Quality Evaluation of Shexiang Tongxin Dropping Pill Based on HPLC Fingerprints Combined with HPLC-Q-TOF-MS/MS Method.

Shexiang Tongxin Dropping Pill (STP) is a composite formula of traditional Chinese medicine that is widely used for the treatment of cardiovascular diseases. It consists of seven medicinal extracts thereof or materials, including Bufonis venenum, synthetic Moschus, Panax ginseng, Bovis calculus artifactus, Bear bile powder, Salvia miltiorrhiza Bge and synthetic borneol. However, it is considerably difficult to evaluate the quality of STP due to its complex chemical compositions. This paper was designed to explore a comprehensive and systematic method combining fingerprints and chemical identification for quality assessment of STP samples. Twenty batches of STP samples were analyzed by high-performance liquid chromatography (HPLC) and high-performance liquid chromatography-quadrupole time-of-flight tandem mass spectrometry. Ten common peaks were detected by HPLC fingerprint similarity evaluation system. Meanwhile, 100 compounds belonging to 4 structural characteristics, including 23 bufadienolides, 36 organic acids, 34 saponins and 7 other types, were systematically identified as the basic components in STP. This study could be used for clarifying the multiple bioactive substances and developing a comprehensive quality evaluation method of STP.

Stereoselective Synthesis of Monofluoroalkenylphosphine Oxides via Photoinduced Decarboxylative Coupling of α-Fluoroacrylic Acids with P(O)H Compounds.

Herein, a practical and efficient method for synthesizing monofluoroalkenyl phosphine oxides via photoinduced decarboxylative/dehydrogenative coupling of α-fluoroacrylic acids with phosphine oxides and phosphonates has been developed. Various α-fluoroacrylic acids and P(O)H compounds containing relevant functional groups, including tetrafluorobenzene and pentafluorobenzene, were converted into corresponding products with excellent E-stereoselectivity in satisfactory yields. This method can be extended to achieve the synthesis of monofluoroalkenyl silanes under similar conditions.

Identification of Antitumor Constituents in Toad Venom by Spectrum-Effect Relationship Analysis and Investigation on Its Pharmacologic Mechanism.

(1) Background: Toad venom (Bufonis Venenum, known as 'Chansu' in Chinese), the secretion of the ear-side gland and skin gland of Bufo gargarizans cantor or Duttaphrynus melanostictus Schneider, has been utilized to treat several diseases in China for thousands of years. However, due to the chemical variability of the components, systematic chemical composition and the key pharmacophores in toad venom have not yet fully understood. Besides, it contains a variety of effective compounds with different physiological activity and chemotypes, mainly including alkaloids, bufogenins, bufotoxins, and so on. The recent pharmacological researches have demonstrated that several bufogenins have remarkable pharmacological effects, such as anti-inflammatory, analgesic effects, and anti-tumor effects. Aim of the study: To identify the bioactive compounds and pharmacophores originating from toad venom based on analyzing spectrum-effect relationship by chemometrics and to explore the anti-cancer mechanism primarily. (2) Materials and methods: Fingerprint of the 21 batches of samples was established using HPLC (High Performance Liquid Chromatography). The anti-tumor activity of extracts were determined by in-vitro assays. Chemometric analysis was used to establish the spectrum-effect model and screen for active ingredients. Pharmacodynamic tests for the screened active compound monomers were conducted with in-vitro assays. Further anti-tumor mechanisms were investigated using western blot and flow cytometry. (3) Results: The established spectrum-effect model has satisfactory fitting effect and predicting accuracy. The inhibitory effect of major screened compounds on lung carcinoma cells A549 were validated in vitro, demonstrating that arenobufagin, telocinobufogenin, and cinobufotalin had significant anti-tumor effects. Through further investigation of the mechanism by western blotting and flow cytometry, we elucidated that arenobufagin induces apoptosis in A549 cells with the enhanced expression of cleaved PARP (poly (ADP-ribose) polymerase). These results may provide valuable information for further structural modification of bufadienolides to treat lung cancer and a method for discovery of anti-tumor active compounds. Conclusions: Our research offers a more scientific method for screening the principal ingredients dominating the pharmacodynamic function. These screened compounds (arenobufagin, etc.) were proven to induce apoptosis by overactivation of the PARP-pathway, which may be utilized to make BRCA (breast cancer susceptibility gene) mutant cancer cells more vulnerable to DNA damaging agents and kill them.

[Correlative study of bufogenin constituents in venom of Bufo bufo gargarizans].

As known,simultaneous determination of various chemical indicators is one of the future trends in quality control of traditional Chinese medicines because of the extremely complex chemical compositions. This project is to screen the quality markers that can accurately control the quality of the Bufonis Venenum by exploring the intrinsic correlation of components. In this study,venom of Bufo bufo gargarizans from 17 different sources were used as research samples,and the contents of 7 bufogenin were determined by HPLC-DAD. Then,the data obtained were analyzed by Spearman correlation analysis and principal component analysis( PCA). In addition,a stepwise regression analysis was used to establish a predictive model for the contents of the seven bufogenin components( independent variable) and the total contents of the bufogenin( dependent variable). The results indicated that there is a significant positive correlation between the contents of telocinobufagin and cinobufotalin,and there is a significant positive correlation between the contents of bufalin,cinobufagin and resibufogenin. In contrast,the contents of telocinobufagin and cinobufotalin are negatively correlated with the contents of bufalin,cinobufagin and resibufogenin. However,the correlation between gamabufotalin and bufotalin and other components are not obvious. Furthermore,further study found that there is a correlation between the sum of the contents of bufalin,cinobufagin and telocinobufagin and the total contents of the bufogenin. In fact,the application of bufalin,cinobufagin and telocinobufagin as the quality control indicators of the Bufonis Venenum can better reflect the quality characteristics of the Bufonis Venenum compared with the previous quality control indicators. The conclusions will provide a reference for the revision of the quality standards of the Bufonis Venenum.

[Modeling of thermal degradation of linarin during concentration process].

Ganmaoling granule, with annual sale of over one billion yuan, is the first brand of domestic cold medicine sales. As the only traditonal Chinese medicine（TCM） quality control indicator of Ganmaoling granule, linarin is thermally unstable. Its content will be changed significantly during the production process, which would then affect the quality of the finished product. In this paper, the law of degradation of linarin was investigated. The experimental results showed that degradation reaction of linarin belongs to the first reaction characteristics. The effective methods to reduce the loss of linarin would be realized fortunately by strictly controlling the heating temperature or shortening the heating time.

[Study on balance of process of alcohol precipitation of ganmaoling granules].

Ganmaoling granule is the first brand of domestic cold medicine sales, but its preparation method and process control parameters are relatively rough. Therefore it is urgent to upgrade the technologies of large varieties of traditional Chinese medicine (TCM). This paper focused on the balance between the remove of impurity and the retention of linarin during the process of alcohol precipitation of Ganmaoling granules. The effects of four factors on the process were investigated via single factor experiments. The results showed that the precipitating period, the initial ethanol concentration and the final ethanol concentration had a great effect on retention of linarin while the initial density of the extract has not. Similarly, the initial ethanol concentration, the final ethanol concentration and the initial extract density have a great effect on the yield of dry extract while the time of alcohol precipitation has not. The parameters of alcohol precipitation of Ganmaoling granules were optimized as 16 h of precipitating period, 95% ethanol as the initial reagent, 70% of the final ethanol concentration, and 1.10 of the initial extract density.

[On-line monitoring of extraction of coldrine by near-infrared spectrum].

Extraction of the four Chinese herbals is the beginning step of the production process of coldrine granules and influences on drug quality significantly. In this paper, the on-line near infrared spectrum was collected during the extraction process of coldrine and then pre-processed by the first derivative. Partial least square regression (PLSR) model was developed for the quantity indicators of linarin, chlorogenic acid and solid content, according to results of both HPLC and weight-loss as reference methods. The correlation coefficient, root mean square error of cross-validation (RMSECV), root mean square error of calibration (RMSEC) and root mean square error of prediction (RMSEP) were used to optimize model parameters and confirm their performance. Correlation coefficients of three quality control indicator models reached more than 0.95.Values of RMSEC of linarin, chloroenic acid and solid content were 0.010 4, 0.009 34 and 0.055 5, respectively. And the values of RMSEP were 0.009 47, 0.142 and 0.008 42, respectively. The models, built on-line analyze data, revealed that the correlation coefficients of predicted values and measured values were greater than 0.97 and values of RSEP of linarin, chloroenic acid and solid content were 8.14%, 8.17% and 9.86%, respectively. The results showed that the NIR method could achieve the on-line detection and real-time monitoring of multi-indexes during the extraction process of coldrine. The technology could be used for drug quality control in the process of practical production, reducing the batch differences and ensuring pharmaceutical quality stability. In addition, it could provide real-time production data for subsequent product quality backtracking.