Rosenthal Immanuel, Wolfram Evelyn, Meier Beat
Zurich University of Applied Science, Institute of Biotechnology, Research Group of Phytopharmacy, Einsiedlerstrasse 31, 8820 Wädenswil, Switzerland,
Pharmeur Bio Sci Notes. 2014;2014:92-102.
The current Ph. Eur. monographs for senna pods, senna leaf and senna leaf dry extract standardised describe a photometric assay based on the Bornträger reaction to determine hydroxyanthracene glycosides, calculated as sennoside B. The method is timeconsuming, unspecific for sennosides and the precision is not adequate for a modern assay.
The photometric method shall therefore be replaced by a modern HPLC method. About 70 % of the total anthrachinone content in herbal drugs of senna species is due to sennoside A and sennoside B. These substances are therefore suitable for the standardisation of Senna products. The Japanese Pharmacopoeia (JP) already describes an HPLC method to determine sennoside A and sennoside B in the monograph for senna leaf. It uses ion-pair chromatography with tetraheptylammoniumbromide. The procedure described in the monograph has a runtime of 70 min.
The adapted and validated method described here uses solid-phase extraction (SPE) which allows a selective sample preparation by using an anion exchange phase. A conventional RP C18 column Tosh TSKgel ODS-80TS (4.6 mm × 150 mm), 5 μm, was used as stationary phase and acetonitrile for chromatography R, water R, phosphoric acid R (200:800:1 V/V/V) as mobile phase. The flow rate was 1.2 mL/min, the column temperature 40 °C, the detection wavelength 380 nm, and the injection volume 20 μL. The runtime is 10 min, the chromatogram shows 2 peaks due to sennoside A/B and 2 additional smaller compounds. One of them is rhein-8-O-glucoside.
The procedure has been successfully validated according to ICH guidelines. We analysed 6 batches of Senna. The pods (Senna angustifolia) showed a total content of sennoside A and B of 1.74-2.76 % m/m and the content of senna leaves was clearly lower with 1.07-1.19 % m/m, respectively.
The suggested method is considered to be suitable to determine sennoside A and sennoside B in senna leaves and senna pods. The consideration is based on the performed validation and on the results for the analysed samples. A short run time and better resolution are clear advantages of the suggested method, compared to other methods.
现行欧洲药典中关于番泻豆荚、番泻叶和标准化番泻叶干提取物的各论描述了一种基于Bornträger反应的光度法，用于测定羟基蒽醌苷，以番泻苷B计算。该方法耗时，对番泻苷缺乏特异性，且精密度不足以满足现代分析要求。
因此，光度法应由现代高效液相色谱法替代。番泻属草药中总蒽醌含量的约70%归因于番泻苷A和番泻苷B。因此，这些物质适用于番泻产品的标准化。日本药典（JP）已在番泻叶各论中描述了一种测定番泻苷A和番泻苷B的高效液相色谱法。它使用含四庚基溴化铵的离子对色谱法。各论中描述的方法运行时间为70分钟。
此处所述的经过调整和验证的方法使用固相萃取（SPE），通过使用阴离子交换相实现选择性样品制备。使用常规反相C18柱Tosh TSKgel ODS - 80TS（4.6 mm×150 mm），5μm，作为固定相，乙腈用于流动相R，水R、磷酸R（200:800:1 V/V/V）作为流动相。流速为1.2 mL/min，柱温40℃，检测波长380 nm，进样体积20μL。运行时间为10分钟，色谱图显示由于番泻苷A/B出现2个峰以及另外2个较小的化合物峰。其中之一是大黄酸 - 8 - O - 葡萄糖苷。
该方法已按照ICH指南成功验证。我们分析了6批番泻。豆荚（狭叶番泻）中番泻苷A和B的总含量为1.74 - 2.76% m/m，而番泻叶的含量明显较低，分别为1.07 - 1.19% m/m。
建议的方法被认为适用于测定番泻叶和番泻豆荚中的番泻苷A和番泻苷B。这一结论基于所进行的验证以及分析样品的结果。与其他方法相比，建议方法运行时间短且分离度更好，具有明显优势。
