AbstractThe ripened seeds of Strychnos nux‐vomica L. have been extensively used as herbal medicines in Asian countries. Dihydroindole‐type alkaloids are not only the active constituents but also the toxicants in Strychnos. However, the simultaneous determination of these alkaloids in both crude and processed Semen Strychni is still lacking. The present study represents the first quantitation and relative quantitation assay of 12 dihydroindole‐type alkaloids in Strychnos nux‐vomica unprocessed and sand‐processed seeds using high‐performance liquid chromatography coupled with diode array detection and mass spectrometry. The relative concentration of ten alkaloids was calculated by semi‐quantification using the internal standard and their amounts in unprocessed and detoxified Semen Strychni were compared. We report here for the first time the significant increase of the two alkaloids, 19‐N‐methyl‐strychnine, and 2,3‐dimethoxy‐19‐N‐methyl‐strychnine, during the processing of Semen Strychni. Our study provides new insight into the true complexity of seed processing procedure and valuable information for assessing the efficacy and safety for clinical applications of Semen Strychni‐containing drugs.

The ripened seeds of Strychnos nux-vomica L. have been extensively used as herbal medicines in Asian countries. Dihydroindole-type alkaloids are not only the active constituents but also the toxicants in Strychnos. However, the simultaneous determination of these alkaloids in both crude and processed Semen Strychni is still lacking. The present study represents the first quantitation and relative quantitation assay of 12 dihydroindole-type alkaloids in Strychnos nux-vomica unprocessed and sand-processed seeds using high-performance liquid chromatography coupled with diode array detection and mass spectrometry. The relative concentration of ten alkaloids was calculated by semi-quantification using the internal standard and their amounts in unprocessed and detoxified Semen Strychni were compared. We report here for the first time the significant increase of the two alkaloids, 19-N-methyl-strychnine, and 2,3-dimethoxy-19-N-methyl-strychnine, during the processing of Semen Strychni. Our study provides new insight into the true complexity of seed processing procedure and valuable information for assessing the efficacy and safety for clinical applications of Semen Strychni-containing drugs.

Rhizoma et Radix Polygoni Cuspidati (RRPC) is commonly prescribed for the treatment of amenorrhea, arthralgia, jaundice and abscess in traditional Chinese medicine. Previous pharmacological studies have indicated that polyphenols are the main pharmacological active ingredients in RRPC. Meanwhile, the poor bioavailability of polyphenols in RRPC implies that those components are probably metabolized by intestinal bacteria before absorption. However, there is rather limited information about RRPC''s metabolites produced by intestinal bacteria and the intestinal absorbed constituents. In the present study, the metabolites were characterized after the aqueous extract of RRPC was incubated with the crude enzyme of human intestinal bacteria in vitro. The metabolic characteristics of glycosides in RRPC were figured out by comparing the metabolic profiles of emodin-8-O-β-d-glucopyranoside and polydatin between aqueous extract of RRPC and equivalent amounts of these two glycosides. The transitional constituents absorbed into blood were investigated in rats via intraduodental administration and portal vein intubation. A total of 38 prototype components and 43 metabolites were detected and characterized in vivo. The overall results demonstrated that the intestinal bacteria played an important role in the metabolism of RRPC, and the main metabolic pathways were hydrolysis in vitro, glucuronidation and sulfation in vivo.

With a great difference in therapeutic effects of Mahuang(MH, the stems of Ephedra sinica) and Mahuanggen(MHG, the roots of Ephedra sinica), chemical differences between MH and MHG should be investigated. In the present study, gas chromatography-mass spectrometry(GC-MS)-based plant metabolomics was employed to compare volatile oil profiles of MH and MHG. The antioxidant activities of volatile oils from MH and MHG were also compared. 32 differential chemical markers were identified according to the variable importance in the projection(VIP) value of orthogonal partial least squares discriminant analysis(OPLS-DA) and P value of Mann-Whitney test. Among them, chemical markers of tetramethylpyrazine(TMP) and α-terpineol were quantified. Their contents were much higher in most MH samples compared with MHG. The antioxidant assay demonstrated that MH had significantly higher free radical-scavenging activity than MHG. Although MH and MHG derived from the same medicinal plant, there was much difference in their volatile oil profiles. MH samples had significantly higher content of two reported pharmacologically important chemical markers of TMP and α-terpineol, which may account for their different antioxidant activities.

A high performance liquid chromatography-tandem mass spectrometry method was developed to investigate the influence of wine-processing on pharmacokinetics of anthraquinones from rhubarb in hyperlipidemic hamsters.

Therapeutic effects of herbal medicines differ greatly due to the use of different anatomical parts or processing methods in traditional Chinese medicine, and Ephedra sinica (ES) is just a case in point. To better understand different traditional uses of the stems (known as Mahuang, MH) and roots (known as Mahuanggen, MHG) of ES, their therapeutic material basis should be investigated. In this study, ephedrine alkaloids were profiled simultaneously with primary metabolites using GC-MS based metabolomics. Ephedrine (E) has been reported to be the major bioactive constituent in MH for the treatment of asthma. The results showed that compared with MH, MHG contained much lower levels of five ephedrine alkaloids, which may well explain that MHG has not been used as an antiasthmatic. Additionally, these pharmacologically important ephedrine alkaloids exhibited strong positive correlation with five primary metabolites. In conclusion, this study facilitates better understanding of different traditional uses of MH and MHG. (C) 2015 Elsevier B.V. All rights reserved.

Objective To investigate apoptotic effects of berberine, a significant alkaloids component existing in Rhizoma coptidis , and its possible acting mechanism in insulinoma cells. Methods Different concentrations of berberine were used to treat mouse insulinoma (MIN6) cells for various period of time. The viability and apoptosis of the cells were analyzed using methylthiazolyldiphenvl-tetrazolium bromide assay, flow cytometry and enzyme-linked immuno sorbent assay. Changes in the relating pro- and anti-apoptosis proteins were detected by western-blotting. Results The half-maximal inhibitory concentration (IC 50 ) of berberine was 5.7 μmol/L on MIN6 cells viability for 16 h. Berberine caused a 20% reduction ( P <0.05) in cell number after only 4-h incubation; which reached 50% after 24 h ( P <0.01). Berberine treatment for 16 h significantly increased the level of DNA fragmentation. The flow cytometry showed the apoptotic rate increased 2.9- and 4.6-fold after treating with berberine (5 μmol/L) for 8 and 16 h, while 3- and 8.7-fold after 10 μmol/L treatment for 8 and 16 h ( P <0.01). Berberine treatment dramatically elevated the expression ratio of Bax to Bcl-2. Meanwhile, berberine notably increased the apoptosis-inducing factors and cytochrome C transforming from the mitochondria to the cytoplasm. Apoptotic protease-activating factor 1 (Apaf-1) was subsequently activated after cytochrome C release. Furthermore, caspase-3 and poly adenosine diphosphate-ribose polymerase were also activated to trigger apoptosis cascade. Conclusion High concentration (5 and 10 μmol/L) of berberine could induce the apoptosis of MIN6 cells through cytochrome C/Apaf-1/caspase-3 and apoptosis inducing factor (AIF) pathway.

Crude rhubarb subjected to different processing procedures will produce different therapeutic effects that are possibly due to processing-induced variation in chemical composition. In this study, a chemometric approach based on ultra fast liquid chromatography with ion trap/time-of-flight mass spectrometry was established to systematically investigate the chemical variations of rhubarb induced by different processing methods. The approach was validated based on pooled quality-control samples from two perspectives: the individual properties of variables and the bulk properties of samples. Orthogonal partial least squares discriminant analysis was introduced to compare the differences between crude and processed rhubarb products. A total of 20 significantly different markers were screened out and unambiguously/tentatively characterized. This research proved that a chemometric method based on ultra fast liquid chromatography with ion trap/time-of-flight mass spectrometry can comprehensively analyze the chemical variation of herbal medicine and provide evidence for a deeper understanding of the pharmacological activities of processed rhubarb products.

As a specific item mentioned in traditional Chinese medicine theory, processing can fulfill different requirements of therapies. Crude and wine-processed rhubarbs are used as drastic and mild laxatives, respectively. In this study, a practical method based on ultra-fast liquid chromatography coupled with diode-array detection and ion trap time-of-flight mass spectrometry was developed to screen and analyze multiple absorbed bioactive components and metabolites in the serum of both normal and acute blood stasis rats after oral administration of crude or wine-processed rhubarbs. A total of 16 compounds, mainly including phase II metabolites, were tentatively identified. Possible explanations for the processing-induced changes in pharmacological effects of traditional Chinese medicines were first explored at serum pharmacochemistry level.

BACKGROUND:Brucine is a widely prescribed glycine antagonist, but a complete understanding of its metabolic pathway is still lacking. The present work represents the first investigation of in vivo metabolism of brucine in rats using LC-ESI-ion trap-TOF-MS.RESULTS:A total of 12 Phase I and five Phase II metabolites were tentatively identified. Brucine can be metabolized by hydrolysis, demethylation and methoxylation, in addition to diverse oxidations in a Phase I manner followed by glucuronidation in Phase II metabolism. Both the renal and biliary routes were observed for the excretion of brucine and its metabolites.CONCLUSION:Our results update the metabolism and disposition data on brucine, which provides basic information for better understanding of the pharmacological and toxicological activities of brucine-containing medicines.

OBJECTIVE To establish the method of thin layer chromatography(TLC) on related substances of Glucosamine Sulfate potassium chloride(GS).METHODS Study on the method of TLC on related substances of GS from the system of expansion agent,the chromogenic method,impurity sources,minimum detection limit,sample loading concentration,specificity and stability.RESULTS TLC could detect the related substances of GS.CONCLUSION The method is simple,sensitive,accurate with specificity and good reproducibility.This method is available for the related substances detection of GS.

Cornu Rhinoceri Asiatici (rhinoceros horn, RH), Cornu Bubali (water buffalo horn, WBH), and Cornu Bovis grunniens (yak horn, YH) are traditional Chinese medicine (TCM), and have been used in China for thousands of years. In this study, ethnopharmacological experiments were used to evaluate and verify the traditional efficacies of horns. Area under curve (AUC) was used to quantify the pharmacological efficacy strength of three horns. Two-dimensional electrophoresis (2-DE) was used to analyze the protein components in horns, as a result, 14 common protein spots in rhinoceros horn, water buffalo horn, and yak horn electrophoresis gels were found by image analysis. Then linear regression analysis was used to establish the correlation between pharmacological efficacies and components in the horns, and five potential active components were selected from the 14 common protein spots. Finally, two protein spots from five were identified by matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS). In this study, a simple method to construct correlation between components and efficacy strength was explored by linear regression analysis, which could be applied to screen potential active components of animal horns.

To establish quality standards for buffalo horn concentrate,the quality information of buffalo horn concentrate including moisture,ash,acid-insoluble ash,water-soluble extract and taurine were evaluated.The content of taurine was determined by HPLC using derivatization method.The results showed that the contents of moisture,total ash,acid-insoluble ash,water-soluble extract and taurine were 1.8%～13.8%,1.0%～6.0%,0.2%～2.2%,3.2%～6.9% and 2.9μg/g～61.5μg/g,respectively.It is indicated that buffalo horn concentrate showed uneven quality,and the methods provided evidences for production and quality control of buffalo horn concentrate.

Cornu Bubali (water buffalo horn, WBH) has been used in Traditional Chinese Medicine (TCM) for thousands of years. In the present study, three peptides with antioxidant properties were purified from aqueous extract of Cornu Bubali (water buffalo horn, WBH) by consecutive chromatographic methods including gel filtration chromatography, ion-exchange chromatography and high performance liquid chromatography. The sequences of the three peptides were identified to be Gln-Tyr-Asp-Gln-Gly-Val (WBH-1, 708Da), Tyr-Glu-Asp-Cys-Thr-Asp-Cys-Gly-Asn (WBH-2, 1018Da) and Ala-Ala-Asp-Asn-Ala-Asn-Glu-Leu-Phe-Pro-Pro-Asn (WBH-3, 1271Da) by matrix assisted laser desorption ionization time-of-flight/time-of-flight mass spectrometry (MALDI-LIFT-TOF/TOF MS). The antioxidant activity of these peptides was tested using 1-diphenyl-2-picrylhydrazyl (DPPH) scavenging assay directly. Methylthiazol tetrazolium (MTT) assay and lactate dehydrogenase (LDH) release assay were also used to evaluate the protection of peptides against hydrogen peroxide (H2O2) induced injury. The results showed that these peptides could reduce the DPPH radical and protect rat cerebral microvascular endothelial cells (rCMECs) against H2O2-induced injury, thus demonstrating that these peptides had antioxidant activity. These results suggest that WBH-1, WBH-2 and WBH-3, isolated from the aqueous extract of water buffalo horn are natural antioxidants and may contribute to the efficacy of WBH.

Cornu Bubali (water buffalo horn, WBH) is an animal-derived product which is widely used in Traditional Chinese Medicine (TCM) for dispelling heat, relieving convulsions and cooling blood. The purpose of this study was to investigate the antipyretic activity of WBH aqueous extract and its potential mechanism. Two hyperthermia models, yeast-induced (infectious) and skimmed milk-induced (noninfectious) hyperthermia were employed to evaluate the antipyretic effect and the results showed that rectal temperature of hyperthermia animals was decreased significantly after oral administration of WBH extract. The production of tumor necrosis factor-alpha (TNF-alpha)-induced prostaglandin E(2) (PGE(2)) in rat cerebral microvascular endothelial cells (rCMECs) was inhibited by WBH extract in the concentrations of 10 microg/ml and 100 microg/ml. The WBH extract protected rCMECs survival from hydrogen peroxide (H(2)O(2))-induced toxicity and inhibited the H(2)O(2)-induced leakage of lactate dehydrogenase (LDH) enzyme release at a dose ranging from 5 microg/ml to 100 microg/ml. It could also increase the superoxide dismutase (SOD) and catalase (CAT) enzyme activities. The results suggest that Cornu Bubali exhibits antipyretic activity on both infectious and noninfectious hyperthermia. The antipyretic activity of WBH may be due to the effects on enhancing antioxidation enzyme activities, decreasing PGE(2) production, and protecting the rCMECs against H(2)O(2)-induced injury.

Objective:To study the antitumor activity and immunity enhancement of polysaccharide from Momordica charantia(MCP) and its potential mechanism.Methods:The model of S180-bearing and H22-bearing tumor mice was built to study the antitumor activity of MCP in vivo.Besides,the stimulation effect of MCP on normal mice lymphocyte proliferation was assayed by MTT method,the phagocytic ability of macrophage RAW264.7 was evaluated with neutral red dye phagocytosis and the stimulation effect of MCP on NO production by macrophage RAW264.7 was measured by Griess reagents.Results:MCP could inhibit the growth of S180 and H22 tumor in vivo and increase the spleen gland index and thymus gland index of tumor-bearing mice.Furthermore,pharmacological experiments showed that MCP can promote normal mice lymphocyte proliferation and the ability of macrophage RAW264.7 in phagocytizing neutral red,and raising the level of NO production.Conclusion:The immune-stimulating effect,such as promoting lymphocyte proliferation and activating macrophage,may result in the antitumor activity of MCP.

Two new bibenzyl derivatives, named dengraols A 1 and B 2, are isolated from stems of Dendrobium gratiosissimum Rchb. (Orchidaceae), together with seven known compounds: 3,5,4'-trihydroxybibenzyl 3, 3,4'-dihydroxy- 5-methoxybibenzyl 4, 3, 4 -dihydroxy-5, 4' - dimethoxy bibenzyl 5, 3, 4' - dihydroxy - 4, 5, 3'-trimethoxybibenzyl (moscatilin) 6 and 5, 4'- dihydroxy -3, 3'-dimethoxybibenzyl (gigantol) 7, 5, 3', 4'-trihydroxy - 3- methoxy-bibenzyl (tristin) 8 and 5, 3'- dihydroxy- 3-methoxybibenzyl (batatasin III) 9. Among the isolated compounds, dengraols A 1 and B 2, moscatilin 6 and gigantol 7 showed inhibitory activity of proliferation on HL-60 cells with IC 50 values at 2.1, 6.4, 0.082 and 10.6 (μM, respectively.

A new bibenzyls compound, Isomoniliformine A, 1-{(7-hydroxy-5-methoxy-6-(18-(hydroxylmethyl)oxiranyl)-16, 18-dimethoxylphenoxy)-phenyl)-2-(11-methoxyphenyl)ethane, was isolated from stems of Dendrobium gratiosissmum Rchb. f. The structure of this unusual bibenzyl compound was determined by chemical and spectroscopic methods, particularly one- and two-dimensional H-1 and C-13 NMR.

OBJECTIVE:To study the chemical constituents of Dendrobium gratiosissimum.METHOD:The constituents were isolated and purified by silica gel, Sephadex LH-20 colum chromatography, and the structures were identified by physicochemical properties and spectral data.RESULT:Nine compounds were obtained as: 3,5,4'-trihydroxybibenzyl (1), 3,4'-dihydroxy-5-methoxybibenzyl (2), 3,4-dihydroxy-4', 5-dimethoxybibenzyl (3), apigenin (4), p-hydroxybenzaldehyde (5), defuscin (6), n-octacostyl ferulate (7), beta-sitosterol (8) and daucosterol (9).CONCLUSION:All these compounds were obtained from this plant for the first time, and compounds 2, 4 and 5 were isolated firstly from this genus.