UPLC-Q-TOF-MS metabolomic study on improvement of acute myocardial ischemia in rats by Dalbergia cochinchinensis heartwood / 中国中药杂志

This paper aimed to study the effect of Dalbergia cochinchinensis heartwood on plasma endogenous metabolites in rats with ligation of the left anterior descending coronary artery, and to analyze the mechanism of D. cochinchinensis heartwood in improving acute myocardial ischemic injury. The stability and consistency of the components in the D. cochinchinensis heartwood were verified by the establishment of fingerprint, and 30 male SD rats were randomly divided into a sham group, a model group, and a D. cochinchinensis heartwood(6 g·kg~(-1)) group, with 10 rats in each group. The sham group only opened the chest without ligation, while the other groups established the model of ligation. Ten days after administration, the hearts were taken for hematoxylin-eosin(HE) staining, and the content of heart injury indexes in the plasma creatine kinase isoenzyme(CK-MB) and lactate dehydrogenase(LDH), energy metabolism-related index glucose(Glu) content, and vascular endothelial function index nitric oxide(NO) was determined. The endogenous metabolites were detected by ultra-high-performance liquid chromatography-time-of-flight-mass spectrometry(UPLC-Q-TOF-MS). The results showed that the D. cochinchinensis heartwood reduced the content of CK-MB and LDH in the plasma of rats to relieve myocardial injury, reduced the content of Glu in the plasma, improved myocardial energy metabolism, increased the content of NO, cured the vascular endothelial injury, and promoted vasodilation. D. cochinchinensis heartwood improved the increase of intercellular space, myocardial inflammatory cell infiltration, and myofilament rupture caused by ligation of the left anterior descending coronary artery. The metabolomic study showed that the content of 26 metabolites in the plasma of rats in the model group increased significantly, while the content of 27 metabolites decreased significantly. Twenty metabolites were significantly adjusted after the administration of D. cochinchinensis heartwood. D. cochinchinensis heartwood can significantly adjust the metabolic abnormality in rats with ligation of the left anterior descending coronary artery, and its mechanism may be related to the regulation of cardiac energy metabolism, NO production, and inflammation. The results provide a corresponding basis for further explaining the effect of D. cochinchinensis on the acute myocardial injury.

Novel SNP markers on ginsenosides biosynthesis functional gene for authentication of ginseng herbs and commercial products / 中国天然药物

Panax ginseng and Panax quinquefolius have similar bioactive components and morphological characteristics, but they are known to have different medicinal values, high-sensitive and accurate method is expected to identify the sources of ginseng products and evaluate the quality, but with a huge challenge. Our established UHPLC-TOF/MS method coupled with orthogonal partial least squares discriminant analysis (OPLS-DA) model based on 18 ginsenosides was applied to discriminate the sources of raw medicinal materials in ginseng products, and nested PCR strategy was used to discover 6 novel single nucleotide polymorphism (SNP) sites in functional dammarenediol synthase (DS) gene for genetic authentication of P. ginseng and P. quinquefolius for the first time. OPLS-DA model could identify the sources of raw ginseng materials are real or not. SNP markers were applied to identify ginseng fresh samples as well as commercial products, and proved to be successful. This established molecular method can tell exact source information of adulterants, and it was highly sensitive and specific even when total DNA amount was only 0.1 ng and the adulteration was as low as 1%. Therefore, this study made an attempt at the exploration of new type SNP marker for variety authentication and function regulation at the same time, and the combination of chemical and molecular discrimination methods provided the comprehensive evaluation and authentication for the sources of ginseng herbs and products.

Discovery of differential sequences for improving breeding and yield of cultivated Ophiocordyceps sinensis through ITS sequencing and phylogenetic analysis / 中国天然药物

To accelerate the breeding process of cultivated Ophiocordyceps sinensis and increase its yield, it is important to identify molecular fingerprint of dominant O. sinensis. In the present study, we collected 3 batches of industrially cultivated O. sinensis product with higher yield than the others and compared their internal transcribed spacer (ITS) sequences with the wild and the reported. The ITS sequence was obtained by bidirectional sequencing and analyzed with molecular systematics as a DNA barcode for rapid and accurate identification of wild and cultivated O. sinensis collected. The ITS sequences of O. sinensis with detailed collection loci on NCBI were downloaded to construct a phylogenetic tree together with the sequences obtained from the present study by using neighbor-joining method based on their evolution relationship. The information on collection loci was analyzed with ArcGIS 10.2 to demonstrate the geographic distribution of these samples and thus to determine the origin of the dominant samples. The results showed that all wild and cultivated samples were identified as O. sinensis and all sequences were divided into seven phylogenetic groups in the tree. Those groups were precisely distributed on the map and the process of their system evolution was clearly presented. The three cultivated samples were clustered into two dominant groups, showing the correlation between the industrially cultivated samples and the dominant wild samples, which can provide references for its optimized breeding in the future.

Discovery of differential sequences for improving breeding and yield of cultivated Ophiocordyceps sinensis through ITS sequencing and phylogenetic analysis / 中国天然药物

To accelerate the breeding process of cultivated Ophiocordyceps sinensis and increase its yield, it is important to identify molecular fingerprint of dominant O. sinensis. In the present study, we collected 3 batches of industrially cultivated O. sinensis product with higher yield than the others and compared their internal transcribed spacer (ITS) sequences with the wild and the reported. The ITS sequence was obtained by bidirectional sequencing and analyzed with molecular systematics as a DNA barcode for rapid and accurate identification of wild and cultivated O. sinensis collected. The ITS sequences of O. sinensis with detailed collection loci on NCBI were downloaded to construct a phylogenetic tree together with the sequences obtained from the present study by using neighbor-joining method based on their evolution relationship. The information on collection loci was analyzed with ArcGIS 10.2 to demonstrate the geographic distribution of these samples and thus to determine the origin of the dominant samples. The results showed that all wild and cultivated samples were identified as O. sinensis and all sequences were divided into seven phylogenetic groups in the tree. Those groups were precisely distributed on the map and the process of their system evolution was clearly presented. The three cultivated samples were clustered into two dominant groups, showing the correlation between the industrially cultivated samples and the dominant wild samples, which can provide references for its optimized breeding in the future.

Simultaneous Determination of Five Active Ingredients in Xialiqi Capsule by UPLC / 中国药房

OBJECTIVE:To establish a method for simultaneous determination of calycosin-7-glucoside,specnuezhenide,ber-berine hydrochloride,palmatine hydrochloride and rosmarinic acid in Xialiqi capsule. METHODS:UPLC was performed on the col-umn of Shield RP18 with mobile phase of nitrile-0.2%formic acid(gradient elution)at a flow rate of 0.2 ml/min,the column tem-perature was 35 ℃,the detection wavelength was 240 nm,and the injection volume was 1 μl. RESULTS:The linear range was 0.619-39.6 μg/ml for calycosin-7-glucoside(r=0.999 9),1.181-115.6 μg/ml for specnuezhenide(r=0.999 9),0.89-57.2 μg/ml for berberine hydrochloride(r=0.999 9),1.16-74 μg/ml for palmatine hydrochloride(r=0.999 9)and 2.47-157.8 μg/ml for rosmarinic acid(r=0.999 9);RSDs of precision,stability and reproducibility tests were lower than 3%,recoveries were 97.98%-102.44%(RSD=1.49%,n=9),97.37%-100.96%(RSD=1.13%,n=9),98.53%-102.30%(RSD=1.40%,n=9),97.06%-102.21%(RSD=1.46%,n=9)and 100.10%-102.25%(RSD=0.68%,n=9),respectively. CONCLUSIONS:The method is simple,rapid and reproducible,and can be used for the quality control of Xialiqi capsule.

A new C21 steroidal saponins from Periplocae Cortex / 中国中药杂志

To study the chemical constituents of Periplocae Cortex, the separation and purification of 70% alcohol extract were carried out by column chromatographies on AB-8 macroporous resin, silica gel and preparative HPLC. The structure of the compounds were identified by NMR and TOF-MS. A new compound was isolated and identified as 21-O-methyl-Δ5-pregnene-3β, 14β, 17β, 21-tetraol-20-one-3-O-β-D-oleandropyranosyl(1-->4)-β-D-cymaropyranosyl-(1-->4)-β-D-cymaropyranosyl (1), named as periplocoside P.

Application of Dry Granulation Technology in Production of Lianhua Qingwen Capsules / 中国中医药信息杂志

Objective To solve the agglomeration problem in the former process, dry granulation technology was used to prepare the granules of Lianhua Qingwen Capsules. Methods Complexity and granule yield coefficient were set as inspection indexes. The optimum subsidiary material and its amount were optimized. The parameters of dry granulation technology were optimized by orthogonal test. Then, granule yield, angle of repose, and bulk density were compared with those of wet granulation technology. Results Starch was set as subsidiary material. The optimum technology is roll pressure of 12 MPa, rotation speed of 5 r/min, and feed speed of 10 r/min. The yield of granules prepared by dry granulation technology was significantly higher than that of wet granulation technology. Conclusion The dry granulation technology can effectively improve the agglomeration of the granulation process of Lianhua Qingwen Capsules, and was suitable for granulating process of Lianhua Qingwen Capsules.

Determination of twelve active compounds in Qili Qiangxin capsules by UPLC-MS / 中国中药杂志

In order to establish an UPLC-MS method for determination of twelve active compounds in Qili Qiangxin capsules including astragaloside, calycosin-7-0-glucoside, ginsenoside Rb1, ginsenoside Re, ginsenoside Rd, ginsenoside Rg1, ginsenoside Rf, periplocin, periplocoside H1, hesperidin, narirutin, isoquercitrin, the chromatographic separations were performedon a Phenomenex UPLC Kinetex C18 column (2.1 mm x 100 mm, 2.6 microm) with gradient elution of acetonitrile and 0.1% aqueous formic acidat a flow rate of 0.4 mL x min(-1). The temperature was set as 40 degrees C and injection volume was 5 microL. The monitoring of all analytes was achieved under the negative ionization mode with TOF-MS and TOF-MS/MS method. The twelve analytes showed good linearity (R2 > 0.9990) within the test ranges, the average recoveries were 98.0%-102%, respectively, and the RSD were less than 3.9%, respectively. The established method is simple, rapid, and sensitive, and can be used for quality control of Qili Qiangxin capsules.

Cloning and sequence analysis of phytoene desaturase gene in Pogostmen cablin / 中草药

Objective: In order to elucidate the carotenoid metabolic pathway in Pogostemon cablin, a phytoene desaturase (PDS) of P. cablin (PcPDS1) gene was cloned and analyzed bioinformatically. Methods: The full length PDS gene sequence was retrieved from transcriptomic database of P. cablin, the full length primer was designed for PCR verification, and PcPDS1 gene was analyzed using several bioinformatical softwares. Results: PcPDS1 (NCBI accession, KC854409) contained 1960 bp length of open reading frame (ORF) and encoded 569 amino acids postulated. Additionally, the physical and chemical properties of PcPDS1-coded protein were analyzed by bioinformatical softwares. Phylogenetic analysis showed that PcPDS1 was tightly clustered with PDS genes in Gentiana lutea, Nicotiana tabacum, and Ipomoea batatas, which is consistent to the phylogenetic relationship of species. Conclusion: PcPDS1 is successfully coloned and molecularly characterized.