摘要:
目的 提取附子总生物碱,采用高效液相色谱法同时测定附子总生物碱提取物中苯甲酰新乌头碱、苯甲酰乌头原碱、苯甲酰乌头次碱、新乌头碱、乌头碱和次乌头碱6个成分的含量。 方法 采用85%的乙醇回流提取附子药材,经大孔吸附树脂柱分离后,制得附子总生物碱提取物,采用Waters Terra C18色谱柱(3.0 mm×100mm, 3.5μm);流动相为A:乙腈,B:水(5 mM醋酸铵),梯度洗脱,A相随时间的变化:15%~26%(0~10 min),26%~30%(10~20min),30%(20~25 min);流速0.5 ml/min;检测波长235 nm;柱温25℃;进样量5 μl。 结果 6个生物碱成分苯甲酰新乌头碱、苯甲酰乌头原碱、苯甲酰次乌头碱、新乌头碱、乌头碱、次乌头碱在20 min内实现完全分离,在11.40~114.0、10.22~102.2、10.36~103.6、6.250~62.50、13.50~135.0、15.12~151.2 μg/ml的线性范围内,线性关系良好(r>0.999)。方法学考察结果表明,日内日间精密度,最低检测限和定量限的范围均符合含量测定的要求,加样回收率(n=6)分别为104.52% (RSD=2.0%)、103.43%(RSD=1.6%)、101.92%(RSD=2.3%)、99.45%(RSD=2.7%)、97.67%(RSD=1.9%)、101.15%(RSD=3.4%)。 结论 该方法简便、准确、可靠,可用于附子提取物中6个乌头碱成分的含量测定。
Abstract:
Objective To extract alkaloids in Radix Aconiti Praeparata (Fuzi) and determine the contents of six alkaloids benzoylmesaconine, benzoylaconine, benzoylhyaconine, mesaaconine, aconine and hyaconine by high performance liquid chromatography (HPLC). Methods The crude herb Fuzi was extracted by 85% ethanol, and separated by macroporous resin column. HPLC condition was as follows:Chromatographic column:Waters Terra C18 (3.0 mm×100 mm, 3.5 μm); Mobile phase:A was acetonitrile, B was water (5 mM ammonium acetate), with gradient elution, the gradient of A phase:15%~26%(0~10 min), 26%~30%(10~20 min), 30%(20~25 min); flowing rate:0.5 ml/min; detection wave:23 5nm; column temperature:25℃; injection volume:5 μl. Results Six alkaloids, including benzoylmesaconine, benzoylaconine, benzoylhyaconine, mesaconine, aconine and hyaconine were separated at baseline within 25 min with good linear (r>0.999 0) between 11.40~114.0, 10.22~102.2, 10.36~103.6, 6.250~62.50, 13.50~135.0, 15.12~151.2 μg/ml. The result of intra-day and inter-day precisions, limits of detection and quantitation were at normal range. Each of the recovery rates (n=6) was 104.52% (RSD=2.0%), 103.43%(RSD=1.6%), 101.92%(RSD=2.3%), 99.45%(RSD=2.7%), 97.67%(RSD=1.9%), 101.15%(RSD=3.4%). The contents of six alkanoids in Fuzi alkaloids extract were determined. Conclusion The method was rapid, simple, reliable and could be used to determine six alkanoids in Fuzi alkanoids extract.
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