HPLC梯度洗脱法同时测定镇痛活络酊中4种成分的含量

Simultaneous determination of four components in Zhentong Huoluo Ding by HPLC

目的 建立同时测定镇痛活络酊中香蒲新苷、异鼠李素-3-O-新橙皮苷、巴西苏木素和（±）原苏木素B含量的高效液相色谱法。方法 采用依利特C18柱;流动相为乙腈（A）-0.5%磷酸溶液（B）,梯度洗脱;流速：1.3 mL/min;柱温：26℃;检测波长λ_1=254 nm（香蒲新苷及异鼠李素-3-O-新橙皮苷）,λ_2=285 nm[巴西苏木素和（±）原苏木素B]。结果 香蒲新苷、异鼠李素-3-O-新橙皮苷、巴西苏木素和（±）原苏木素B的线性范围分别为4.62~92.40μg/mL（r=0.999 2）、5.70~114.00μg/mL（r=0.999 6）、8.95~179.00μg/mL（r=0.999 4）、7.79~155.80μg/mL（r=0.999 7）;香蒲新苷、异鼠李素-3-O-新橙皮苷、巴西苏木素和（±）原苏木素B的平均加样回收率分别为97.81%、99.16%、97.23%、96.74%,其RSD分别为1.19%、0.84%、1.04%、1.08%。结论 所建立的镇痛活络酊样品中,4种成分（香蒲新苷、异鼠李素-3-O-新橙皮苷、巴西苏木素和（±）原苏木素B）的含量测定方法准确、可靠。

Objective To establish an HPLC method for the simultaneous determination of 4 main components in Zhentong Huoluo Ding, including typhaneoside, isorhamnetin-3-O-neohesperidoside, hrazilin and （ ＋ ） protosappanin B. Methods A stable HPLC method was established and the chromatography was accomplished on an Hypersil C18 column, with a mobile phase consisting of acetonitrile （A） and 0. 5% phosphoric acid solution （B） in a gradient elu- tion. The flow rate was set at 1.3 mL/min, and the column temperature was set at 26 ℃. Typhaneoside and isorhanme- tin-3-O-neohesperidoside were detected at 254 nm, while brazilin and （ ＋ ） protosappanin B were detected at 285 nm. Results Typhaneoside,isorhamnetin-3-O-neohesperidoside, hrazilin and （ ＋ ） protosappanin B had good linearity at 4. 62 -92.40 μg/mL （r =0. 999 2） ,5.70 - 114. 00 μg/mL （r =0. 999 6） ,8. 95 - 179. 00 μg/mL （r =0. 999 4） and 7.79 - 155.80 μg/mL （ r = 0. 999 7 ） , respectively; the average recovery rates were 97. 81% （ RSD -- 1. 19% ） , 99.16% （RSD=0.84% ） ,97.23% （RSD= l.04% ） and96.74% （RSD=1.08% ） ,respectively. Conclusion The developed HPLC method is accurate and reliable for the simultaneous determination of typhaneoside, isorhamnetin-3-O- neohesperidoside,brazilin and （ ± ） protosappanin B in Zhentong Huoluo Ding.