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January-March 2017
Volume 13 | Issue 49
Page Nos. 1-198

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EDITORIAL  

Pharmacognosy Magazine-Completion of 12 years of successful publishing p. 1
Mueen Ahmed
DOI:10.4103/0973-1296.197631  
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ORIGINAL ARTICLES Top

Antidermatophytic and protease-inhibiting activities of zerumbone: A natural sesquiterpene from the rhizome of Zingiber zerumbet (L.) Roscoe ex J.E; Smith p. 2
Madhavankutty Jyothilakshmi, Mathew Jyothis, Gokulanathan Nair Hari Narayanan, Mukalel Sankunni Latha
DOI:10.4103/0973-1296.197649  
Context: Due to increase in the number of patients with impaired immunity, incidence of dermatophytoses has increased considerably. Antidermatophytic agents with anti-inflammatory and protease-inhibiting activities will help in restricting inflammatory response associated with dermatophytoses. Aims: The present study aims to evaluate antidermatophytic and protease-inhibiting activities of zerumbone. Cytotoxicity was tested using Chang liver cell line as a preliminary step in toxicity study. Methods and Materials: Minimum inhibitory concentration (MIC) and minimum fungicidal concentration (MFC) of zerumbone purified from the rhizome of Zingiber zerumbet were determined against Epidermophyton floccosum var. nigricans, Microsporum canis, Microsporum gypseum, and Trichophyton rubrum. MIC was determined according to Clinical and Laboratory Standards Institute (CLSI) method M38-A2. Protease-inhibiting property was tested using trypsin as the enzyme. In vitro cytotoxic effect was studied using the MTT assay. Results: MIC of zerumbone was 8 mg/L against E. floccosum and M. canis and 16 mg/L for M. gypseum and T. rubrum. MFC of zerumbone was 64 mg/L against E. floccosum and M. canis and 128 mg/L for M. gypseum and T. rubrum. Zerumbone exhibited remarkable protease-inhibiting activity. In the MTT assay, IC50 values were 150 and 0.31 µg, respectively, for zerumbone and reference drug. Statistical Analysis Used: For protease inhibition, assay and cytotoxicity assay control and tests were done in triplicate and the results are expressed as mean ± SD, where n = 3. Conclusions: Zerumbone is a novel candidate for use in dermatophytoses therapy because of the combined antifungal, anti-inflammatory (unpublished results), and protease-inhibiting properties. Cytotoxicity of zerumbone was found to be very low compared with the reference drug. Abbreviations used: CFU: colony forming unit, CLSI: Clinical and Laboratory Standards Institute, COX: cyclooxygenase, DMSO: dimethyl sulphoxide, EDTA: ethylene diamine tetra acetic acid, FT-IR: Fourier transform–infra red spectroscopy, HPLC: high-performance liquid chromatography, LOX: lipoxygenase, IMTECH: Institute of Microbial Technology, LCMS: liquid chromatography mass spectrometry, MTT: 3-(4,5 dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide, MTCC: microbial type culture collection, MFC: minimum fungicidal concentration, MIC: minimum inhibitory concentration, MPO: myeloperoxidase, NMR: nuclear magnetic resonance spectroscopy, PAR: proteinase-activated receptor, PBS: phosphate-buffered saline, TCA: trichloro acetic acid
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Wheatgrass extract ameliorates hypoxia-induced mucin gene expression in A549 cells p. 7
Ju hwan Sim, Moon-Hee Choi, Hyun-Jae Shin, Ji-Eun Lee
DOI:10.4103/0973-1296.197660  
Background: Wheatgrass is known to have antioxidant, antiaging, and anti-inflammatory effect. However, its protective effect against hypoxia is not yet evaluated. Objective: In this study, we evaluated the protective and anti-inflammatory effect of wheatgrass against the hypoxia in airway epithelial cells. Materials and Methods: A549 human lung adenocarcinoma cells were incubated in a hypoxic condition (CO2 5%/O2 1%) for 24 hr in the presence of different concentration of wheatgrass 50, 75, 100, and 150 μg/mL, and the magnitude of each immunologic response produced by the A549 cells was compared. The mRNA expression level of mucin gene (MUC), 5A, 5B, 8, GM-CSF, TNF-α, and VEGF were evaluated by using real-time polymerase chain reaction. The MUC proteins level before and after knocking out the hypoxia-inducible factor (hif)-1α via short interfering (si) RNA transfection were assessed by immunoblot analysis. Accordingly, the involved cell signaling pathway was evaluated by immunoblot analysis. Results: The inflammatory cytokines (GM-CSF, TNF- α) and the expressions of MUC 5A, 5B, and 8 were augmented by hypoxia. The augmented MUC expression was decreased by the wheatgrass extract administration. Hif-1α gene expression after hypoxia exposure was decreased by wheatgrass. Knockdown of hif-1α by siRNA reduced the mucin gene expression and which was more enhanced by wheatgrass extract. Conclusion: Theses results suggest that wheatgrass may be useful in the treatment of sinonasal disease by inhibiting mucus hypersecretion in airway epithelium. Abbreviations used: A549: human lung adenocarcinoma cells, GM-CSF: granulocyte-macrophage colony stimulating factor, HIF: hypoxia inducible factor, IL: interleukin, MUC: mucin, MTT: 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide, TNF: tumor necrosis factors, VEGF: vascular endothelial growth factor, si RNA: short interfering RNA
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Using UHPLC and UV-vis fingerprint method to evaluate substitutes for Swertia mileensis: An endangered medicinal plant p. 13
Jie Li, Ji Zhang, Hang Jin, Yuan-Zhong Wang, Heng-Yu Huang
DOI:10.4103/0973-1296.197655  
Background: Millions of people are killed by viral hepatitis every year in the world, whereas many relevant medicines are too expensive to purchase. Swertia mileensis, a medicinal plant for hepatitis in the system of traditional Chinese medicine, has been vanishing gradually because of overexploitation. Objective: To find substitutes of S. mileensis and reduce the cost of purchasing drugs for hepatitis patients, the similarity of phytochemical constituents between S. mileensis and other three Swertia species was compared. Materials and Methods: Both ultra high performance liquid chromatographies and ultraviolet-vis fingerprints of four Swertia species were developed. Methanol extracts of the stems and leaves were used as samples to establish the fingerprint. The calibration curve was drawn for quantitative analysis of swertiamarin. The data of ultra high performance liquid chromatographies were evaluated statistically using similarity analysis and principal component analysis. Results: The result shows a significant difference at area of 204–290 nm in the ultraviolet fingerprint. Swertiamarin, the only one common peak, was defined in chromatographic fingerprints of four Swertia species. The quantitative analysis suggested that the highest concentration of swertiamarin is in S. davidii. The similarity indexes between different samples were almost under 0.60. In the principal component analysis, separate points not only represent the distinction among different species, but also perform chemical discrepancies in content between stems and leaves of one same species. Conclusions: S. angustifolia, S. davidii, and S. punicea are not suitable as substitutes of S. mileensis because of their remarkable differences in entirety and local part. In order to address issues about substitutes and high cost of purchasing drugs, more studies need to undertake. Abbreviation used: UHPLC: Ultra high performance liquid chromatography, UV-vis: Ultraviolet-vis, HBV: Anti-hepatitis virus, DNA: Deoxyribonucleic acid, PCA: Principal component analysis, D-GaIN: D-Galactosamine, BCG: Bacille Calmette-Guerin, LPS: Lipopolysaccharide
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Nardostachys jatamansi root extract modulates the growth of IMR-32 and SK-N-MC neuroblastoma cell lines through MYCN mediated regulation of MDM2 and p53 p. 21
Snehal Suryavanshi, Prerna Raina, Rashmi Deshpande, Ruchika Kaul-Ghanekar
DOI:10.4103/0973-1296.197645  
Aim: The present study evaluated the effect of ethanolic extract of Nardostachys jatamansi roots (NJet) on MYCN mediated regulation of expression of MDM2 and p53 proteins in neuroblastoma cell lines, IMR-32 and SK-N-MC. Materials and Methods: The effect of NJet on cell viability was determined by MTT; and on growth kinetics was evaluated by trypan blue dye exclusion method and soft agar assay. The expression of p53, MDM2 and MYCN proteins in response to NJet treatment was evaluated by immunoblotting. Results: NJet decreased the viability of neuroblastoma cells without affecting the viability of non-cancerous, HEK-293 cells. It altered the growth kinetics of the cancer cells in a dose-dependent manner. NJet down regulated the expression of MYCN and MDM2 proteins with a simultaneous increase in the expression of tumor suppressor protein p53. Conclusions: The present data demonstrated that NJet regulated the growth of IMR-32 and SK-N-MC through reduction in MYCN expression that lead to down regulation of MDM2 protein and increase in p53 expression. These preliminary results warrant further in depth studies to explore the therapeutic potential of Nardostachys jatamansi in the management of neuroblastoma. Abbreviations used: NJet: N. jatamansie thanolic extract; MTT: 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenylthiazolium bromide; FBS: Fetal bovine serum; FITC: Fluorescein isothiocyanate
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Antioxidant and nephroprotective activities of the extract and fractions of Homonoia riparia Lour p. 25
Seena kanniparambil Xavier, Shoja Muhammed Haneefa, Devkar Raviraj Anand, Picheswara Rao polo, Rajalekshmi Maheshwari, Chandrashekara Shastry Shreedhara, Manganahalli Manjunath Setty
DOI:10.4103/0973-1296.197647  
Background: Homonoia riparia is a plant, which is widely used in the indigenous system of medicine for the treatment of urolithiasis, renal disorders and inflammatory conditions. This is the first report on the antioxidant and nephroprotective activities of whole plant of H. riparia. Objective: The present study aims at investigating the in vitro antioxidant and nephroprotective activity of the methanol extract and its different fractions of H. riparia. Methods: Petroleum ether (HRPE), Ethyl acetate (HREA), Butanol (HRBU), aqueous fractions (HRAQ) were prepared from the crude methanol extract of H. riparia (HRM) using liquid partitioning. Total phenolic content, flavonoid content and antioxidant activity assay were performed according to suitable methods. Nephroprotective activities were evaluated by MTT assay using Human Embryonic Kidney cells against cisplatin induced toxicity. Quantification of gallic acid was performed using validated HPTLC method. Results: The studies showed that extract and fractions possess significant nephroprotective activity against cisplatin induced renal toxicity. All the extracts/fractions of whole plant of Homonoia riparia was found to be significantly reducing cisplatin induced toxicity (< 0.05). The highest activity was observed with HRBU and HRAQ with a percentage viability of 293.09 ± 4.3 and 345.07 ± 3.2 at a concentration of 200 µg/ml. Gallic acid was detected in the HRM/fractions using HPTLC. Abbreviations used: HPTLC: High Performance Thin Layer Chromatography, DPPH: 1,1-diphenyl-2-picrylhydrazyl, ABTS: 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) diammonium salt, MTT: 3-(4,5-dimethylthiazolyl-2-yl)-2,5- diphenyl tetrazolium bromide, GAE: Gallic acid equivalents, QE: Quercetin equivalents, HEK: Human Embryonic Kidney, HRM: Methanol extract of H. riparia, HRPE: Petroleum ether fraction of H. riparia, HREA: ethyl acetate fraction of H. riparia, HRBU: Butanol fraction of H.riparia, HRAQ: Aqueous fraction of H. riparia, DMEM: Dulbecco's minimum essential medium, FBS: Foetal bovine serum, DMSO: Dimethyl sulfoxide, ANOVA: One way analysis of variance.
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Isolation and characterisation of a proanthocyanidin with antioxidative, antibacterial and anti-cancer properties from fern Blechnum orientale p. 31
How-Yee Lai, Yau-Yan Lim, Kah-Hwi Kim
DOI:10.4103/0973-1296.197659  
Background:Blechnum orientale Linn. (Blechnaceae), a fern, is traditionally used in the treatment of various ailments, such as skin diseases, stomach pain, urinary bladder complaints, and also as a female contraceptive. Previously, we reported a strong radical scavenging activity, antibacterial activity and cytotoxicity against HT29 colon cancer cells by aqueous extract of B. orientale. Objective: In this study, we attempted to isolate and identify the active compound from the aqueous extract of B. orientale. Materials and Methods: Aqueous extract of B. orientale was subjected to repeated MCI gel chromatography, Sephadex-LH-20, Chromatorex C18 and semi-preparative high performance liquid chromatography and was characterized using nuclear magnetic resonance and electrospray ionization mass-spectrometry spectroscopic methods. Antioxidant activity was determined using 2, 2-diphenyl-1-picrylhydrazyl radical scavenging assay. Antibacterial assays were conducted using disc diffusion whereas the minimum inhibitory concentration (MIC) and minimum bactericidal concentration were determined using the broth microdilution assay. Cytotoxicity was assessed using thiazolylblue tetrazoliumbromide. Results: A polymeric proanthocyanidin consisting of 2-12 epicatechin extension units and epigallocathecin terminal units linked at C4-C8 was elucidated. Bioactivity studies showed strong radical scavenging activity (IC50 = 5.6 ± 0.1 µg/mL), antibacterial activity (MIC = 31.3-62.5 µg/mL) against five gram-positive bacteria and selective cytotoxicity against HT29 colon cancer cells (IC50 = 7.0 ± 0.3 µg/mL). Conclusion: According to our results, the proanthocyanidin of B. orientale demonstrated its potential as a natural source of antioxidant with antibacterial and anti-cancer properties. Abbreviation used: CC: Column chromatography, DP: degree of polymerization, DPPH: 2,2-diphenyl-1-picrylhydrazyl, ESI-MS: electronsprayionisation mass-spectrometry, MBC: Minimum bactericidal concentration, MIC : Minimum inhibitory concentration, MTT: Thiazolyl Blue Tetrazolium Bromide, MRSA: methicillin-resistant Staphylococcus aureus, NMR: nuclear magnetic resonance, TLC: thin layer chromatography, PD: prodelphinidin
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Specific PCR identification between Peucedanum praeruptorum and Angelica decursiva and identification between them and adulterant using DNA barcode p. 38
Bang-Xing Han, Yuan Yuan, Lu-Qi Huang, Qun Zhao, Ling-Ling Tan, Xiang-Wen Song, Xiao-Mei He, Tao Xu, Feng Liu, Jian Wang
DOI:10.4103/0973-1296.197658  
Background: The traditional Chinese medicine (TCM) Qianhu and Zihuaqianhu are the dried roots of Peucedanum praeruptorum and Angelica decursiva, respectively. Since the plant sources of Qianhu and Zihuaqianhu are more complex, the chemical compositions of P. praeruptorum andA. decursiva are significantly different, and many adulterants exist because of the differences in traditional understanding and medication habits. Therefore, the rapid and accurate identification methods are required. Objective: The aim was to study the feasibility of using DNA barcoding to distinguish between Traditional Chinese medicine Qianhu (Peucedanum praeruptorum), Zihuaqianhu (Angelica decursiva), and common adulterants, based on internal transcribed spacer (ITS) sequences, as well as specific PCR identification between P. praeruptorum and A. decursiva. Materials and Methods: The ITS sequences of P. praeruptorum, A. decursiva, and adulterant were studied, and a phylogenetic tree was constructed. Based on the ITS barcode, the specific PCR primer pairs QH-CP19s/QH-CP19a and ZHQH-CP3s/ZHQH-CP3a were designed for P. praeruptorum and A. decursiva, respectively. The amplification conditions were optimized, and specific PCR products were obtained. Results: The results showed that the phylogenetic trees constructed using the BI and MP methods were consistent, and P. praeruptorum and A. decursiva sequence haplotypes formed their own monophyly. The experimental results showed that in PCR products, the target bands appeared in the genuine drug and not in the adulterant, which suggests the high specificity of the two primer pairs. Conclusion: The ITS sequence was ideal DNA barcode to identify P. praeruptorum, A. decursiva, and adulterant. The specific PCR is a quick and effective method to distinguish between P. praeruptorum and A. decursiva. Abbreviations used: TCM: The traditional Chinese medicine, P.: Peucedanum, A.: Angelica, ITS: The internal transcribed spacer, PCR: Polymerase chain reaction, NCBI: National Center for Biotechnology Information, NI: Number of individuals, HN: Haplotype number; GAN: Gen Bank accession numbers, L.: Ligusticum, O.: Ostericum, A.: Angelica, P.: Pimpinella, BI: Bayesian inference, MP: Maximum parsimony, AIC: Akaike Information Criterion, MCMC: Markov Chains Monte Carlo, TBR: Tree bisection-reconnection, LPP: Length of PCR product, PRP: PCR reaction procedure, SNP: Single nucleotide polymorphisms, PP: Posterior probability, BS: Bootstrap.Qun Zhao
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Anticancer potential of nutraceutical formulations in MNU-induced mammary cancer in Sprague Dawley rats p. 46
Gummalla Pitchaiah, Annapurna Akula, Vishala Chandi
DOI:10.4103/0973-1296.197652  
Background: Nutraceuticals help in combating some of the major health problems of the century including cancer, and 'nutraceutical formulations' have led to the new era of medicine and health.Objective: To develop different nutraceutical formulations and to assess the anticancer potential of nutraceutical formulations in N-methyl-N-nitrosourea (MNU)-induced mammary cancer in Sprague Dawley rats. Materials and Methods: Different nutraceutical formulations were prepared using fine powders of amla, apple, garlic, onion, papaya, turmeric, and wheat grass with and without cow urine distillate. Total phenolic content, acute oral toxicity, and microbial load of nutraceutical formulations were assessed. The anticancer potential of nutraceutical formulations was evaluated against MNU-induced mammary cancer in female Sprague Dawley rats. Results: Improvement in total phenolic content was significant (P < 0.001) after self-fortification process. Toxicity studies showed that the nutraceutical formulations were safe to use in animals. Microbial load was within the limits. Significant longer tumor-free days (P < 0.01), lower tumor incidence (P < 0.01), lower tumor multiplicity (P < 0.05) and tumor burden (P < 0.01) were observed for nutraceutical formulation-treated groups. Conclusion: Combination of whole food-based nutraceuticals acted synergistically in the prevention of mammary cancer. Further, the process of fortification is novel and enhanced the anticancer potential of nutraceutical formulations. Abbreviations used: HMNU: N-methyl-N-nitrosourea, CAM: Complementary and Alternative Medicine, NF: Nutraceutical Formulation, SFNF: Self-Fortitfied Nutraceutical Formulation, NFCUD: Nutraceutical Formulation fortified with Cow Urine Disstillate, SFNFCUD: Self-Fortified Nutraceutical Formulation fortified with Cow Urine Disstillate, CPCSEA: Committee for the Purpose of Control and Supervision of Experiments on Animals, OECD: Organisation for Economic Co-operation and Development, TPC: Total Phenolic Content, ANOVA: Analysis of Variance, GAE: Gallic Acid Equivalent, cfu/g: Colony forming unit per g
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Comparative pharmacokinetic profiles of three protoberberine-type alkaloids from raw and bile-processed Rhizoma coptidis in heat syndrome rats p. 51
Yuan Zi-min, Chen Yue, Gao Hui, Lv Jia, Chen Gui-rong, Jing Wang
DOI:10.4103/0973-1296.197632  
Background: The Bile-processed Rhizoma coptidis (BRC), which has a colder drug property than Rhizoma coptidis (RC), is widely used for the treatment of heat syndrome. We compared the pharmacokinetics of the protoberberine-type alkaloids in BRC and RC in rats with heat syndrome to elucidate the bile-processing mechanism. Material and Methods: We established a rapid and sensitive method for simultaneously determining three alkaloids: berberine, palmatine, and jatrorrhizine, in rat plasma based on ultra-performance liquid chromatography/tandem mass spectrometry. The separation was carried out on a Waters ACQUITY BEA C18 column. The mobile phase consisted of acetonitrile (containing 0.1% formic acid) and water (containing 0.1% formic acid and 10 mmol/L ammonium acetate) and carbamazepine was used as an internal standard. The detection was carried out in a multiple reaction monitoring mode (MRM) using electrospray ionization in the positive ion mode. Results: Pharmacokinetic profiles indicated that the Cmax of berberine and palmatine increased two times and the Tmax of the three alkaloids decreased three times after bile processing. AUC0→∞ and AUC0→t of the alkaloids were similar between RC and BRC. Conclusion: The results suggest that bile processing could increase the absorption rate of alkaloids. This study broadens our understanding of Chinese herbal medicine processing. Abbreviation Used: RC: Rhizoma coptidis, BRC: Bile-processed Rhizoma coptidis, HPLC : high-performance liquid chromatography, UPLC-MS/MS: ultra-performance liquid chromatography-mass spectrometry/ mass spectrometry, LC-MS: liquid chromatography-mass spectrometry, MRM : multiple reaction monitoring mode, QC: quality control, RE: relative error, RSD: relative standard deviation, Cmax: maxium of drug concentration, Tmax: time for maxium of drug concentration, AUC: area under concentration-time curve, LLOQ: Linearity and lower limits of quantification, t1/2: half-life, Clz: body clearance
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Chemical components from Aloe and their inhibition of indoleamine 2, 3-dioxygenase p. 58
Ya Nan Sun, Lin Ying Li, Wei Li, Jong Seong Kang, Inkyu Hwang, Young Ho Kim
DOI:10.4103/0973-1296.197633  
Background: In Korea, Aloe is routinely ingested as a traditional medicine or as a component of health beverages. Objective: To research the inhibition of indoleamine 2, 3-dioxygenase (IDO) activities of components from Aloe. Materials and Methods: the compounds were isolated by a combination of silica gel and YMC Rp-18 column chromatography, and their structures were identified by analysis of spectroscopic data (1D, 2D-NMR, and MS). All of the isolated compounds were examined for their ability to inhibit IDO, which actively suppresses immune functions by catalyzing the rate limiting reaction in the conversion of tryptophan to kynurenine. Results: In this phytochemical study, 18 known compounds were isolated from aqueous dissolved Aloe exudates. All of the isolated compounds were examined for their ability to inhibit IDO activities for a series of anthraquinone derivatives (1-7) isolated from the Aloe extract; the IC50 values of these compounds ranged from 39.41 to 53.93 µM. Enzyme kinetic studies of their modes of inhibition indicated that all of the compounds were uncompetitive inhibitors. Conclusion: The aqueous dissolved Aloe exudate can be used as a source of novel natural IDO inhibitors and merit testing as therapeutic agents in the treatments of cancer and immunopathologic diseases, such as autoimmune, inflammatory, and allergic disorders. Abbreviation used: IDO: inhibit indoleamine 2, 3-dioxygenase, TMS: tetramethylsilane, HMQC: heteronuclear multiple quantum correlation, HMBC: heteronuclear multiple bond correlation, COSY: 1H-1H correlation spectroscopy, ESI-MS: Electrospray ionization mass spectrometry, DMSO: dimethyl sulfoxide
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Simultaneous determination of eight phenolic acids, five saponins and four tanshinones for quality control of compound preparations containing Danshen-Sanqi herb-pair by HPLC-DAD p. 64
Hong Yao, Xiaomei Huang, Shaoguang Li, Youjia Wu, Xinhua Lin, Peiying Shi
DOI:10.4103/0973-1296.197651  
Background: The herb-pair, Salviaemiltiorrhizae (Danshen, DS) and Panaxnotoginseng (Sanqi,SQ), often occurs in traditional Chinese medicine prescriptions used for the treatment of cardiovascular diseases in clinics in Asian areas. Many commercial preparations containing the DS-SQ herb-pair were produced by various manufactures with the different production process. The raw materials were from different sources, which raised a challenge to control the quality of the herb-pair medicines. Objective: In this paper, a high-performance liquid chromatography (HPLC) method was developed to simultaneously determine seventeen bioactive components, including 8 phenolic acids, 4 tanshinones, and 5 saponins, for quality control of compound preparations containing DS-SQ herb-pair. The chromatographic separation was studied on an UltimateTM XB-C18 column (150 mm × 4.6 mmi.d., 3.5 μm) with a mobile phase composed of 0.5% aqueous acetic acid and acetonitrile using a gradient elution in 70 min. Results: The optimum detection wavelength was set at 288 nm for phenolic acids and tanshinones, and 203 nm for saponins. The method was validated sufficiently by examining the precision, recoveries, linearity, range, LOD and LOQ, and was successfully applied to quantify the seventeen compounds in five commercial preparations containing DS-SQ herb-pair. Conclusions: It is the first time to report the rapid and simultaneous analysis of the seventeen compounds with the base-line separation of peaks for ginsenoside Rg1 and Re in 70 min by routine HPLC. This HPLC method could be considered as good quality criteria to control the quality of preparations containing DS-SQ herb-pair. Abbreviations used: DS: Salviae miltiorrhizae; SQ: Panaxnotoginseng; HPLC: high-performance liquid chromatography; DAD: diode array detector; LOD: limit of detection; LOQ: limit of quantification; TCMs: Traditional Chinese medicines; GDDP: Guanxin Danshen dripping pills; FDDP: Fufang Danshen dripping pills; FDT: Fufang Danshen tablets; FDC: Fufang Danshen capsules; GP: Guanxin pills
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Obtaining a dry extract from the Mikania laevigata leaves with potential for antiulcer activity p. 76
Mariana Viana Pinto, Ezequiane Machado Oliveira, Jose Luiz Rodrigues Martins, Jose Realino de Paula, Elson Alves Costa, Edemilson Cardoso da Conceição, Maria Teresa Freitas Bara
DOI:10.4103/0973-1296.197640  
Background: Mikania laevigata leaves are commonly used in Brazil as a medicinal plant.Objective: To obtain hydroalcoholic dried extract by nebulization and evaluate its antiulcerogenic potential.Materials and Methods: Plant material and hydroalcoholic extract were processed and analyzed for their physicochemical characteristics. A method using HPLC was validated to quantify coumarin and o-coumaric acid. Hydroalcoholic extract was spray dried and the powder obtained was characterized in terms of its physicochemical parameters and potential for antiulcerogenic activity.Results: The analytical method proved to be selective, linear, precise, accurate, sensitive, and robust. M. laevigata spray dried extract was obtained using colloidal silicon dioxide as adjuvant and was shown to possess 1.83 ± 0.004&#37; coumarin and 0.80 ± 0.012% o-coumaric acid. It showed significant antiulcer activity in a model of an indomethacin-induced gastric lesion in mice and also produced a gastroprotective effect.Conclusion: This dried extract from M. laevigata could be a promising intermediate phytopharmaceutical product. Abbreviations used: DE: M. laevigata spray dried extract, HE: hydroalcoholic extract
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Anti-nociceptive effect of 7-methoxy coumarin from Eupatorium Triplinerve vahl (Asteraceae) p. 81
Binoy Varghese Cheriyan, Parimala Kadhirvelu, Jagan Nadipelly, Jaikumar Shanmugasundaram, Vijaykumar Sayeli, Viswanathan Subramanian
DOI:10.4103/0973-1296.197650  
Aim: To evaluate the anti-nociceptive activity of 7-methoxy coumarin isolated from ethyl acetate fraction of the alcoholic extract of Eupatorium triplinerve Vahl. Materials and Methods: The shade dried leaves of E. triplinerve were extracted with ethyl alcohol and the extract was condensed. This extract was fractionated with n-hexane, ethyl acetate, and n-butanol. The ethyl acetate fraction was subjected to column chromatography which yielded a crystalline compound-A, which was investigated for spectral characteristics. Pharmacological studies: The isolated compound-A was subjected to behavioral studies and anti-nociceptive evaluation in mice by acetic acid induced writhing and formalin induced nociception. Results: The spectral studies indicated that the structure of compound-A complies with 7- methoxy coumarin. Pre-treatment with 7-methoxy coumarin reduced the number of abdominal constrictions in mice and decreased the time spent in paw licking and biting response in formalin assay. There were no significant behavioral changes. Conclusion: A dose dependent anti-nociceptive action of 7- methoxy coumarin was revealed by the present experiments which support the traditional use of E. triplinerve in pain and inflammatory disorders. Abbreviation used: TLC-Thin layer chromatography, Kg-kilogram, g-gram, TXB2-Thromboxane B2, UV-Ultraviolet, IgE-Immunoglobulin E, s.c-subcutaneous, p.o-oral route
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Effect of absolute from Hibiscus syriacus L. flower on wound healing in keratinocytes p. 85
Seok Won Yoon, Kang Pa Lee, Do-Yoon Kim, Dae Il Hwang, Kyung-Jong Won, Dae Won Lee, Hwan Myung Lee
DOI:10.4103/0973-1296.197644  
Background: Proliferation and migration of keratinocytes are essential for the repair of cutaneous wounds. Hibiscus syriacus L. has been used in Asian medicine; however, research on keratinocytes is inadequate. Objective: To establish the dermatological properties of absolute from Hibiscus syriacus L. flower (HSF) and to provide fundamental research for alternative medicine. Materials and Methods: We identified the composition of HSF absolute using gas chromatography-mass spectrometry analysis. We also examined the effect of HSF absolute in HaCaT cells using the XTT assay, Boyden chamber assay, sprout-out growth assay, and western blotting. We conducted an in-vivo wound healing assay in rat tail-skin. Results: Ten major active compounds were identified from HSF absolute. As determined by the XTT assay, Boyden chamber assay, and sprout-out growth assay results, HSF absolute exhibited similar effects as that of epidermal growth factor on the proliferation and migration patterns of keratinocytes (HaCaT cells), which were significantly increased after HSF absolute treatment. The expression levels of the phosphorylated signaling proteins relevant to proliferation, including extracellular signal-regulated kinase 1/2 (Erk 1/2) and Akt, were also determined by western blot analysis. Conclusion: These results of our in-vitro and ex-vivo studies indicate that HSF absolute induced cell growth and migration of HaCaT cells by phosphorylating both Erk 1/2 and Akt. Moreover, we confirmed the wound-healing effect of HSF on injury of the rat tail-skin. Therefore, our results suggest that HSF absolute is promising for use in cosmetics and alternative medicine. Abbreviations used: HSF: Hibiscus syriacus L. flower, Erk 1/2: extracellular signal-regulated kinase 1/2, EGF: epidermal growth factor, GC/MS: gas chromatography-mass spectrometry, DMEM: dulbecco's modified eagle medium, FBS: fetal bovine serum, BSA: bovine serum albumin, p-Akt: phosphorylation of Akt, p-Erk 1/2: phosphorylation of Erk 1/2
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Osteoblasts proliferation and differentiation stimulating activities of the main components of Epimedii folium p. 90
Mingming Liu, Haiyan Xu, Yong Ma, Jian Cheng, Zhen Hua, Guicheng Huang
DOI:10.4103/0973-1296.197654  
Background: Osteoporosis is a disease of bones that leads to an increased risk of fracture. Epimedii Folium is commonly used for treating bone fractures and joint diseases for thousands of years in China. Methods: This study was aimed to screen active components, which might have the potency to stimulate osteoblasts proliferation and differentiation in Epimedii Folium. An HPLC method was established to analyze the main components in Epimedii Folium. The MTT and ALP methods were utilized for the assay of osteoblasts proliferation and differentiation activity. Bavachin, a flavonoid compound was treated as the positive control. Results: Totally eight compounds have been identified by comparing their retention time with correspondent standard substances. Icariside I and icariside II significantly stimulated cell proliferation and osteoblasts differentiation. All these compounds were found with a characterized flavonoid structure in each of their molecule backbones. Conclusion: These results lead to a hypothesis that flavonoid monoglycoside structure might be crucial to exhibit the activity. The structure–effect relationship of these compounds with flavonoid monoglycoside structure in mouse primary calvarial osteoblasts needs to be explored in further research. Abbreviations used: HPLC: High performance liquid chromatography, MTT: Methylthiazolyldiphenyl - tetrazolium bromide, ALP: Alkaline phosphatase
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Evaluate the antigenotoxicity and anticancer role of β-sitosterol by determining oxidative DNA damage and the expression of phosphorylated mitogen-activated protein kinases', c-fos, c-jun, and endothelial growth factor receptor p. 95
Ramalingam Sharmila, Ganapathy Sindhu
DOI:10.4103/0973-1296.197634  
Background: Plant sterols are the major source of micronutrients and have not shown any obvious side effects in human. β-sitosterol is one of the most prevalent phytosterols which have been recorded in ancient medicinal history for its use in the treatment of many chronic diseases,especially cancer. The modulations of mitogen-activated protein kinases' (MAPKs') play a crucial role in the development of human renal cell carcinoma. Objective: The aim of the current study is to evaluate the antigenotoxic and anticancer role of β-sitosterol against renal carcinogen. Materials and Methods: The extent of DNA damage was assessed by the comet assay. The status of p-p38 MAPK,p-c-Jun N-terminal kinase,p-extracellular-signal regulating kinase (ERK),c-fos,c-jun,and endothelial growth factor receptor (EGFR) were analyzed by western blot and polymerase chain reaction techniques. To further confirm the inhibition of ERK-2 by β-sitosterol,molecular docking study was performed. Results: Extensive DNA damage in acute study and a significant increase in levels of p-MAPKs',c-fos,c-jun,and EGFR was observed in N-diethylnitrosamine (200 mg/kg bw) and ferric nitrilotriacetate (9 mg/kg bw) alone treated rats. Rats which are pretreated with 20 mg/kg bw of β-sitosterol reduced the DNA damage and restored the elevated levels of above-mentioned markers (p < 0.05). The binding free energy obtained for β-sitosterol for ERK-2 was found to be-5.578. Conclusion: Therefore,it has been concluded that β-sitosterol has a strong potential against genotoxic as well as suppress neoplastic transformation in experimental renal cancer. Abbreviation used: AP-1: Activator protein-1,DEPC: Diethyl pyrocarbonate,EDTA: Ethylenediaminetetraacetic acid,EGFR: Endothelial growth factor receptor,ERK: Extracellular-signal regulating kinase,Fe-NTA: Ferric nitrilotriacetate,GAPDH: Glyceraldehyde-3-phosphate dehydrogenase,HBSS: Hank's balanced salt solution,JNK: c-Jun N-terminal kinase,MAPK: Mitogen-activated protein kinase,DEN: N-diethylnitrosamine,RCC: Renal cell carcinoma,SDS-PAGE: Sodium dodecyl sulfate polyacrylamide gel electrophoresis
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Effects of potassium sulfate [K2SO4] on the element contents, polyphenol content, antioxidant and antimicrobial activities of milk thistle [Silybum marianum] p. 102
Gulsum Yaldiz
DOI:10.4103/0973-1296.197641  
Background:Silybum marianum L. (Milk thistle) is native to the Mediterranean basin and is now widespread throughout the world. It's sprout is used as a herbal medicine for the treatment of liver disease for centuries. The seeds of milk thistle contain silymarin, an isomeric mixture of flavonolignans [silybin, silychristin, and silydianin. Silymarin acts as a strong anti-hepatotoxic. Objectives: The objective of this study was to evaluate the influences of potassium sulfate [K2SO4] fertilizer doses on polyphenol content, some nutrient elements, antioxidant and antimicrobial activities of milk thistle at experimental fields of Ordu University in Turkey. Methods: The antimicrobial activities of seed ethanol extracts and seed oil were tested in vitro against Pseudomonas aeruginosa (P. aeruginosa), Escherichia coli, (E. coli) Staphylococcus aureus (S. aureus), Aspergillus niger (A. niger) and Candida albicans (C. albicans) using the disc diffusion method. Free radical scavenging activity of the ethanolic extracts of milk thistle was determined spectrophotometrically by monitoring the disappearance of 2, 2-diphenyl-1-picrylhydrazil (DPPH·) at 517 nm according to the method described by Brand-Williams et al.[17] The phenolic contents in the ethanolic extracts of milk thistle were determined according to the procedure described by Slinkard and Singleton[19] with a slight modification of using a Folin-Ciocalteu phenolic reagent. The amount of total flavonoid in the ethanolic extracts was measured by aluminum chloride [AlCl3] colorimetric assay. The ions in aerosol samples were determined by using Dionex ICS 1100 Series ion chromatography. Results: Seed and seed oils obtained from obvious doses of potassium sulfate [0, 30, 60, 90 and 120 kg ha -1 fertilizer applications showed antimicrobial activities against E. coli, A. niger and P. aeruginosa. The application of 90 kg ha-1 of K2SO4 on seed oil resulted in the highest antimicrobial activities. At 100 µg mL-1and 200 µg mL-1, except the highest potassium application [120 kg ha -1 extract, all extracts showed high and similar DPPH scavenging activity. The highest phenolic compounds were obtained with 30 kg ha-1 of K2SO4, whereas the use of 60 kg ha-1 caused the highest total flavonoid content. This plant is a good source of K+, Ca+2, PO4-3, and Cl-1. Conclusion: In this study, increasing doses of potassium sulfate had significant effect on element, polyphenol content, antioxidant and antimicrobial activities of the milk thistle. Abbreviations used: AlCl3: aluminum chloride, Ca+2: calcium, Cl-: chloride, Cr: chromium CE: catechol equivalents, DPPH: 2,2-diphenylpicrylhydrazyl, ABTS: 2,2'-azino-bis-3-ethylbenzthiazoline-6-sulphonic acid, DAP: diamonyum fosfat, F-: fluoride, Fe: iron, K2SO4: potassium sulfate, K+ : potassium, Li+: lithium, Mg+2 : magnesium, NH4+ : amonyum, Na+: sodium, NO2-: nitrite, NO3-: nitrate, Ni: nickel, NaNO2: sodium nitrite, NaOH: sodium hidroksit. ND: Not detectable, PO4-3: phosphorus, Zn: zinc
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Antioxidant and hepatoprotective potential of phenol-rich fraction of Juniperus communis Linn. leaves p. 108
Akash Ved, Amresh Gupta, Ajay Kumar Singh Rawat
DOI:10.4103/0973-1296.197648  
Background: Juniperus communis Linn. is an important plant in India traditional system of medicine which is widely used by different tribes in many countries. Objective: In the present study, the antioxidant, cytotoxic and hepatoprotective activities of Juniperus communis leaves were investigated against various models. Materials and Methods: ethanolic extract (70% v/v) of J. communis leaves was successively extracted using hexane and ethyl acetate to prepare various fractions. Total phenol content was resolute by the Folin-Ciocalteau's process. The antioxidant properties of the different fractions/extract of leaves of J. communis were examined by 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging activity and Fe2+ chelating ability. Cytotoxic activity was examined by cell viability assay on HepG2 cells. Hepatoprotective activity of ethyl acetate fraction (EAF) evaluated against PCM-Paracetamol-induced hepatic damage in Wistar albino rats. Results: Total phenol content was found maximum 315.33 mg/GAE/g in EAF. Significant scavenging activity were found for EAF (IC50 = 177 μg/ml) as compared to standard BHT (IC50 = 138 μg/ml), while EAF showed good Fe2+ chelating ability having an IC50 value of 261 mg/ML compared to standard ethylenediaminetetraacetic acid (7.7 mg/mL). It was found that EAF treated group shows remarkable decrease in serum Aspartate aminotransferase, serum Alanine aminotransferase, total bilirubin, direct bilirubin, and alkaline phosphatase level in treatment group as compared to the hepatotoxic group. Conclusion: EAF of J. communis leaves is found to be potent antioxidant and hepatoprotective without any cytotoxicity and it can also be included in nutraceuticals with notable benefits for mankind or animal health. Abbreviations used: HepG2: Liver hepatocellular carcinoma, BHT: Butylated hydroxytoluene, PCM: Paracetamol, IC50: Half maximal inhibitory concentration, RSA: Radical Scavenging Activity, WST: Water-soluble tetrazolium.
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Effects of insecticidal ketones present in mint plants on GABAA receptor from mammalian neurons p. 114
Mariela Eugenia Sánchez-Borzone, Leticia Delgado Marin, Daniel Asmed García
DOI:10.4103/0973-1296.197638  
Background: The genus Mentha, an important member of the Lamiaceae family, is represented by many species commonly known as mint. The insecticidal activity of Mentha oil and its main components has been tested and established against various insects/pests. Among these, the ketone monoterpenes that are most common in different Mentha species demonstrated insect toxicity, with pulegone being the most active, followed by carvone and menthone. Considering that the GABAA receptor (GABAA-R) is one of the main insecticide targets on neurons, and that pulegone would modulate the insect GABA system, it may be expected that the insecticidal properties of Mentha ketones are mediated by their interaction with this receptor. Objective: In order to discern the pharmacological actions of these products when used as insecticides on mammalian organisms, we evaluated the pharmacologic activity of ketones, commonly present in Mentha plants, on native GABAA-R from rats. Materials and Methods: Determination of ketones effects on allosterically enhanced benzodiazepine binding, using primary cultures of cortical neurons, which express functional receptors and MTT assay to evaluate their cell toxicity. Results: Our results seem to indicate that ketone components of Mentha, with proven repellent or insecticide activity, were able to behave as GABAA-R negative allosteric modulators in murine cells and consequently could exhibit convulsant activity in mammalians. Only pulegone at the highest assayed concentration (2 mM) showed a significant reduction in cell viability after exposure for 24 hr. Conclusion: The present results strongly suggest that the ketone components of Mentha are able to exhibit convulsant activity in mammalian organisms, but functional assays and β€͸in vivo experiments would be necessary to corroborate this proposed action. Abbreviations used: GABA: gamma aminobutyric acid, GABAA-R: GABAA receptor, MTT: 1-(4,5-dimethylthiazol-2-yl)-3,5-diphenylformazam, DMEM: Dulbecco's modified minimum essential mèdium, [3H]TBOB: [3H] t-Butylbicycloorthobenzoate
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Cytotoxic activities of naturally occurring oleanane-, ursane-, and lupane-type triterpenes on HepG2 and AGS cells p. 118
Heejung Yang, Hyun Woo Kim, Young Choong Kim, Sang Hyun Sung
DOI:10.4103/0973-1296.196308  
Background: It is well known that the naturally occurring modified triterpenes in plants have a wide diversity of chemical structures and biological functions. The lupane-oleanane-and ursane-type triterpenes are the three major members of natural triterpenes with a wide range of biological properties. A systematic approach is necessary to review their structures and biological activities according to the backbones and the different substituents. Objective: Thirty lupane-(L1-7)oleanane-(O1-14)and ursane-type (U1-9) triterpenes with structural diversity were examined to evaluate their cytotoxic activities against two cancer cell lineshuman hepatocellular carcinoma (HepG2) and AGS cells. Materials and Methods: They were isolated from Hedera helixJuglans sinensisand Pulsatilla koreana using a series of column chromatography methods and were treated to evaluate their cytotoxic activities against HepG2 and AGS human gastric adenocarcinoma cell. Furthertwo triterpenes showing the most potent activities were subjected to the apoptotic screening assay using flow cytometry. Results: The polar groupssuch as an oxo group at C-1a free hydroxyl at C-2C-3or C-23and a carboxylic moiety at C-28as well as the type of backboneexplicitly increased the cytotoxic activity on two cancer cells. O5 and U5 showed significantly the potent cytotoxic activity in comparison to other glycosidic triterpenes. In annexin-V/propidium iodide (PI) staining assaythe percentage of late apoptosis (annexin-V+/PI+) 12 and 24 h after treatment with O5 and U5 at 25 μM increased from 14.5% to 93.1% and from 46.4% to 49.1%respectivelyin AGS cells. The cytotoxicity induced by O5 showed a significant difference compared to U5 for 12 and 24 h. Conclusion: In the studywe can suggest the potent moieties which influence their cytotoxic activities against two cancer cells. The polar groups at C-1C-2C-3C-23and C-28 and the linkage of sugar moieties influenced the different cytotoxic activities.
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Vincamine alleviates amyloid-β 25-35 peptides-induced cytotoxicity in PC12 cells p. 123
Jianfeng Han, Qiumin Qu, Jin Qiao, Jie Zhang
DOI:10.4103/0973-1296.196309  
Objective: Vincamine is a plant alkaloid used clinically as a peripheral vasodilator that increases cerebral blood flow and oxygen and glucose utilization by neural tissue to combat the effect of aging. The main purpose of the present study is to investigate the influence of vincamine on amyloid-β 25-35 (Aβ25-35) induced cytotoxicityto gain a better understanding of the neuroprotective effects of this clinically used anti-Alzheimer's disease drug. Materials and Methods: Oxidative stress was assessed by measuring malondialdehydeglutathioneand superoxide dismutase (SOD) levels. Cell viability was assessed by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. Cell apoptosis detection was performed using an Annexin-V-FITC Apoptosis Detection Kit. The production of reactive oxygen species (ROS) was determined using an ROS Assay Kit. Western blot detection was carried out to detect the protein expression. Results: Our studies showed that pretreatment with vincamine could reduce Aβ25-35 induced oxidative stress. Vincamine markedly inhibited cell apoptosis dose-dependently. More importantlyvincamine increased the phosphatidylinositol-3 kinase (PI3K)/Akt and Bcl-2 family protein ratios on preincubation with cells for 2 h. Conclusion: Above observation led us to assume that one possible mechanism of vincamine protects Aβ25-35-induced cell death could be through upregulation of SOD and activation of the PI3K/Akt pathway. Abbreviation used: Aβ25-35: Amyloid-β 25-35; AD: Alzheimer's disease; BCA: Bicinchoninic acid; GSH: glutathione; PBS: Phosphate buffered solution; SDS: Sodium dodecylsulphate; SOD: Superoxide dismutase
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Therapeutic effect of Dendrobium candidum on lupus nephritis in mice p. 129
Qiang Wang, Peng Sun, Rui Wang, Xin Zhao
DOI:10.4103/0973-1296.197653  
Context: Dendrobium candidum (D.candimum) widely is a functional drug. The curative effect of D. candidum on lupus nephritis has been studied in vivo. Materials and Method: The DBA/2 and B6D2F1 mice were used for this in vivo experiment. The 50% effective dose (ED50) was used to check the effective concentration for this study. Then the SCr, BUN, TC, TG, IL-6, IL-12, TNF-α, and IFN-γ levels were determined by kits. The output of urine protein was determined by means of Coomassie Brilliant Blue, and the auto-antibody dsDNA was determined with titer plate technology and indirect immunofluorescence. The NF-κB, IκB-α, TGF 'β1, Fas, and FasL expressions were measured by RT-PCR and western blot assay. The component analysis of D. candidum was determined by nuclear magnetic resonance. Results: Based on the ED50 result at 329 mg/kg, 200 and 400 mg/kg doses were chosen for this study. SCr, BUN, TC and TG levels of 400 mg/kg D. candidum mice were lower than control mice, TP and ALB levels were higher than control mice. The control and 400 mg/kg treated mice tested positive for dsDNA at the end of sixth and tenth week after the experiment began. The glomerular number of 400 mg/kg treated mice was more than control group. Treatment with 400 mg/kg D. candidum reduced IL-6, IL-12, TNF-α and IFN-γcytokine levels as compared to control mice. D. candidum decreased NF-κb, TGF 'β1, Fas, FasL and increased IκB-α expressions in kidney tissue. There were 11 compounds in dry D. candidum, these compounds might make the curative effects of lupus nephritis. Conclusion: D. candidum showed a potential curative effect on lupus nephritis. It could be used as a health medicine on lupus nephritis. Abbreviations used: LN: Lupus nephritis, SLE: systemic lupus erythematosus, D. candidum: Dendrobium candidum; IL-6: interleukin-6, IL-12: interleukin-12, TNF-α: tumor necrosis factor alpha, IFN-γ: Interferon-gamma, SCr: serum creatinine, BUN: blood urea nitrogen, TC: total cholesterol, TG: triglyceride, TP: total protein, ALB: albumin.
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High performance liquid chromatography-diode array detector method for the simultaneous determination of five compounds in the pulp and seed of sea buckthorn p. 136
Lu Zhao, E Wen, Halmuart Upur, Shuge Tian
DOI:10.4103/0973-1296.197656  
Context: Sea buckthorn (Hippophae rhamnoides L.) as a traditional Chinese medicinal plant has various uses in Xinjiang. Objective: A reversed-phase rapid-resolution liquid-chromatography method with diode array detector was developed for simultaneous determination of protocatechuic acid, rutin, quercetin, kaempferol, and isorhamnetin in the pulp and seed of sea buckthorn, a widely used traditional Chinese medicine for promoting metabolism and treating scurvy and other diseases. Settings and design: Compounds were separated on an Agilent ZORBAX SB-C18 column (4.6 mm × 250 mm, 5 μm; USA) with gradient elution using methanol and 0.4% phosphoric acid (v/v) at 1.0 mL/min. Detection wavelength was set at 280 nm. Materials and Methods: The fruits of wild sea buckthorn were collected from Wushi County in Aksu, Xinjiang Province. Statistical performances: The RSD of precision test of the five compounds were in the range of 0.60-2.22%, and the average recoveries ranged from 97.36% to 101.19%. Good linearity between specific chromatographic peak and component qualities were observed in the investigated ranges for all the analytes (R2 > 0.9997). Results: The proposed method was successfully applied to determine the levels of five active components in sea buckthorn samples from Aksu in Xinjiang. Conclusions: The proposed method is simple, fast, sensitive, accurate, and suitable for quantitative assessment of the pulp and seed of sea buckthorn. Abbreviation used: PR: protocatechuic acid, RU: rutin, QU: quercetin, KA: kaempferol, IS: isorhamnetin, HPLC: high-performance liquid chromatography, HPLC-DAD: high performance liquid chromatographydiode array detector, LOD: linearity and limit of detection, LOQ: limit of quantitation, RSD: relative standard deviation
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Depolarizing effects of daikenchuto on interstitial cells of cajal from mouse small intestine p. 141
Hyungwoo Kim, Hyun Jung Kim, Dongki Yang, Myeong Ho Jung, Byung Joo Kim
DOI:10.4103/0973-1296.196312  
Background: Daikenchuto (DKT; TJ-100, TU-100)a traditional herbal medicineis used in modern medicine to treat gastrointestinal (GI) functional disorders. Interstitial cells of Cajal (ICCs) are the pacemaker cells of the GI tract and play important roles in the regulation of GI motility. Objective: The objective of this study was to investigate the effects of DKT on the pacemaker potentials (PPs) of cultured ICCs from murine small intestine. Materials and Methods: Enzymatic digestions were used to dissociate ICCs from mouse small intestine tissues. All experiments on ICCs were performed after 12 h of culture. The whole-cell patch-clamp configuration was used to record ICC PPs (current clamp mode). All experiments were performed at 30-32°C. Results: In current-clamp modeDKT depolarized and concentration-dependently decreased the amplitudes of PPs. Y25130 (a 5-HT3 receptor antagonist) or SB269970 (a 5-HT7 receptor antagonist) did not block DKT-induced PP depolarizationbut RS39604 (a 5-HT4 receptor antagonist) did. Methoctramine (a muscarinic M2 receptor antagonist) failed to block DKT-induced PP depolarizationbut pretreating 4-diphenylacetoxy-N-methylpiperidine methiodide (a muscarinic M3 receptor antagonist) facilitated blockade of DKT-induced PP depolarization. Pretreatment with an external Ca2+-free solution or thapsigargin abolished PPsand under these conditionsDKT did not induce PP depolarization. FurthermoreGinseng radix and Zingiberis rhizomes depolarized PPswhereas Zanthoxyli fructus fruit (the third component of DKT) hyperpolarized PPs. Conclusion: These results suggest that DKT depolarizes ICC PPs in an internal or external Ca2+-dependent manner by stimulating 5-HT4 and M3 receptors. Furthermorethe authors suspect that the component in DKT largely responsible for depolarization is probably also a component of Ginseng radix and Zingiberis rhizomes. Abbreviation used: DKT: DaikenchutoGI: GastrointestinalICCs: Interstitial cells of CajalPPs: Pacemaker Potentials.
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Antiprotozoal constituents from Annona cherimola Miller, a plant used in mexican traditional medicine for the treatment of diarrhea and dysentery p. 148
Fernando Calzada, Jose Correa-Basurto, Elizabeth Barbosa, David Mendez-Luna, Lilian Yepez-Mulia
DOI:10.4103/0973-1296.197636  
Background:Annona cherimola Miller (Annonaceae) is a medicinal plant frequently recommended in Mexican traditional medicine for the treatment of gastrointestinal disorders such as diarrhea and dysentery.Objective: This work was undertaken to obtain information that support the traditional use of A. cherimola, on pharmacological basis using in vitro and computational experiments.Material and Methods: Bioassay-guided fractionation of the ethanol extract of the leaves of A. cherimola afforded five phenolic compounds:caffeic acid, quercetin, kaempferol, nicotinflorin, and rutin.Results: The in vitro antiprotozoal assay showed that kaempferol was the most potent antiamoebic and antigiardial compound with IC50 values of 7.9 μg/mL for Entamoeba histolytica and 8.7 μg/mL for Giardia lamblia. Computational molecular docking study showed that kaempferol interacted in a region different than metronidazole in the enzyme pyruvate:ferredoxin oxidoreductase (PFOR).Conclusion: Considering that PFOR is a target of metronidazole; kaempferol may be a lead compound for the development of novel antiprotozoal agent. Also, these findings give support to the use of A. cherimola in the traditional medicine from México for the treatment of diarrhea and dysentery. Abbreviations used: PFOR:Pyruvate:ferredoxin oxidoreductase, G:lamblia:Giardia lamblia, E:histolytica:Entamoeba histolytica
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Trichosanthes kirilowii exerts androgenic activity via regulation of PSA and KLK2 in 22Rv1 prostate cancer cells p. 153
Soo-Jin Jeong, Ji-Yoon Choi, Mi-Sook Dong, Chang-Seob Seo, Hyeun-Kyoo Shin
DOI:10.4103/0973-1296.197657  
Background: The androgen comprises a group of hormones that play roles in male reproductive activity as well as personal characteristics. Objective: We investigated the androgenic activity of various herbal medicines in human prostate cancer 22Rv1 cells. Materials and Methods: Herbal extracts of Trichosanthes kirilowii (TK), Asarum sieboldii (AS), Sanguisorba officinalis (SO), and Xanthium strumarium (XS) were selected to have androgenic effects based on a preliminary in vitro screening system. Results: TK, AS, SO, and XS enhanced the proliferation of 22Rv1 cells without having cytotoxic effects. All tested herbal extracts increased androgen receptor (AR)-induced transcriptional activity in the absence or presence of dihydrotestosterone (DHT). In an AR-binding assay, TK, but not AS, SO, or XS, produced a significant inhibition of AR binding activity, indicating it has androgenic activity. Additionally, TK treatment positively regulated mRNA expression of the AR-related molecular targets prostate-specific antigen (PSA) and kallikrein 2 (KLK2) compared with untreated control. Conclusion: Taken together, TK-enhanced AR-mediated transcriptional activity might be an attractive candidate drug for treating androgen-related diseases. Abbreviations used: BPH: benign prostatic hyperplasia; AR: androgen receptor; DHT: dihydrotestosterone; PSA: prostate-specific antigen; TK: Trichosanthes kirilowii; AS: Asarum sieboldii; SO: Sanguisorba officinalis; XS: Xanthium strumarium; ATCC: American Type Culture Collection; FBS: fetal bovine serum; PBS: phosphate-buffered saline; SD: standard deviation; ARE: androgen-responsive element; KLK: kallikrein
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Technological evaluation of emulsions containing the volatile oil from leaves of Plectranthus Amboinicus Lour p. 159
Pablo Queiroz Lopes, Fabíola Bernardo Carneiro, Ana Braz Letícia de Sousa, Sócrates Golziodo Santos, Elquio Eleamen Oliveira, Luiz Alberto Lira Soares
DOI:10.4103/0973-1296.197646  
Background: Plectranthus amboinicus Lour is a species which is widespread throughout tropical countries where it is widely used against respiratory tract disorders such as bronchodilator, antitussive, and expectorant conditions. Objective: This study aims to characterize the essential oil of P. amboinicus (PaEO) and produce and evaluate emulsions containing PaEO. Materials and Methods: The essential oil was characterized by physical-chemical analyses for density, refractive index, 90% ethanol solubility, color, appearance, and identification by gas chromatography coupled to mass spectrometry detection. The emulsions were prepared following a hydrophile-lipophile balance [HLB] spreadsheet design from two nonionic surfactants (Span 80® and Tween 20®) producing HLB values ranging from 4.3 to 16.7. The products were stored at room temperature at 5°C. The emulsion stabilities were tested both in the long and short-term.Results: The PaEO was obtained by steam distillation and the total extraction was reached after 3 hours yielding of 0.2% (w/w). This essential oil was characterized by physicochemical analyses for density [1.5 g.ml-1], refraction index [0.9167], ethanol 90% solubility [1:2], color, and appearance (yellow/clear). Nineteen components were identified in the oil, among them the sesquiterpenes: carvacrol [33.50%], p-cymene [28.20%] and γ-terpinene [14.77%]. The emulsions obtained successfully showed, for the first time, HLB values for essential oils from Plectranthus amboinicus [15.7].Conclusion: The experimental data shows a relationship between HLB values of the surfactant mixtures contributing to the emulsified systems production containing phytopharmaceuticals. Such an approach is of great importance to the development of lipid carriers for therapeutic drugs. Abbreviations used: PaEO: essential oil of P. amboinicus, HLB: hydrophilic-lipophilic balance, CI: Creaming Index, MET: micro-emultocrit technique
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Feeble antipyretic, analgesic, and anti-inflammatory activities were found with regular dose 4'-O-β-D-glucosyl-5-O-methylvisamminol, one of the conventional marker compounds for quality evaluation of Radix Saposhnikoviae p. 168
Jing-Ming Yang, Hua Jiang, Hong-Liang Dai, Zi-Wei Wang, Gui-Zhi Jia, Xiang-Cai Meng
DOI:10.4103/0973-1296.197637  
Introduction: 4'-O-β-D-glucosyl-5-O-methylvisamminol (GML) is a conventional marker compound for quality control of Radix Saposhnikoviae. Despite that, neither pharmacodynamic or pharmacokinetic information is available with regard to GML. As such, the aim of thisstudy was to assess the conventional evaluation indices for the quality of Radix Saposhnikoviae. Materials and methods: Pyretic animal model, hot plate test, and ear edema model were established to evaluate and compare the antipyretic, analgesic, and anti-inflammatory effect of the chromone derivativescimifugin, prime-O-glucosylcimifugin (PGCN), and GML in Radix Saposhnikoviae. High performance liquid chromatography separation and analysis was used to obtain pharmacokinetic parameters. Simulated gastric fluid and simulated intestinal fluid was used to investigate the metabolite profiles of PGCN and GML in gastrointestinal tract. Results: Cimifugin exerted a marked dose-dependent antipyretic, analgesic, and anti-inflammatory effect,whereas the effects of PGCN were relatively lower. GML had feeble pharmacodynamic effects. Pharmacokinetic study showed that only cimifugin was detected in the plasma sample of cimifugin and PGCN-treated animals, with drug concentration in the former much higher than the latter. No components were traced in the plasma samples from GML-treated rats. Stability study showed that PGCN and GML was predominantly biotransformed into cimifugin and 5-O-methyvisammiol, respectively. The latter was proven to be extremely unstable in liver tissue homogenate and plasma.Conclusions: A feeble antipyretic, analgesic, and anti-inflammatory activities was observed when GML was orally delivered. Given that Radix Saposhnikoviae extract is generally administered orally, we speculate that this compound might be a nonpharmacolagically active agent in real usage. Thus, it might be unscientific to evaluate the quality of Radix Saposhnikoviae based on the content of GML. Abbreviations used: AUC:area under concentration-time curve, DNP:2,4-Dinitrophenol, HPLC:high performance liquid chromatography, HPLC-MS:high performance liquid chromatography- mass spectrography, GML:4'-O-β-D-glucosyl-5-O-methylvisamminol, MVL:5-O-methyvisammiol, PGCN:prime-O-glucosylcimifugin, SGF:alkaline phosphatase. SIF:simulated intestinal fluid
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Rapid analysis of components in Coptis chinensis Franch by ultra-performance liquid chromatography with quadrupole time-of-flight mass spectrometry p. 175
Peng-peng Tian, Xiao-xu Zhang, Hong-ping Wang, Pu-ling Li, Yu-xin Liu, Shao-jing Li
DOI:10.4103/0973-1296.197635  
Background: Coptis chinensis Franch is a traditional Chinese medical herb.Objective: In this article, ultra-performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry was used to rapidly, qualitatively, and comprehensively identify the components in Coptis chinensis Franch. Materials and Methods: Chromatographic separation was achieved on an Agilent Zorbax RRHD Eclipse Plus C18 column. The mobile phase consisted of 0.1% formic acid water (A) and 0.1% formic acid acetonitrile (B) with a gradient program. Qualitative analysis was performed on an Agilent 6540 quadrupole time-of-flight mass spectrometer, which was equipped with a Dual AJS ESI source operating in negative mode. Results: A total of 30 alkaloid and non-alkaloid components of Coptis chinensis Franch were identified in only 14 min.Conclusion: This study helped to provide a basis for the quality control of Coptis chinensis Franch. Abbreviations used: Q-TOF-MS: quadrupole time-of-flight mass spectrometry, UPLC: ultra-performance liquid chromatography, pos: positive, neg: negative.
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Combinative method using multi-components quantitation and HPLC fingerprint for comprehensive evaluation of Gentiana crassicaulis p. 180
Jiuhua Song, Fengzheng Chen, Jiang Liu, Yuanfeng Zou, Yun Luo, Xiaoyan Yi, Jie Meng, Xingfu Chen
DOI:10.4103/0973-1296.197639  
Background: Gentiana crassicaulis () is an important traditional Chinese herb. Like other herbs, its chemical compounds vary greatly by the environmental and genetic factors, as a result, the quality is always different even from the same region, and therefore, the quality evaluation is necessary for its safety and effective use. In this study, a comprehensive method including HPLC quantitative analysis and fingerprints was developed to evaluate the quality of Cujingqinjiao and to classify the samples collected from Lijiang City of Yunnan province. A total of 30 common peaks including four identified peaks, were found, and were involved for further characterization and quality control of Cujingqinjiao. Twenty-one batches of samples from Lijiang City of Yunnan Province were evaluated by similarity analysis (SA), hierarchical cluster analysis (HCA), principal component analysis (PCA) and factor analysis (FA) according to the characteristic of common peaks.Results: The obtained data showed good stability and repeatability of the chromatographic fingerprint, similarity values were all more than 0.90. This study demonstrated that a combination of the chromatographic quantitative analysis and fingerprint offered an efficient way to quality consistency evaluation of Cujingqinjiao. Consistent results were obtained to show that samples from a same origin could be successfully classified into two groups.Conclusion: This study revealed that the combinative method was reliable, simple and sensitive for fingerprint analysis, moreover, for quality control and pattern recognition of Cujingqinjiao. Abbreviations used: SA: Similarity analysis, HCA: Hierarchical cluster analysis, PCA :Principal component Analysis, FA :Factor analysis
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Green biosynthesis, characterization, in vitro antidiabetic activity, and investigational acute toxicity studies of some herbal-mediated silver nanoparticles on animal models p. 188
Kalakotla Shanker, Gottumukkala Krishna Mohan, Md Ashwaq Hussain, Naradala Jayarambabu, Poka Lakshmi Pravallika
DOI:10.4103/0973-1296.197642  
Diabetes is a metabolic disorder characterized by hyperglycemia, altered carbohydrate, lipid and protein metabolism. In recent studies, Nanoscience and nanotechnology are blazing fields for researchers; for researchers; of late there has been a prodigious excitement in the field of nanopharmacology to study silver nanoparticle (SNP) synthesis using natural products. Biological methods have been used to synthesize SNPs using medicinally active plants having an antidiabetic role, and this made us to assess the biologically synthesized SNPs from the seed extract of Psoralea corylifolia using 1 mM silver nitrate solution. The synthesized herbal&-mediated SNPs (HMSNPs) were subjected to various characterization techniques such as X&-ray diffraction analysis (XRD), energy dispersive X&-ray (EDX) analysis, transmission electron microscope (TEM), and differential light scattering (DLS), respectively. In the current study the HMSNPs were tested to observe the in vitro antidiabetic activity and possible toxic effects in healthy female albino mice by following OECD guidelines&-425. Huge data from biochemical, cellular, mouse, and chemical inhibitor studies have recognized protein tyrosine phosphatase 1B (PTP1B) as a major negative regulator of insulin signaling. In addition, corroboration suggests that insulin action can be enhanced by the inhibition of PTP1B. Keeping in view of the above fact, the PTP1B assay was done to determine the PTP1 B inhibitory effect of HMSNPs. It can be concluded that medicinal plants can be a good source for the synthe sis of HMSNPs. This study can be used for the development of valuable nanomedicines to treat various ailments, and it also highlights the safety and biocompatibility of SNPs within a biological cell; in vivo parameters need to be considered for further discoveries. Abbreviations used: HMSNPs: Herbal mediated silver nanoparticles, XRD: X&-ray diffraction, EDX: Energy dispersive X&-ray analysis, TEM: Transmission electron microscope, PTP1B: Protein tyrosine phosphotase 1B, OECD: Organization for economic cooperation and development
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Quality-by-Design: Multivariate model for multicomponent quantification in refining process of honey p. 193
Xiaoying Li, Zhisheng Wu, Xin Feng, Shanshan Liu, Xiaojie Yu, Qun Ma, Yanjiang Qiao
DOI:10.4103/0973-1296.196310  
Objective: A method for rapid analysis of the refining process of honey was developed based on near-infrared (NIR) spectroscopy. Methods: Partial least square calibration models were built for the four components after the selection of the optimal spectral pretreatment method and latent factors. Results: The models covered the samples of different temperatures and time pointstherefore the models were robust and universal. Conclusions: These results highlighted that the NIR technology could extract the information of critical process and provide essential process knowledge of the honey refining process. Abbreviation used: NIR: Near-infrared; 5-HMF: 5-hydroxymethylfurfural; RMSEP: Root mean square error of prediction; R: correlation coefficients; PRESS: prediction residual error-sum squares; TCM: Traditional Chinese medicine; HPLC: High-performance liquid chromatography; HPLC-DAD: HPLC-diode array detector; PLS: Partial least square; MSC: multiplicative scatter correction; RMSECV: Root mean square error of cross validation; RPD: Residual predictive deviation; 1D: 1st order derivative; SG: Savitzky-Golay smooth; 2D: 2nd order derivative.
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