Department of Pathophysiology and Allergy Research, Medical University of Vienna, Vienna, Austria.
J Ethnopharmacol. 2013 Sep 16;149(2):562-9. doi: 10.1016/j.jep.2013.07.022. Epub 2013 Jul 24.
Sini Tang (SNT) is a traditional Chinese herbal formulation consisting of three different herbs: Aconitum carmichaelii (Fuzi), Zingiber officinale (Ganjiang), and Glycyrrhiza uralensis (Gancao). For this study, we modified this mixture by adding the bark of Cinnamomum cassia (Rougui, ). Aconitum carmichaelii contains aconitine and its derivatives, all of which are highly toxic alkaloids. These compounds are commonly detoxified with pyrolytic and hydrolytic pretreatments, such as Heishunpian, which requires repeated soaking in salt water, boiling until the roots turn black, and drying in the oven. We now demonstrate that Glycyrrhiza uralensis, which is often used in Traditional Chinese Medicine for detoxification, reduces the concentration of free aconitine in decoctions by forming a complex between liquiritin and aconitine.
Aqueous extracts of SNT, each individual herb or herbal mixture, and methanolic extracts of individual herbs were tested for free aconitine by HPLC coupled with a diode array detector. A detected complex was investigated by NMR and UV/vis spectroscopy. The continuous variations method and (1)H-NMR titrations provided the complex stoichiometry and binding constant. A 2D-ROESY experiment was performed to obtain the structural details of the formed complex.
A fast and simple HPLC method was developed to determine the amounts of aconitine and its derivatives found in herbal extracts. The Heishunpian pretreatment led to nearly complete pyrolysis and hydrolysis of the toxic compounds. However, in some batches, considerable amounts of aconitine remained. The addition of Glycyrrhiza uralensis to Aconitum carmichaelii, or liquiritin to free aconitine, led to a complexation with aconitine. The complex possessed a 1:1 stoichiometry and a binding constant of ca. 3000 L/mol to 4000 L/mol in mixtures of aqueous methanol.
A new HPLC based method allows the concentration of toxic aconitine and other diester diterpene alkaloids in herbal extracts to be rapidly determined. This method provides a starting point for the development of routine quality control procedures. The complexation of free aconitine by adding an excess of Glycyrrhiza uralensis or free liquiritin to SNT formulations will make these formulations safer.
四逆汤（SNT）是一种由三种不同草药组成的中药配方：乌头（Fuzi）、生姜（Ganjiang）和甘草（Gancao）。在这项研究中，我们通过添加肉桂皮（Rougui，）来修改这种混合物。乌头含有乌头碱及其衍生物，都是剧毒生物碱。这些化合物通常通过热解和水解预处理来解毒，例如黑顺片，需要反复浸泡在盐水中，煮沸直到根部变黑，然后在烤箱中干燥。我们现在证明，甘草常用于中药解毒，通过与乌头碱形成复合物来降低汤剂中游离乌头碱的浓度。
用高效液相色谱法（HPLC）与二极管阵列检测器耦合，测试 SNT、各单味草药或草药混合物的水提取物以及各单味草药的甲醇提取物中的游离乌头碱。通过 NMR 和紫外/可见光谱研究检测到的复合物。连续变化法和（1）H-NMR 滴定法提供了复合物的化学计量和结合常数。进行二维 ROESY 实验以获得形成复合物的结构细节。
开发了一种快速而简单的 HPLC 方法来测定草药提取物中发现的乌头碱及其衍生物的含量。黑顺片预处理导致有毒化合物几乎完全热解和水解。然而，在一些批次中，仍有相当数量的乌头碱残留。甘草加入乌头，或游离甘草酸加入游离乌头碱，都会导致与乌头碱形成复合物。该复合物具有 1:1 的化学计量比，在水甲醇混合物中的结合常数约为 3000 L/mol 至 4000 L/mol。
一种新的基于 HPLC 的方法可以快速测定草药提取物中有毒乌头碱和其他二酯二萜生物碱的浓度。该方法为常规质量控制程序的开发提供了起点。通过向 SNT 制剂中添加过量的甘草或游离甘草酸来结合游离乌头碱，将使这些制剂更安全。
