Analysis of hydroxylation and O-glycosylation on lysine sites in deer-hide gelatin / 中国中药杂志

Nano-LC MS/MS was used to analyze trypsin digested deer-hide gelatin(DHG) samples, hydroxylation and O-glycosylation on lysine sites of DHG were comprehensive identified by using PEAKS Studio software. The sites, sorts and amounts of hydroxylation and O-glycosylation on Type Ⅰ collagen α1 chain(COL1 A1) and α2 chain(COL1 A2) of DHG were revealed. As a result, 5 284 peptides were identified from DHG samples, which were mainly from COL1 A1 and COL1 A2. Among these peptides, there were 449 peptides with hydroxylysine, 442 with galactosyl-hydroxylysine, 449 with glucosyl-galactosyl-hydroxylysine. The major modified sites of hydroxylation and O-glycosylation in DHG were shown as follow: α1-9 N and α2-5 N in N-telopeptides, α1-87, α1-174, α1-930, α2-87, α2-174, α2-933 in triple helix domain, and α1-16 C in C-telopeptides. These hydroxylation and O-glycosylation were correlated with the formation and stability of collagen molecules and collagen fibrils. It is feasible for the collagens and peptides dissolving from deer skin collagen fibrils under high temperature and pressure decocting, high temperature and pressure also might destroy inter-molecular covalent cross-linking and help those glycol-peptides formations. The present study provided ideas and strategies for the in-depth investigation on DHG chemical constituents, and showed good theoretical significance and application value.

Chemical constituents of triterpenoid saponins from Glycyrrhiza glabra / 药学学报

Ten triterpenoid saponins were isolated and purified from the water extract of Glycyrrhiza glabra by polyamide resin combined with macroporous resin column chromatography, ODS medium pressure column chromatography and semi-preparative RP-HPLC. Their structures were elucidated by physicochemical properties, NMR and MS spectra, and determined as 3β-O-[β-D-glucuronpyranosyl-(1→2)-β-D-glucuronpyranosyl]-30β-O-β-D-glucuronpyranosyl-oleanane-11-oxo-12(13)-ene (1), 3β-O-[β-D-glucuronpyranosyl-(1→2)-β-D-glucuronpyranosyl]-30β-O-α-L-rhamnopyranosyl-oleanane-11-oxo-12(13)-en-22β,30-diol (2), uralsaponin C (3), licorice-saponin A3 (4), licorice-saponin P2 (5), 22β-acetoxyl-glycyrrhizin (6), macedonoside A (7), 29-hydroxyl-glycyrrhizin (8), licorice-saponin G2 (9), glycyrrhizin (10). Compounds 1 and 2 are two new compounds and named as licorice-saponin R3 and licorice-saponin S3.

Collagen derived species-specific peptides for distinguishing donkey-hide gelatin (Asini Corii Colla) / 中草药·英文版

Objective: As an important food therapy product with traditional Chinese medicine (TCM) applications, donkey-hide gelatin (Asini Corii Colla, ACC) has been used for thousands of years. However, till now few effective strategy had been proposed to distinguish ACC from other animal hide gelatins, especially closely related horse- and mule-hide gelatins, which was an embarrassment of ACC quality control. Methods: Combined mass spectrometry and bioinformatic methods have been applied to identify and verify two ACC-specific peptides (Pep-1 and Pep-2) capable of distinguishing ACC from other closely related animal gelatins with high selectivity. Results: It confirmed that these two peptides could be not only used for distinguishing ACC from highly homologous horse-hide and mule-hide gelatins as well as other animal hide gelatins. Conclusion: The present study provides a simple method for species-specific peptides discovery, which can be used for assessing the quality of animal gelatin products, and ensure they are authenticable and traceable.

Comparative analysis of chemical constituents of deer hide and deer hide gelatin by "peptidomics-modifications" methods / 药学学报

Collagen is the main constituent of gelatinous Chinese medicine, with deer hide gelatin (Cervi Corii Colla, DHG) made from deer hide (DH) through a complex thermal and high-pressure processing procedure. During this procedure some amino acids in collagen undergo hydroxylation and deamidation. In the present study, comparative analysis of proteins and peptides in DH and DHG was carried out using "peptidomics-modifications" methods. Nano-LC-MS/MS was used to analyze proteins and peptides in DH and DHG, and the number and sites of modification were determined as well. The amount of hydroxylation and deamidation that occurred in DHG was significantly greater than that in DH, suggesting that under thermal and high-pressure processing these modifications occurred more frequently on certain amino acids in collagen, and might be correlated with hydrophobicity. The occurrence and mechanism of hydroxylation and deamidation in DH processing procedures should be explored in further research. The present study provides important evidence of the chemical constituents and the correlation of processing procedures with these modifications, and also suggests some investigative ideas for DHG processing optimization and improvement of quality standards.

Analysis of quality difference based on Astragalus membranaceus var. mongholicus in genuine region / 中国中药杂志

This work is to establish the fingerprint of Astragalus membranaceus var. mongholicus by HPLC-ELSD method, and to analyze the simulated wildness degree of A. membranaceus var. mongholicus in the genuine region of Inner Mongolia, Ningxia and Gansu. Compared with wild A. membranaceus var. mongholicus, the quality differences of A. membranaceus var. mongholicus in the genuine region were analyzed by identification of chromatographic peaks and similarity evaluation, cluster analysis(CA), principal components analysis(PCA) and orthogonal partial least squares discriminant analysis(OPLS-DA). HPLC fingerprints of A. membranaceus var. mongholicus in different genuine regions are established. The qualitative analysis of mass spectrometry identified 18 components. The similarity evaluation shows that the similarity of 32 batches of A. membranaceus var. mongholicus samples was 0.688-0.993. Among them, the similarity of samples in Shanxi, Inner Mongolia, Ningxia is 0.688-0.993, 0.835-0.989, 0.934-0.988, respectively and the similarity of samples in Gansu is 0.729-0.876 except No. 25 sample. The results of CA show that the samples of A. membranaceus var. mongholicus can be grouped into four categories according to the production area except the No. 11 and No. 25 samples. The results of PCA indicate that 32 batches of A. membranaceus var. mongholicus samples can be clustered according to quality and origin, and the quality of A. membranaceus var. mongholicus in Inner Mongolia is the closest to the wild breed. The results of OPLS-DA indicate that there are six components that can distinguish the wild and domestic A. membranaceus var. mongholicus, which are malonylastragaloside Ⅰ, astragaloside Ⅰ, calycosin-7-O-β-D-glycoside-6″-O-malonate, calycosin-7-O-β-D-glycoside, formononetin-7-O-β-D-glycoside-6″-O-malonate, and astrapterocarpan-3-O-β-D-glycoside-6″-O-malonate. The established method can be used to analyze differences between A. membranaceus var. mongholicus origin and planting environment, and can provide references for the protection and replacement of wild A. membranaceus var. mongholicus resources, and the cultivation, processing and production of A. membranaceus var. mongholicus.

Textual Research on Origin of Alismatis Rhizoma in China Pharmacopoeia / 中国实验方剂学杂志

This paper origin studies the origin of Alismatis Rhizoma in Chinese pharmacopoeia， and puts forward some suggestions for modification. Through the changes in the records of the source of Alismatis Rhizoma in the various versions of the Chinese Pharmacopoeia and the records of Flora of China and Materia Medica of China，it is found that the source of Alismatis Rhizoma in the Chinese Pharmacopoeia is confused. Specifically， the Chinese name of Alismatis Rhizoma does not correspond to the Latin name. As a common Chinese herbal medicine，Alismatis Rhizoma has a large market circulation. Many classic Chinese medicine prescriptions released by China Food and Drug Administration contain Alismatis Rhizoma. The development of the classic Chinese medicine prescriptions will further increase the market circulation of Alismatis Rhizoma. As a major national move to promote the development of traditional Chinese medicine， the study for classic Chinese medicine prescriptions requires defining the origin of the medicinal materials used，and the confused origin of Alismatis Rhizoma recorded in the Chinese Pharmacopoeia seriously hinder the development of the classics. Therefore，in order to regulate the origin of Alismatis Rhizoma， ensure the clinical efficacy and promote the development of classic Chinese medicine prescriptions，the confused origin of Alismatis Rhizoma in the Chinese Pharmacopoeia has to be resolved as soon as possible. Based on the analysis of the changes of Alismatis Rhizoma's producing areas in the past dynasties， it is found that the producing areas of Alismatis Rhizoma have continuous changed from Wei and Jin dynasties to present， and finally formed the current situation of Sichuan as the main producing area. In comparison of chemical composition，origin and market circulation of Alismatis Rhizoma in Sichuan Province that is the most productive， and Fujian Province that is the best quality， it is found that the two species are different in every aspects. Nowadays，Alisma plantago-aquatica occupies the majority of the market， which doesn't conform to Alisma orientale as specified in the 2015 edition of the Chinese Pharmacopoeia. Therefore， through textual research and analysis， it is suggested that both A. plantago-aquatica and A. orientale. Shall be used as the origin of Alismatis Rhizoma. In the 2015 edition of Chinese Pharmacopoeia，Cassiae Semen，Schizonepetae Herba，Aisaematis Rhizoma，Fibraureae Caulis and Ajugae Herba have the same problem. This paper provides ideas for the revision of sources of traditional Chinese medicine.

Comparative study of water characteristic components of Glycyrrhiza uralensis from three geographical regions by chemical pattern recognition combined with muti-component qualitative and quantitative analysis / 中国中药杂志

Thirty-two batches of cultivated and wild Glycyrrhiza uralensis were obtained from three geographical regions. Comparative study of water characteristic components of G. uralensis from three geographical origins was conducted by PCA,OPLS-DA chemical pattern recognition combined with LC-TOF/MS and muti-component analysis. The similarity of fingerprints of 32 batches of medicinal materials ranged from 0. 903 to 0. 999. Patterns recognition could be used to distinguish cultivated G. uralensis in Gansu and Xinjiang areas from cultivated and wild plants in Inner Mongolia. Then a total of thirty-one common constituents were identified by LC-TOF/MS analysis coupled with standard compounds information. The contents of four flavonoid glycosides and five saponins were determinated by HPLC and compared using One-way ANOVA. The results showed that there was no significant difference in the contents of 5 triterpenoid saponins among the three regions,but the contents of 4 flavonoid saponins showed the trend of Inner Mongolia >Gansu≈Xinjiang( P<0. 05). In the same Inner Mongolia region,the contents of 4 flavonoid glycosides and 5 triterpenoid saponins in wild plant was significantly higher than that in cultivated plants( P<0. 01). In addition,the contents of liquiritin,isoliquiritin,licorice-saponin A_3,22β-acetoxyl-glycyrrhizic acid and uralsaponin B in Gansu and Xinjiang were obviously lower than those in Inner Mongolia,but the contents of glycyrrhizic acid,the main component of G. uralensis,were not different in the three geographical regions. In Inner Mongolia,the contents of liquiritin,isoliquiritin,licorice-saponin A_3,licorice-saponin G_2 and glycyrrhizic acid in wild plants were significantly higher than those in cultivated plants. In conclusion,qualitative/quantitative analysis of multi-index components combined with pattern recognition could effectively evaluate the quality of cultivated and wild licorice in different regions. It was helpful for us to understand the reality of licorice in different regions,and provided scientific basis for the development and comprehensive utilization of licorice resources.

Design and implementation of Xinshenghua granule traceability system based on data authenticity / 中国中药杂志

The quality of traditional Chinese medicine affects the clinical efficacy and the market research at home or abroad, which is closely related to the standardization of production. At present, Chinese market still exist in drug quality and nonstandard production phenomenon. In addition to the existing GMP production standards, it is still necessary to ensure the authenticity and reliability of traditional Chinese medicine by means of retrospect. Based on the Xinshenghua granule, starting from the authenticity of the whole process of production, this study designs and constructs the traditional Chinese medicine traceability system which is equipped with the Internet platform by using two-dimensional code as a trace tool. By making authentic record of the production process and quality transfer, it is convenient for consumers to know the drug information. The production traceability system provides guarantee for the standardization of traditional Chinese medicine development, in order to obtain the production source，the testing quality, whereabouts and responsibility.

HPLC characteristic fingerprints of Xinshenghua Keli / 中国中药杂志

Xinshenghua Keli is known as the "preferred prescription of postpartum", with large demand in the field of gynecologic medicine. However, the quality of the preparation is uneven in the market, so its clinical efficacy cannot be guaranteed. In order to improve and establish its quality control standard, high performance liquid chromatography (HPLC) was used to establish the fingerprint of Xinshenghua Keli. The detection was performed on Agilent 5 HC-C₁₈ (2) column(4.6 mm×250 mm, 5 microns) with methanol-0.1% formic acid solution as mobile phase for gradient elution, at a flow rate of 1 mL·min⁻¹ with column temperature of 25 °C. The injection volume was 10 μL and detection wavelength was set at the maximum value between 210.0 nm and 400.0 nm by Photo-Diode Array (PDA) detector. The fingerprint of 12 batches of high-quality Xinshenghua Keli was established and 43 common peaks were identified. The similarities of crowned products, 10 batches of ordinary ones made by Jiangsu Rongyu Pharmaceutical and 10 batches produced by different manufacturers were evaluated. The composition identification and source analysis for the common peaks were performed by comparing the retention time of herbal medicines and ultraviolet absorption spectrum, along with high performance liquid chromatography-mass spectrometry (HPLC-MS) technology. The established fingerprint of Xinshenghua Keli, has proven to have good precision, stability and repeatability through the methodology validation, so it can be used to comprehensively evaluate the quality of Xinshenghua Keli.

Multi-index determination and optimization of liquirtin separated from polyamide resin / 中国中药杂志

To optimize the separation process of liquirtin from glycyrrhiz by static, dynamic adsorption and desorption experiments on polyamide resin, with liquirtin, isoliquiritin and glycyrrhizic acid as the study index. The optimum process conditions were that the pH of solution was regulated to be 7.0, the concentration of liquirtin was 1.296 g x L(-1), the volume of loading buffer was 3 BV. After absorption, efforts shall be made to elute resin with water, 10%, 20%, 30% ethanol (3 BV for each), collect 20% ethanol eluted fraction, and recover solvents. The results showed lower contents of such impurities as isoliquiritin and isoliquiritin in extracts sepaprated under this process conditions, as well as an increase in purity of liquirtin from 4.86% to 88.5%. The method was simple and feasible, it could obtain a higher purity in extracts from liquirtin and provide basis for industrialized separation and preparation of liquirtin.