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   2014| October-December  | Volume 10 | Issue 40  
    Online since September 26, 2014

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Hplc-ESI-MS/MS analysis of the water-soluble extract from Ziziphi spinosae semen and its ameliorating effect of learning and memory performance in mice
Yanqing Zhang, Longdong Qiao, Mengyuan Song, Lijuan Wang, Junbo Xie, Hua Feng
October-December 2014, 10(40):509-516
DOI:10.4103/0973-1296.141777  PMID:25422554
Background: Ziziphi Spinosae Semen (ZSS), the seed of Ziziphus jujuba var. spinosa (Bunge) Hu ex H. F. Chow., is a traditional herb for insomnia and anxiety in eastern Asia. However, few researches have been concerned with its effect on ameliorating memory and learning performance. Objective: To investigate the constituents of ZSS water soluble extract and its ameliorating learning and memory in mice. Materials and Methods: A new high performance liquid chromatography coupled with tandem mass spectrometry (HPLC-ESI-MS/MS) method was developed and validated to determine the main constituents in the extract. The effect of ZSS water soluble extract on memory and learning performance was investigated in mice by Y-maze and passive avoidance test. Results: The extract could significantly decrease the number of errors (NOE), and increase the transfer latency time (TLT) and electrical stimuli time (EST). In addition, spinosin, jujuboside A (JuA) and jujuboside B (JuB) were simultaneously identified and quantified in the extract. Their contents in the extract were as followed: Spinosin (223.51mg/g), JuA (63.76mg/g) and JuB (26.29mg/g). Conclusion: The extract played a promising role in ameliorating memory in mice with alcohol induced memory retrieval disorders, and might help to improve learning capacity to some extent. Spinosin, JuA and JuB were the predominant constituents, which might be mainly responsible for the definite activity.
  18 2,972 131
Effects of Astragaloside IV combined with the active components of Panax notoginseng on oxidative stress injury and nuclear factor-erythroid 2-related factor 2/heme oxygenase-1 signaling pathway after cerebral ischemia-reperfusion in mice
Xiao-Ping Huang, Yong-Yuan Qiu, Bei Wang, Huang Ding, Ying-Hong Tang, Rong Zeng, Chang-Qing Deng
October-December 2014, 10(40):402-409
DOI:10.4103/0973-1296.141765  PMID:25422538
Background: Astragalus and Panax notoginseng are traditional Chinese Medicines used for the treatments of ischemic cerebrovascular disease, being often combined together in China and achieving a good effect. Objective: The objective of this study is to investigate the effects of astragaloside-IV (AST-IV) (the effective component of Astragalus) combined with ginsenoside Rg1, ginsenoside Rb1, notoginsenoside R1 (the effective components of P. notoginseng) on oxidative stress injury after cerebral ischemia-reperfusion in mice, and to explore the mechanisms through nuclear factor-erythroid 2-related factor-2/heme oxygenase-1 (Nrf2/HO-1) signaling pathway. Materials and Methods: C57BL/6 mice were randomly grouped after treated for 3 days, the model of cerebral ischemia-reperfusion injury was established, and the brain tissues were detected. Results: AST-IV combined with ginsenoside Rg1, ginsenoside Rb1, notoginsenoside R1 could increase significantly the survival rate of nerve cell; decrease the contents of malondialdehyde, nitric oxide, increase the activity of superoxide dismutase and the level of glutathione; Nrf2 was down-regulated in the cytoplasm while up-regulated in nucleus, nuclear translocation rate raised as well as HO-1 messenger ribonucleic acid and protein expressions increased. The effects of four active components combination were better than those of the active components alone. Conclusion: Active components of Astragalus and P. notoginseng had the effects against cerebral ischemia-reperfusion injury, which were related to the antioxidative stress after cerebral ischemia-reperfusion. AST-IV combined with ginsenoside Rg1, ginsenoside Rb1, notoginsenoside R1 could strengthen the antagonism effects on ischemia-reperfusion and oxidative stress injury, the mechanism underlying might be associated with jointly activating Nrf2/HO-1 signaling pathway after cerebral ischemia-reperfusion.
  16 2,694 102
Two new cyclopeptides from the co-culture broth of two marine mangrove fungi and their antifungal activity
Song Huang, Weijia Ding, Chunyuan Li, Daniel G Cox
October-December 2014, 10(40):410-414
DOI:10.4103/0973-1296.141781  PMID:25422539
Background: The strategy that co-cultivation two microorganisms in a single confined environment were recently developed to generate new active natural products. In the study, two new cyclic tetrapeptides, cyclo (D-Pro-L-Tyr-L-Pro-L-Tyr) (1) and cyclo (Gly-L-Phe-L-Pro-L-Tyr) (2) were isolated from the co-culture broth of two mangrove fungi Phomopsis sp. K38 and Alternaria sp. E33. Their antifungal activity against Candida albicans, Gaeumannomyces graminis, Rhzioctonia cerealis, Helminthosporium sativum and Fusarium graminearum was evaluated. Materials and Methods: Different column chromatographic techniques with different solvent systems were used to separate the constituents of the n-butyl alcohol extract of the culture broth. The structures of compounds 1 and 2 were identified by analysis of spectroscopic data (one-dimensional, two-dimensional - nuclear magnetic resonance, mass spectrometry) and Marfey's analytic method. Dilution method was used for the evaluation of antifungal activity. Results: Compounds 1 and 2 were identified as cyclo (D-Pro-L-Tyr-L-Pro-L-Tyr) and cyclo (Gly-L-Phe-L-Pro-L-Tyr), respectively. Compounds 1 and 2 showed moderate to high antifungal activities as compared with the positive control. Conclusions: Compounds 1 and 2 are new cyclopeptides with moderate antifungal activity being worthy of consideration for the development and research of antifungal agents.
  16 2,846 91
Preliminary identification of the absorbed bioactive components and metabolites in rat plasma after oral administration of Shaoyao-Gancao decoction by ultra-performance liquid chromatography with electrospray ionization tandem mass spectrometry
Ping Wang, Quan-wei Yin, Ai-hua Zhang, Hui Sun, Xiu-hong Wu, Xi-jun Wang
October-December 2014, 10(40):497-502
DOI:10.4103/0973-1296.141774  PMID:25422552
Background: Shaoyao-Gancao decoction (SGD), a traditional Chinese medicine formula, has been used for the treatment of abdominal pain and dysmenorrhea disease in Asia over long period of time. Its effectiveness has been confi rmed in clinic, but its active constituents remain unclear. Materials and methods: In this paper, a rapid, sensitive and reliable ultra-performance liquid chromatography-electrospray ionization/ quadrupole-time-of-fl ight high-defi nition mass spectrometry (UPLC-ESI-Q-TOF-MS) in positive and negative ion mode were established to characterize the active constituents of SGD in vitro. The analysis was performed on a Waters UPLCTM HSS T3 (2.1 × 100 mm, 1.8 µm) using gradient elution system. Automated MetaboLynxTM technique was employed to screen for the potentially bioactive components in rat plasma after oral administration of SGD. MS/MS fragmentation behavior was proposed for aiding the structural identifi cation of the components. Results: Based on the developed method of fi ngerprint analysis, an injection run of the plasma sample was fi nished in 15.0 min. A total of 12 compounds including 9 prototype components such as gallicacid, albifl orin, liquiritin, pallidifl orin, liquiritigenin, isoLiquiritigenin, formononetin, isolicofl avonol, licoricone, C9H10O3 and 2 metabolites such as liquiritigenin-4'-O-glucuronide, formononetin glucuronide were identifi ed or tentatively characterized. Of note, 3 ingredients were identified from Radix Paeoniae Alba, and 9 were from Radix Glycyrrhizae. Conclusion: The compounds found in dosed plasma could be the effective substances of SGT for treating dysmenorrheal, and may provide important experimental data for further pharmacological and clinical research of SGD. Furthermore, this work has demonstrated that the feasibility of the UPLC-ESI-Q-TOF-MS for rapid and reliable characterization of identifi cation and structural elucidation of the chemical constituents and their metabolites from herbal medicines.
  15 2,827 126
The treatment of Uygur medicine Dracocephalum moldavica L on chronic mountain sickness rat model
Dilinuer Maimaitiyiming, Guangmei Hu, Ainiwaer Aikemu, Shi Wen Hui, Xiangyang Zhang
October-December 2014, 10(40):477-482
DOI:10.4103/0973-1296.141817  PMID:25422549
Aim: Dracocephalum moldavica L, a traditional Uygur medicine, possesses some key cardiac activities. However, till date, no reports are available on the use of D. moldavica against chronic mountain sickness (CMS), which is a medical condition that affects the residents of high altitude. The present study was designed to explore the treatment efficacy of D. moldavica on CMS. Materials and Methods: 80 of the 100 Sprague Dawley rats enrolled were bred in simulated high altitude environment and the remaining 20 rats were kept in the plains. Water and alcohol extracts of D. moldavica were prepared. CMS rat model was prepared, and the rat hearts were removed for histopathological analysis. Blood samples were taken for hematological and biochemical analyses. Rat pulmonary artery pressure was determined to study the treatment efficacy. Results: In the CMS model group, the levels of interleukin-6 (IL-6), C-reactive protein (CRP), and malondialdehyde (MDA) were found to be significantly higher than the control group; while the concentrations of SOD and GSH-Px decreased. D. moldavica could improve these levels, decrease pulmonary artery pressure, and improve the cardiac pathological state. Conclusions: The study results show that IL-6, CRP, MDA, SOD and GSH-Px participate and mediate the formation of CMS and D. moldavica is found to possess noticeable effects on CMS. The present study explored the basics of high altitude sickness and laid the foundation for further progress of Uygur medicines on the treatment of altitude sickness. Further preclinical and clinical studies with more sample size are recommended.
  14 2,157 73
Effects of ginsenosides-Rb 1 on exercise-induced oxidative stress in forced swimming mice
Bo Qi, Lan Zhang, Zhiqun Zhang, Jiangqiong Ouyang, Hui Huang
October-December 2014, 10(40):458-463
DOI:10.4103/0973-1296.141818  PMID:25422546
Background: The fleshy root of Panax ginseng C.A. Meyer (ginseng) is one of the most well-known and valued herbs in traditional Chinese medicine. Ginsenosides are considered mainly responsible for the pharmacological activities of ginseng. The purpose of this study was to investigate the effects of ginsenoside-Rb 1 (G-Rb 1 ) on swimming exercise-induced oxidative stress in male mice. Materials and Methods: A total of 48 animals were randomly divided into four groups, with twelve mice in each group. The first, second and third groups were designed as G-Rb 1 treatment groups, got 25, 50 and 100 mg/kg bodyweight of G-Rb 1 , respectively. The fourth group was designed as the control group, got physiologic saline. The mice were intragastrically administered once daily for 4 weeks. The weight-loaded forced swimming test was conducted on the final day of experimentation. Then the exhaustive swimming time, blood lactate, serum creatine kinase (CK), malondialdehyde (MDA) and antioxidant enzymes in liver of mice were measured. Results: The results showed that G-Rb 1 could prolong the exhaustive swimming time and improve exercise endurance capacity of mice, as well as accelerate the clearance of blood lactate and decrease serum CK activities. Meanwhile, G-Rb 1 could decrease MDA contents and increase superoxide dismutase, catalase, glutathione peroxidase activities in liver of mice. Conclusions: The study suggested that G-Rb 1 possessed protective effects on swimming exercise-induced oxidative stress in mice.
  13 2,645 84
Changes in regional cerebral blood flow with Chaihu-Shugan-San in the treatment of major depression
Juan Qiu, Sui-Yu Hu, Guang-Qing Shi, Su-e Wang
October-December 2014, 10(40):503-508
DOI:10.4103/0973-1296.141775  PMID:25422553
Background: Chaihu-Shugan-San (CHSGS) is a well-known Chinese traditional prescription used for depression. Objective: To observe the regional cerebral blood flow (rCBF) changes in patients with major depression and to investigate rCBF and clinical response to CHSGS. Materials and Methods: A total of 33 unmedicated patients with major depression and 12 healthy comparison subjects underwent single photon emission computed tomography (SPECT) imaging. A total of 33 unmedicated patients with major depression all met the diagnostic criteria of stagnation of liver qi of traditional Chinese medicine and were divided into two groups: CHSGS group (n = 20) and fluoxetine group (n = 13). SPECT imaging was restudied in posttreatment. Results: SPECT detected abnormalities in all (100.0%) patients both in CHSGS group and fluoxetine group. All healthy subjects were normal results. The depressed patients showed rCBF decreased in the multiple regions. The semiquantitative values of bilateral frontal and left temporal lobes both in CHSGS group and fluoxetine group were lower than that in healthy group (P < 0.05). Reexamined SPECT after 8 weeks treatment with CHSGS showed the consistency between the increase in perfusion defects and the improvement of clinical cerebral symptoms. The semiquantitative values increased in posttreatment, when compared with pretreatment (P < 0.05). Conclusion: SPECT represents a sensitive tool to detect the major depressive disorder, which show the rCBF decreased. rCBF perfusion defects can be reversed and clinical symptoms can be improved by CHSGS treatment. CHSGS treatment is effective, well-tolerated, and safe for depression. By semiquantitative analysis, SPECT can objectively detect rCBF changes that is useful for guiding treatment.
  12 2,105 61
Comparison of cytotoxic activities of extracts from Selaginella species
Juan Li, Xiang Lei, Ke-li Chen
October-December 2014, 10(40):529-535
DOI:10.4103/0973-1296.141794  PMID:25422557
Background: Selaginella species are resurrection plants, which are known, possess various molecular bioactivities depending on species, but only a few species have been detailed observe in the advanced research. Objective: The objective of the following study is to compare the chemical profiles of different species of Selaginella and to investigate cytotoxicity and induction of apoptosis activities of some species of Selaginella. Materials and Methods: The high-performance liquid chromatography (HPLC) method was developed for chemical analysis. Ethyl acetate, ethanol and water-soluble extracts from seven Selaginella species were submitted to 3-(4,5-dimenthyl thizol-2-yl)-2,5-diphenyl tetrazolium bromide assay, flow cytometry, deoxyribonucleic acid (DNA) laddering analysis and caspase-3 expression using Bel-7402, HT-29 and HeLa cells. Results: The HPLC analysis revealed two major common peaks, which were identified as amentoflavone and robustaflavone and another three main peaks in their chromatograms. The results showed that S. labordei, Selaginella tamariscina and Selaginella uncinata had relatively stronger activities on Bel-7402 and HeLa cells and Selaginella moellendorfii had moderate antiproliferation activities, but Selaginella remotifolia and Selaginella pulvinata had almost no inhibitory activities. The main active components were in the ethyl acetate extracts which had abundant biflavonoids. The effects of these extracts on cell proliferation and apoptosis in different cells were not the same, they were more apparent on HeLa cells than on HT-29 cells. The assay of DNA laddering analysis and caspase-3 expression further confirmed that inducing cell apoptosis was one of antitumor mechanisms and antitumor activities of Selaginella species were related to apoptosis induced by caspase family. Conclusion: S. labordei, S. tamariscina and S. uncinata would be potential antitumor agents.
  12 2,761 76
Molecular authentication of the traditional medicinal plant Peucedanum praeruptorum and its substitutes and adulterants by dna - barcoding technique
Jing Zhou, Wencai Wang, Mengqi Liu, Zhenwen Liu
October-December 2014, 10(40):385-390
DOI:10.4103/0973-1296.141754  PMID:25422535
Background: Peucedanum praeruptorum L., a traditional Chinese medicine known as Qian-hu, is commonly used for dispelling wind-heat and expectorant and loss of energy. However, due to similar morphological characters and high market demand, there are many substitutes and adulterants of P. praeruptorum. DNA barcoding is an approach to identify species based on sequences from a short, standardized DNA region. Objective: To authenticate P. praeruptorum from its substitutes and adulterants. Materials and Methods: The differential identification of P. praeruptorum and 13 regional substitutes and 23 adulterants was investigated by means of DNA sequence analysis of internal transcribed spacer (ITS) region of the nuclear ribosomal DNA (nrDNA), a bootstrap neighbor-joining (NJ) tree according to Kimura's 2-parameter method was also calculated. Results: The data showed that P. praeruptorum, its substitutes and adulterants could be easily distinguished at the DNA level, while almost all species were well resolved, and successfully identified on the NJ tree. Conclusion: The ITS sequence can be used for the identification of P. praeruptorum and to distinguish it from common substitutes and adulterants.
  11 3,705 312
An improved ultra-performance liquid chromatography-electrospray ionization/quadrupole-time-of-flight high-definition mass spectrometry method for determining ingredients of herbal Fructus corni in blood samples
Changsong Xue, Aihua Zhang, Hui Sun, Ying Han, Di Zou, Yuying Wang, Xiuhong Wu, Xijun Wang
October-December 2014, 10(40):422-429
DOI:10.4103/0973-1296.141796  PMID:25422541
Background: Fructus Corni (FC), a well-known traditional Chinese medicine (TCM), derived from the dry ripe sarcocarp of Cornus offi cinalis Sieb. et Zucc (Cornaceae), has been widely prescribed to treat disease in China for centuries. It has attracted increasingly much attention as one of the most popular and valuable herbal medicine in clinic. However, the systematic analysis of the chemical constituents of FC is diffi cult to determine and remain unclear. Materials and Methods: In this work, a rapid, sensitive, and reliable ultra-performance liquid chromatography-electrospray ionization/ quadrupole-time-of-fl ight high-defi nition mass spectrometry (UPLC-ESI/QTOF/MS) with automated data analysis (MetaboLynx TM ) in negative ion mode were established to characterize the chemical constituents of FC and simultaneously identify components in blood after oral administration of FC, respectively. The analysis was performed on a Waters UPLC TM HSS T 3 (2.1 × 100 mm, 1.8 μm) using gradient elution system. MS/MS fragmentation behaviors were proposed for aiding the structural identifi cation of the components. Results: With optimized conditions, a total of 34 peaks were obtained from FC, 23 of which were tentatively characterized by comparing the retention time and mass spectrometry data and retrieving the reference literatures. Of note, the 25 compounds were identifi ed after oral administration of FC, which might be the potential active components in vivo. Conclusion: The present study demonstrates the potential of UPLC-ESI/QTOF/MS approach for the rapid and reliable characterization of the metabolites of natural products.
  11 2,656 74
Chemical analysis of raw and processed Fructus arctii by high-performance liquid chromatography/diode array detection-electrospray ionization-mass spectrometry
Kunming Qin, Qidi Liu, Hao Cai, Gang Cao, Tulin Lu, Baojia Shen, Yachun Shu, Baochang Cai
October-December 2014, 10(40):541-546
DOI:10.4103/0973-1296.141806  PMID:25422559
Background: In traditional Chinese medicine (TCM), raw and processed herbs are used to treat the different diseases. Fructus Arctii, the dried fruits of Arctium lappa l. (Compositae), is widely used in the TCM. Stir-frying is the most common processing method, which might modify the chemical compositions in Fructus Arctii. Materials and Methods: To test this hypothesis, we focused on analysis and identification of the main chemical constituents in raw and processed Fructus Arctii (PFA) by high-performance liquid chromatography/diode array detection-electrospray ionization-mass spectrometry. Results: The results indicated that there was less arctiin in stir-fried materials than in raw materials. however, there were higher levels of arctigenin in stir-fried materials than in raw materials. Conclusion: We suggest that arctiin reduced significantly following the thermal conversion of arctiin to arctigenin. In conclusion, this finding may shed some light on understanding the differences in the therapeutic values of raw versus PFA in TCM.
  7 2,480 77
Protective effects of methanolic extract form fruits of Lycium ruthenicum Murr on 2,2'-azobis (2-amidinopropane) dihydrochloride-induced oxidative stress in LLC-PK1 cells
Jia-Le Song, Yang Gao, Jianguo Xu
October-December 2014, 10(40):522-528
DOI:10.4103/0973-1296.141790  PMID:25422556
Background: Fruits of Lycium ruthenicum Murr is a health food and also used as a folk to treat heart disease, abnormal menstruation and menopause in Tibetan, China. However; whether L. ruthenicum Murr fruits methanolic extracts (LFME) protect LLC-PK1 porcine renal tubules cells from AAPH-induced oxidative damage has not been investigated. Objective: To investigate the protective effects of L. ruthenicum Murr fruits methanolic extracts (LFME) against 2, 2'- azobis (2-amidinopropane) dihydrochloride (AAPH)-induced oxidative damage in renal proximal tubule LLC-PK1 cells. Materials and Methods: LLC-PK1 cells were co-incubated with AAPH (1mM) and different concentrations of LFMW together for 24 h. Cell viability was determined by MTT assay. Total intercellular reactive oxygen species (ROS) levels and lipid peroxidation were measured using a fluorescent probe 2', 7'-dichlorfluorescein-diacetate (DCFH-DA) and the TBA reactive substance (TBARS) assay, respectively. The endogenous antioxidant enzymes including catalase (CAT), superoxide dismutase (SOD), glutathione peroxidase (GSH-px) and intercellular glutathione (GSH) levels were determined using commercial assay kits according to the manufacturer's instructions. Results: LFME did not show a significant cytotoxic effect and increased the viability of LLC-PK1 cells in a concentration-dependent manner. LFME also decreased the total intercellular levels of ROS, reduced lipid peroxidation and increased the GSH levels as well as the activities of endogenous antioxidant enzymes to protect LLC-PK1 cells against AAPH-induced oxidative damage. Conclusion: The results from the present study indicated that LFME is an effective ROS scavenger to protect LLC-PK1 cells against AAPH-induced oxidative damage through decreasing ROS generation, reducing lipid peroxidation and up-regulation of endogenous GSH levels and antioxidant enzymes.
  6 2,570 54
A rapid UPLC method for simultaneous determination of eleven components in 'Ge-Gen-Qin-Lian' decoction
Rui An, Lisha You, Yizhu Zhang, Xinhong Wang, Yuemin Ma
October-December 2014, 10(40):464-469
DOI:10.4103/0973-1296.141821  PMID:25422547
Background: 'Ge-Gen-Qin-Lian' Decoction derived from 'Shang-Han-Lun' compiled by Zhang Zhongjing. It is widely used in the treatment of acute gastroenteritis, bacillary dysentery, virus diarrhea. This paper describes a sensitive and specific assay for the determination of the 11-marker compounds using ultra performance liquid chromatography (UPLC). Objective: To develop an UPLC method for simultaneous determination of 11 bioactive compounds in 'Ge-Gen-Qin-Lian' preparations. Materials and Methods: The chromatography analysis was performed on an Agilent Proshell 120 EC-C 18 column (4.6 × 50 mm, 2.7 μm) at 30°C with a gradient elution of methanol, 0.5% formic acid and 0.5% ammonium acetate at a flow rate 1.0 ml/min and UV detected at 270 nm. Results: All calibration curves showed good linear regression (r ≥ 0.9993) within tested ranges. Limits of detection (LOD) and limits of quantification (LOQ) fell in the range between 0.0691-1.04 μg/ml and 0.23-3.43 μg/ml, respectively. The mean recovery of each herbal medicine ranged from 96.60 to 102.11%. Conclusion: The method was validated for repeatability, precision, stability, accuracy, and selectivity. The validated method was successfully applied to simultaneous analysis of these active components in 'Ge-Gen-Qin-Lian' decoction.
  6 2,149 71
Cytochrome C oxidase subunit I barcodes provide an efficient tool for Jinqian Baihua She (Bungarus parvus) authentication
Zhi Chao, Jing Liao, Zhenbiao Liang, Suhua Huang, Liang Zhang, Junde Li
October-December 2014, 10(40):449-457
DOI:10.4103/0973-1296.141816  PMID:25422545
Objective: To test the feasibility of DNA barcoding for accurate identification of Jinqian Baihua She and its adulterants. Materials and Methods: Standard cytochrome C oxidase subunit I (COI) gene fragments were sequenced for DNA barcoding of 39 samples from 9 snake species, including Bungarus multicinctus, the officially recognized origin animal by Chinese Pharmacopoeia, and other 8 adulterate species. The aligned sequences, 658 base pairs in length, were analyzed for divergence using the Kimura-2-parameter (K2P) distance model with MEGA5.0. Results: The mean intraspecific K2P distance was 0.0103 and the average interspecific genetic distance was 0.2178 in B. multicinctus, far greater than the minimal interspecific genetic distance of 0.027 recommended for species identification. A neighbor-joining (NJ) tree was constructed, in which each species formed a monophyletic clade with bootstrap supports of 100%. All the data were submitted to Barcode of Life Data system version 3.0 (BOLD, http://www.barcodinglife.org) under the project title "DNA barcoding Bungarus multicinctus and its adulterants". Ten samples of commercially available crude drugs of JBS were identified using the identification engine provided by BOLD. All the samples were clearly identified at the species level, among which five were found to be the adulterants and identified as Dinodon rufozonatum. Conclusion: DNA barcoding using the standard COI gene fragments provides an effective and accurate means for JBS identification and authentication.
  6 3,032 113
Chromatographic fingerprinting analysis of Zhizhu Wan preparation by high-performance liquid chromatography coupled with photodiode array detector
Hui Sun, Xi Chen, Aihua Zhang, Tetsuro Sakurai, Jinzhong Jiang, Xijun Wang
October-December 2014, 10(40):470-476
DOI:10.4103/0973-1296.141819  PMID:25422548
Background: Traditional Chinese medicine (TCM) formula has been used for over 1000 years and most of them contain complicate chemical constituents. Chromatographic fingerprinting has been widely accepted as a crucial method for qualitative and quantitative analyses for TCM. Zhi Zhu Wan (ZZW), a classical Chinese medical formula, has been commonly used for the treatment of gastrointestinal disease, which pose a serious challenge to its quality control. Materials and Methods: In this work, a sensitive and reliable method of high-performance liquid chromatography coupled with photodiode array detector (HPLC-PDA) was developed to control the quality of ZZW for chemical fingerprint analysis and quantitative analysis of four major bioactive constituents, including hesperidin, naringin, neohesperidin, and atractylenolide I. The chromatographic separation was performed on a Waters Symmetry C18 column (4.6 mm × 250 mm, 5 μm particle size), with an aqueous 0.095% phosphate acid and acetonitrile mobile phase gradient. Results: Optimization of other experimental conditions was validated with satisfactory accuracy, precision, repeatability, and recovery. In quantitative analysis, the four components showed good regression (R > 0.9994) within test ranges, and the recovery method ranged from 99.32% to 100.630%. HPLC fingerprints of the ZZW samples were compared by performing similarity analysis. Conclusion: The results indicated that the newly developed HPLC-PDA fingerprint method would be suitable for quality control of ZZW.
  5 2,000 60
The effects of the water-extraction of Astragali Radix and Lycopi herba on the Pathway of TGF-smads-UPP in a rat model of Diabetic Nephropathy
Xiao Fu, Bing Song, Guo-wei Tian, Jing-Lin Li
October-December 2014, 10(40):491-496
DOI:10.4103/0973-1296.141773  PMID:25422551
Background: Astragali Radix and Lycopi Herba were widely used in clinical practice for treating the diabetic nephropathy (DN), but their therapeutic mechanisms were not clear. Objective: To observe the effects of the water-extraction of Astragali Radix and Lycopi Herba on the signaling pathway of TGF-Smads-UPP in streptozotocin (STZ)-induced DN. Materials and Methods: Sprague-Dawley (SD) rats were randomly divided into the normal control (NC) group and the model group. The NC group was fed with a standard diet and the other five diabetic groups received a high-fat diet. After 4 weeks, five diabetic groups were treated with STZ (30mg/kg i.p.). The NC group rats were treated with citrate buffer. Tail random blood glucose (RBG) was measured 72h later using a strip-operated blood glucose sensor and monitored every 2 weeks until drug intervention. Rats with RBG levels less than 16.7mmol/L were excluded from the diabetic groups. At the end of 4 weeks after STZ injection, 24h microalbuminuria was collected and detected. The microalbuminuria was measured by radioimmunoassay (RIA). The blood glucose was tested using a blood glucose meter. The kidney was dissected from each SD rat. Proteins and mRNA of TGF-β1, Smads and Smurf were tested by western-blot and real-time PCR analysis, and 26S proteasome activity was measured by an ELISA kit. Results: The water-extraction of Astragali Radix and Lycopi Herba significantly lowered fasting glucose and urine albumin in diabetic rats through inhibition of TGF-β1 mRNA and protein expression in the STZ-induced diabetic rats, and regulation of the Smad3, Smad7, Smurf1, Smurf2 mRNA and protein expression, as well as elevated 26S proteasome activity to play control effect in DN. Conclusion: 0.9 g/ml water-extraction of Astragali Radix and Lycopi Herba group has significant therapeutic effects on the STZ-induced diabetic rats, and this regulation depends on TGF-Smads-UPP signaling pathway.
  5 2,210 74
Classification and quantification analysis of peach kernel from different origins with near-infrared diffuse reflection spectroscopy
Wei Liu, Zhen-Zhong Wang, Jian-Ping Qing, Hong-Juan Li, Wei Xiao
October-December 2014, 10(40):441-448
DOI:10.4103/0973-1296.141814  PMID:25422544
Background: Peach kernels which contain kinds of fatty acids play an important role in the regulation of a variety of physiological and biological functions. Objective: To establish an innovative and rapid diffuse reflectance near-infrared spectroscopy (DR-NIR) analysis method along with chemometric techniques for the qualitative and quantitative determination of a peach kernel. Materials and Methods: Peach kernel samples from nine different origins were analyzed with high-performance liquid chromatography (HPLC) as a reference method. DR-NIR is in the spectral range 1100-2300 nm. Principal component analysis (PCA) and partial least squares regression (PLSR) algorithm were applied to obtain prediction models, The Savitzky-Golay derivative and first derivative were adopted for the spectral pre-processing, PCA was applied to classify the varieties of those samples. For the quantitative calibration, the models of linoleic and oleinic acids were established with the PLSR algorithm and the optimal principal component (PC) numbers were selected with leave-one-out (LOO) cross-validation. The established models were evaluated with the root mean square error of deviation (RMSED) and corresponding correlation coefficients (R2 ). Results: The PCA results of DR-NIR spectra yield clear classification of the two varieties of peach kernel. PLSR had a better predictive ability. The correlation coefficients of the two calibration models were above 0.99, and the RMSED of linoleic and oleinic acids were 1.266% and 1.412%, respectively. Conclusion: The DR-NIR combined with PCA and PLSR algorithm could be used efficiently to identify and quantify peach kernels and also help to solve variety problem.
  4 2,345 68
Simultaneous quantification of eleven bioactive components of male flowers of Eucommia ulmoides oliver by HPLC and their quality evaluation by chemical fingerprint analysis with hierarchical clustering analysis
Yanxia Ding, Deqiang Dou, Yangjing Guo, Qin Li
October-December 2014, 10(40):435-440
DOI:10.4103/0973-1296.141813  PMID:25422543
Background: Eucommia ulmoides Oliv (EU), a dioecious perennial angiosperm, is one of the oldest tonics in Chinese traditional medicine. The tea of male flowers of EU has been become popularities and seen as aspirational health care tea in China. There were no enough marks and effective method to control the quality of male flowers of EU. Objective: A simple and efficient HPLC method was developed for the simultaneous determination of 11 bioactive compounds (4 iridoids, 1 phenylpropanoid, 6 flavonoids). HPLC chromatographic fingerprint and hierarchical cluster analysis were used to evaluate and classify the samples of male flowers of EU which came from different locations in China. Materials and Methods: Samples were separated on a Thermal hypersil gold column (250 mm × 4.6 mm, 5 μm) and detected by an ultraviolet detector. The UV wavelength was set at 206, 236, and 206 nm. Mobile phase consisted of methanol (B) and phosphoric acid-water (0.5%) (C) using a gradient elution. Analytes were performed at 25°C with a flow rate of 1.0 mL/min. Results: In quantitative analysis, the eleven components showed good regression (r 2 > 0.9996) within linear ranges, and their recoveries were in the range of 98.65-102.31%. In the chromatographic fingerprint, 16 peaks were selected as the characteristic peaks to assess the similarities of different samples. Hierarchical cluster analysis (HCA) was also applied to differentiate the samples based on the area of all the common peaks. The samples which had higher similarity in HPLC fingerprint were classified as a cluster. Conclusion: This study will provide methodological reference for the quality control and sample classification of male flowers of E. ulmoides.
  3 2,565 94
Accumulation and trends in distribution of three triterpenoids in various parts of Achyranthes coynei determined using RP-UFLC analysis
Vinayak Upadhya, Gireesh M Ankad, Sandeep R Pai, Harsha V Hegde, Sanjiva D Kholkute
October-December 2014, 10(40):398-401
DOI:10.4103/0973-1296.141761  PMID:25422537
Background: Achyranthes coynei Sant. (Family: Amaranthaceae) is a rare endemic medicinal plant used by local traditional practitioners to treat various diseases. The plant has been reported for promising antibacterial and antioxidant activities. However, the plant is not explored for its phytocompounds, especially triterpenoids. Objective: To study the accumulation and trends in distribution of triterpenoids: betulinic, oleanolic and ursolic acids (BA, OA and UA) in leaf, stem, root and inflorescence of A. coynei. Materials and Methods: Extraction was achieved using refluxing and reversed phase-ultra flow liquid chromatographic (RP-UFLC) technique was employed for determination. Separation of triterpenoids was achieved on a Hibar 250-4.6 mm, 5 μ, Lichrospher 100, C18e column using methanol and water (90:10) as mobile phase (pH adjusted to 5.0 using GAA) in an isocratic mode. Results: Oleanolic acid was higher in leaf (0.172 ± 0.009%) followed by stem (0.035 ± 0.002%) and root (0.028 ± 0.001%). Ursolic acid was accounted to be highest in the inflorescence (0.099 ± 0.005%). The contents of BA and UA were lower than OA in leaf and stem while it was remained undetected in roots of A. coynei. Conclusion: The triterpenoids: BA, OA and UA were detected, quantified and reported for the first time from A. coynei. In the present study leaves were found to be the major source of BA and OA, whereas inflorescence was for UA.
  2 2,335 70
Application of microscopy technique and high performance liquid chromatography for quality assessment of Polygonum multiflorum Thunb. (Heshouwu)
Li Liang, Zhongzhen Zhao, Tingguo Kang
October-December 2014, 10(40):415-421
DOI:10.4103/0973-1296.141788  PMID:25422540
Background: The technique of microscopy has been applied for identification of Chinese materia medica (CMM) since decades. However, very few scientific publications report the combination of conventional microscopy and high performance liquid chromatography (HPLC) techniques for further application to quality assessment of CMM. Objective: The objective of this study is to analyze the quality of the dried root tuber of Polygonum multiflorum Thunb. (Heshouwu) and to establish the relationships between 2,3,5,4'- tetrahydroxystilbene-2-O-β-glucoside, combined anthraquinone (CAQ) and quantity of clusters of calcium oxalate. Materials and Methods: In this study, microscopy and HPLC techniques were applied to assess the quality of P. multiflorum Thunb., and SPSS software was used to establish the relationship between microscopic characteristics and chemical components. Results: The results showed close and direct correlations between the quantity of clusters of calcium oxalate in P. multiflorum Thunb. and the contents of 2,3,5,4'- tetrahydroxystilbene-2-O-β-glucoside and CAQ. From these results, it can be deduced that Polygoni Multiflori Radix with a higher quantity of clusters of calcium oxalate should be of better quality. Conclusion: The established method can be helpful for evaluating the quality of CMM based upon the identification and quantitation of chemical and ergastic substance of cells.
  2 2,440 60
Er-Xian Decoction, a traditional Chinese herbal formula, intervening early in hypothalamic-pituitary axis of male rats with delayed puberty
Zheng Zhu, LiHong Li, Xin Jin, JianWei Fang, DongFang Zhang
October-December 2014, 10(40):517-521
DOI:10.4103/0973-1296.141780  PMID:25422555
Background: Er-Xian Decoction (EXD) is one of the traditional Chinese medicine (TCM) with unique effect on osteoporosis, menopausal syndrome and delayed puberty in China for many years. Objective: We aim to evaluate the potential activity of starting hypothalamic-pituitary-testicular (HPT) axis of male rats with delayed puberty. Materials and Methods : Delayed puberty model of male Sprague-Dawley (SD) rats were established with soy isoflavones (90 mg·kg -1 ) and were treated by EXD extract at doses of 5, 10 g·kg -1 or Testosterone undecanoate (TU) for 8 weeks. Body weight, body length, testis weight, T, E 2 and luteinizing hormone (LH) in serum, gonadotropin releasing hormone (GnRH) in hypothalamus, follicle stimulating hormone (FSH) and LH in pituitary gland were determined by ELISA. Immunohistochemistry was used to detect LH in pituitary gland . Results: Soy isoflavones could significantly decrease body weight, body length, testicular organ coefficient T in serum, GnRH in hypothalamus, FSH and LH in pituitary gland. Both of EXD and TU could improve the condition. E 2 and LH in serum of all groups were non-significance of difference (P > 0.05). The immunohistochemical results were well consistent with LH in pituitary gland. Conclusion: The results of the present research indicate that EXD extract is effective to start the HPT axis in puberty and can significantly improve sexual developmental inhibition caused by soy isoflavones.
  2 2,287 71
A novel approach for the efficient extraction of silybin from milk thistle fruits
Caihong Tan, Xianrong Xu, Yaqi Shang, Xianli Fu, Guohua Xia, Huan Yang
October-December 2014, 10(40):536-540
DOI:10.4103/0973-1296.141799  PMID:25422558
Background: Milk Thistle fruit is an important herb popularly consumed worldwide for a very long time. Silybin is the main bioactive constituent of the herb, and it has been approved by US Food and Drug Administration (FDA) as a medicine to treat liver diseases. Presently, using conventional technology, the meal of Milk Thistle fruit is used as the raw material to extract silybin. Objective: To investigate the necessity of detaching husk from kernel of the herb and also to propose a novel approach to enhance the extraction technology in pharmaceutical practices. Materials and Methods: The husk of Milk Thistle fruit was detached from the kernel of the herb using an automatic huller specially designed for this application. The husk and the meal of Milk Thistle fruit was subsequently refluxed, separately, with production rate of silybin as index for comparison of their extraction effect. Results: The highest production rate was achieved under optimized condition. The husk was extracted 2 times (3 hrs each) using ethyl acetate, and the ratio of solvent to raw material was 8:1. The extract was allowed to be crystallized out. Conclusion: The separation of kernel from the husk of Milk Thistle fruit and using only the husk as raw material can largely enhance the extraction of silybin.
  2 2,318 80
Simultaneous quantification and splenocyte-proliferating activities of nucleosides and bases in Cervi cornu Pantotrichum
Ying Zong, Yu Wang, Hang Li, Na Li, Hui Zhang, Jiaming Sun, Xiaohui Niu, Xiaochen Gao
October-December 2014, 10(40):391-397
DOI:10.4103/0973-1296.141757  PMID:25422536
Background: Cervi Cornu Pantotrichum has been a well known traditional Chinese medicine, which is young horn of Cervus Nippon Temminck (Hualurong: HLR). At present, the methods used for the quality control of Cervi Cornu Pantotrichum show low specificity. Objective: To describe a holistic method based on chemical characteristics and splenocyte-proliferating activities to evaluate the quality of HLR. Materials and Methods: The nucleosides and bases from HLR were identified by high performance liquid chromatography electrospray ionization mass spectrometry (HPLC-ESI-MS), and six of them were chosen to be used for simultaneous HPLC quantification according to the results of proliferation of mouse splenocytes in vitro. Results: In this study, eight nucleosides and bases have been identified. In addition, uracil, hypoxanthine, uridine, inosine, guanosine, and adenosine were chosen to be used for simultaneous HPLC quantification. Simultaneous quantification of these six substances was performed on ten groups of HLR under the condition of a TIANHE Kromasil C 18 column (5 μm, 4.6 mm × 250 mm i.d.) and a gradient elution of water and acetonitrile. Of the ten groups, HLR displayed the highest total nucleoside contents (TNC, sum of adenosine and uracil, 0.412 mg/g) with the strongest splenocyte-proliferating activities. Conclusion: These results suggest that TNC (such as particularly highly contained adenosine and uracil) in HLR has a certain correlation with the activity of splenocyte-proliferating, and it may be used as a quality control for HLR. This comprehensive method could be applied to other traditional Chinese medicines to ameliorate their quality control.
  1 3,110 298
Exploring processing adjuvants' influence on traditional Chinese medicine compatibility of Aconiti Radix Cocta and Pinelliae rhizome using rapid resolution liquid chromatography-quadrupole time-of-flight mass spectrometry
Yuming Wang, Yubo Li, Xiuxiu Zhang, Yanyan Xu, Hui Wang, Yanjun Zhang
October-December 2014, 10(40):483-490
DOI:10.4103/0973-1296.141771  PMID:25422550
Background: It is known that when crude Pinelliae rhizome and Pinelliae rhizoma preparatum are combined with Aconiti Radix Cocta respectively, the toxicity of the combination varies. However, the component's transformation between different compatibility have remained unclear. Objective: In this paper, a novel approach using rapid resolution liquid chromatography-quadrupole time-of-flight mass spectrometry (RRLC-Q-TOF-MS) coupled with multivariate statistical analysis was established for exploring the influence of processing adjuvants (PAs) on the compatibility of Aconiti Radix Cocta and Pinelliae rhizome. Materials and Methods: In order to obtain information about the representative markers between different groups, an exhaustive study of different protocols based on adding or removing different PAs step by step was carried out and the influence of PAs on compatibility was investigated. Results: It was found that lime can facilitate diester diterpenoid alkaloids with high toxicity in Aconiti Radix Cocta to be converted into low-toxic or non-toxic derivatives. Glycyrrhizae Radix et Rhizoma had no remarkable effect on the process. Conclusion: The established method in this study will be of great significance to process research mechanism and study on traditional Chinese Medicine compatibility and clinical application.
  1 2,263 66
Mycelial fermentation characteristics and antiproliferative activity of Phellinus vaninii Ljup
Wei Hu, Shuai Liu, Yuexin Zhang, Deng Xun, Chunping Xu
October-December 2014, 10(40):430-434
DOI:10.4103/0973-1296.141812  PMID:25422542
Background: The mycelial fermentation of higher fungi were investigated to posses various bioactivities. Materials and Methods: The mycelial growth and pellet morphology in a 5-L bioreactor were investigated. The mycelial broth containing biomass and extracellular products harvested from the fermentor was tested for antiproliferative activity of colon cancer LoVo cells using 3-(4,5-Dimethylthiazol-2-yl)-2,5-Diphenyltetrazolium Bromide assay. Results: The maximum mycelial concentration in a 5-L bioreactor was 12.5 g/L after 8 days cultivation. Further investigation in the mycelial pellets during the fermentation period revealed that the mean diameter of the pellet morphology was positively correlated with mycelial biomass (R2 = 0.82, P < 0.05) and broth viscosity (R2 = 0.90, P < 0.01), significantly. The ethyl acetate extract showed the most significant effects, increasing the inhibition rate up to 87.5% after 48 h at concentration of 1000 μg/mL. Conclusion: The results demonstrated the feasibility of P. vaninii Ljup mycelial fermentation for large-scale production of bioactive and medicinal compounds.
  - 1,960 87