, 0.100—0.121, 0.103—0.122, 0.231—0.260, 0.043—0.065, 0.055—0.069, 0.061—0.079, 0.031—0.043 mg/g, respectively. Conclusion The method is accurate, sensitive, credible, and repeatable, which can be applied to the quality control of YQG.