A validation protocol for the HPTLC standardization of herbal products: application to the determination of acteoside in leaves of Plantago palmata Hook. f.s

Formal validation, that is the study of the analytical performances of a method, is recognized as the best safeguard against the generation and publication of data with low reliability.Although the topic of HPTLC validations has been largely investigated, there is still a need for a general validation method applicable whenever a blank matrix cannot be reconstituted, notably herbs and their extracts.This work proposes two validation schemes aiming at generate linearity, accuracy and precision data in a minimal number of HPTLC plates, taking the standardization of Plantago palmata as an example with both UV and visible (post-chromatographic derivatization with a sulphuric acid-vanillin reagent) detections. A major problem associated with HPTLC determinations is underlined, namely the low range of linearity which makes spiking studies quite difficult as care must be taken to avoid overloading, whereas keeping the analyte detectable in blank extracts and avoiding spikes too close to endogenous levels. A second problem is the use of general post-chromatographic derivatization reagents that compromise the selectivity of the method by reacting with compounds that may not be resolved from the compound of interest. The use of such reagents is clearly not without danger, especially given the relatively low resolution of planar chromatography.In conclusion, the retained validation protocol effectively yields the main validation data whereas allowing to pinpoint major analytical drawbacks. It was not possible to simultaneously validate aucubin and acteoside assays as both analytes are present at too different levels/detectabilities.     

Measurement of free hydroxytyrosol in microdialysates from blood and brain of anesthetized rats by liquid chromatography with fluorescence detection

Hydroxytyrosol [4-(2-hydroxyethyl)-1,2-benzenediol] is a well known natural polyphenolic component with antioxidative effects from olive oil and an aglycone of acteoside. In order to examine the in vivo metabolism of acteoside to hydroxytyrosol and the distribution of hydroxytyrosol in the blood and brain, microdialysis coupled to a liquid chromatographic system was developed to evaluate the pharmacokinetics of free-form hydroxytyrosol in rat blood and brain. Probes were implanted in the jugular vein and the brain hippocampus for blood and brain sampling purposes. Hydroxytyrosol in the microdialysis samples was separated by a reversed-phase C18 column and eluted with a mobile phase containing acetonitrile – 2% acetic acid (pH 2.6) (12:88, v/v), using a flow rate for the mobile phase of 1 mL/min. Fluorescence detection for hydroxytyrosol was set at 281 nm and 316 nm for excitation and emission wavelengths, respectively. Hydroxytyrosol and endogenous interference could be resolved within 10 min by the developed chromatographic method. The results indicated that acteoside was metabolized immediately to hydroxytyrosol in vivo and eliminated rapidly from the blood, and hydroxytyrosol could enter the brain. The blood-to-brain distribution ratio was defined by dividing the area under concentration versus time (AUC) ratio of AUC_brain/AUC_blood, which represents the AUC for brain and blood. The results suggested that the P-glycoprotein was not involved in the brain efflux transport of hydroxytyrosol.     

苯丙素苷Acteoside,Isoacteoside和Ligupurpuroside J的全合成

发展了一条合成苯丙素苷类化合物的通用路线,并依据此路线完成了苯丙素苷毛蕊花糖苷（Acteoside）和异毛蕊花糖苷（Isoacteoside）的全合成及紫茎女贞苷J（Ligupurpuroside J）的首次全合成.其中的关键步骤是应用金（Ⅰ）催化的鼠李糖邻炔基苯甲酸酯给体与多羟基裸露的2-苯乙基葡萄糖苷进行区域选择性糖苷化反应,成功构建天然苯丙素苷中常见的α-（1→3）糖苷键.该合成路线减少了保护基的使用,简洁高效.     

Simultaneous Determination of Acteoside, Astragaloside IV and Icariside-I in the Traditional Chinese Medicinal Preparation Shenbao by HPLC–MS

A liquid chromatography–electrospray (ES)–mass spectrometric method for the simultaneous determination of Acteoside, Astragaloside IV and Icariside-I in the Traditional Chinese Medicinal Preparation Shenbao tablets is described. The samples were separated on an Alltima C₁₈ column by linear gradient elution using water–acetonitrile as the mobile phase. Some operational parameters of the ESI interface were optimized. The method is linear over the range of 0.1–10μg mL⁻¹ for Acteoside, Astragaloside IV, and 0.03–3μg mL⁻¹ for Icariside-I. The method has a precision (%CV) of <20%, and an accuracy (%RE) of < ± 10%. It can be used as a complementary method for quality control of Shenbao Tablets while HPLC–UV can be used for the other main components (Icariin, Icariside-II, Psoralen, Isopsoralen, and Osthol).