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   2016| May  | Volume 12 | Issue 46  
    Online since May 11, 2016

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Effect of withinia somnifera and shilajit on alcohol addiction in mice
Priya Bansal, Sugato Banerjee
May 2016, 12(46):121-128
DOI:10.4103/0973-1296.182170  PMID:27279696
Background: Alcohol addiction is a social problem leading to both loss of health and economic prosperity among addicted individuals. Common properties of anti-addictive compounds include anti-anxiety, anticonvulsants, anti-depressant, and nootropic actions primarily through modulation of gamma-aminobutyric acid (GABA) and serotonergic systems. Objective: Here, we screen ashwagandha and shilajit known ethnopharmacologically as nervine tonic and adaptogenic herbs for possible anti-addictive potential. Materials and Methods: Effect of ashwagandha churna and shilajit was measured on ethanol withdrawal anxiety using elevated plus maze. Role of ashwagandha and shilajit on chronic ethanol consumption (21 days) was measured using two bottle choice protocol of voluntary drinking. We also measured the effect of the above herbs on corticohippocampal GABA, dopamine, and serotonin levels. Results: Both ashwagandha and shilajit were found to reduce alcohol withdrawal anxiety in a dose-dependent manner. These herbs alone or in combination also decreased ethanol intake and increased water intake significantly after 21 days of chronic administration. Chronic administration of ashwagandha was found to significantly increase GABA and serotonin levels whereas shilajit altered cortico-hippocampal dopamine in mice. Conclusion: These central nervous system active herbs alone or in combination reduced both alcohol dependence and withdrawal thus showing promising anti-addictive potential. SUMMARY
  • Withinia Somnifera alone and in combination with Shilajeet prevented ethanol withdrawal and alcohol addiction
Abbreviations used: GABA: Gama aminobutyric acid, CNS: Central Nervous System, CPP:Condition place preference, DA: Dopamine, 5-HT: 5-hydroxytryptamine, NMDA:N-methyl-D-aspartate Sugato Banerjee
  5,531 141 -
In vitro and In vivo antioxidant activity of flavonoid extracted from mulberry fruit (Morus alba L.)
Sivakumar Thasma Raman, Ajay Krishna Palani Gounder Ganeshan, Cheng Chen, Chao Jin, Shao-Hui Li, Hui-Juan Chen, Zhongzheng Gui
Apr-Jun 2016, 12(46):128-133
DOI:10.4103/0973-1296.177910  PMID:27076749
Background: Many plants possess antioxidants that exhibit additive or synergistic activities. Objective: In this study, an ethanol-extracted flavonoid extracted from mulberry fruit (FEM) was evaluated for the antioxidant activity in vitro and the hemolysis in red blood cell (RBC) and lipid peroxidation in liver in vivo. Materials and Methods: Antioxidant activities in vitro were measured by quantifying its 2,2-diphenyl-1-picrylhydrazyl (DPPH) scavenging activity, reducing power, and Fe2+-chelating ability. FEM inhibits hemolysis in RBCs and effects of lipid peroxidation in the liver were estimated. Results: The total content of flavonoid compounds was 187.23 mg of quercetin equivalents per grams dried material. In the in vitro assays, FEM demonstrated a strong antioxidant effect, especially in DPPH scavenging activity and reducing power. Mouse RBC hemolysis induced by H2O2was significantly inhibited by FEM in a dose- and time-dependent manner. The effects of FEM on lipid peroxidation in liver, mitochondria, and microsome were investigated. The percentage of inhibition at high concentration (100 μg/mL) of FEM was 45.51%, 39.36%, and 42.78% for liver, mitochondria, and microsomes, respectively. These results suggest that the FEM possesses a strong antioxidant activity both in vivo and in vitro.
  5,457 211 8
Quercetin suppresses the migration and invasion in human colon cancer Caco-2 cells through regulating toll-like receptor 4/Nuclear Factor-kappa B pathway
Mingyang Han, Yucheng Song, Xuedong Zhang
May 2016, 12(46):237-244
DOI:10.4103/0973-1296.182154  PMID:27279714
Objective: The migration and invasion features, which were associated with inflammatory response, acted as vital roles in the development of colon cancer. Quercetin, a bioflavonoid compound, was widely spread in vegetables and fruits. Although quercetin exerts antioxidant and anticancer activities, the molecular signaling pathways in human colon cancer cells remain unclear. Hence, the present study was conducted to investigate the suppression of quercetin on migratory and invasive activity of colon cancer and the underlying mechanism. Materials and Methods: The effect of quercetin on cell viability, migration, and invasion of Caco-2 cells was analyzed by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay, wound-healing assay, and transwell chambers assay, respectively. The protein expressions of toll-like receptor 4 (TLR4), nuclear factor-kappa B (NF-κB) p65, mitochondrial membrane potential-2 (MMP-2), and MMP-9 were detected by Western blot assay. The inflammatory factors, such as tumor necrosis factor-α (TNF-α), cyclooxygenase-2 (Cox-2), and interleukin-6 (IL-6), in cell supernatant were detected by enzyme-linked immunosorbent assay. Results: The concentration of quercetin <20 μM was chosen for further experiments. Quercetin (5 μM) could remarkably suppress the migratory and invasive capacity of Caco-2 cells. The expressions of metastasis-related proteins of MMP-2, MMP-9 were decreased, whereas the expression of E-cadherin protein was increased by quercetin in a dose-dependent manner. Interestingly, the anti-TLR4 (2 μg) antibody or pyrrolidine dithiocarbamate (PDTC; 1 μM) could affect the inhibition of quercetin on cell migration and invasion, as well as the protein expressions of MMP-2, MMP-9, E-cadherin, TLR4, and NF-κB p65. In addition, quercetin could reduce the inflammation factors production of TNF-α, Cox-2, and IL-6. Conclusion: The findings suggested for the 1st time that quercetin might exert its anticolon cancer activity via the TLR4- and/or NF-κB-mediated signaling pathway. SUMMARY
  • Quercetin could remarkably suppress the migratory and invasive capacity of Caco-2 cells
  • The expressions of metastasis-related proteins of mitochondrial membrane potential-2 (MMP-2), MMP-9 were decreased, whereas the expression of E-cadherin protein was increased by quercetin in a dose-dependent manner
  • The anti-toll-like receptor 4 (TLR4) antibody or pyrrolidine dithiocarbamate affected the inhibition of quercetin on cell migration and invasion, as well as the protein expressions of MMP.2, MMP-9, E-cadherin, TLR4, and nuclear factor.kappa B p65
  • Quercetin could reduce the inflammation factors production of tumor necrosis factor-α, cyclooxygenase-2, and interleukin-6.
Abbreviations used: MTT: 3-(4,5-dimethylthiazol-2-yl)- 2,5-diphen yltetrazolium bromide, TLR4: Toll-like receptor 4, NF-κB: Nuclear factor-kappa B, MMP-2: Mitochondrial membrane potential-2, MMP-9: Mitochondrial membrane potential-9, TNF-α: Tumor necrosis factor-α, Cox-2: Cyclooxygenase-2, IL-6: Interleukin-6, ELISA: Enzyme-linked immunosorbent assay, PDTC: Pyrrolidine dithiocarbamate, ROS: Reactive oxygen species, DMSO: Dimethyl sulfoxide, FBS: Fetal bovine serum, DMEM: Dulbecco modified Eagle medium, OD: Optical density, IPP: Image Pro-plus, PBS: Phosphate buffered saline, SD: Standard deviation, ANOVA: One-way analysis of variance, SPSS: Statistical Package for the Social Sciences, ECM: Extracellular matrix, TLRs: Toll-like receptors, LPS: Lipopolysaccharide.
  3,795 131 1
Quantitative analysis and In vitro anti-inflammatory effects of gallic acid, ellagic acid, and quercetin from radix sanguisorbae
Chang-Seob Seo, Soo-Jin Jeong, Sae-Rom Yoo, Na-Ri Lee, Hyeun-Kyoo Shin
Apr-Jun 2016, 12(46):104-108
DOI:10.4103/0973-1296.177908  PMID:27076745
Background: Radix Sanguisorbae has long been used to treat diarrhea, enteritis, duodenal ulcers, and internal hemorrhage. Objective: We investigated the in vitro anti-inflammatory effects of Radix Sanguisorbae and performed quantitative analyses of three marker components, namely gallic acid, ellagic acid, and quercetin, using high-performance liquid chromatography coupled with a photodiode array detector. Materials and Methods: The three marker components were separated using a reversed-phase Gemini C18 analytical column maintained at 40°C by the gradient elution with two solvent systems. We examined the biological effects of the three marker compounds, gallic acid, ellagic acid, and quercetin, by determining their anti-inflammatory activities in the murine macrophage cell line RAW 264.7. Results: All of the marker compounds exhibited inhibitory effects on prostaglandin E2 production in lipopolysaccharide (LPS)-stimulated RAW 264.7 macrophages, with no cytotoxicity. Particularly, ellagic acid significantly inhibited production of the proinflammatory cytokines tumor necrosis factor alpha and interleukin-6 in LPS-treated RAW 264.7 cells. Conclusion: Our results suggest that ellagic acid is the most potent bioactive phytochemical component of radix Sanguisorbae in the treatment of inflammatory diseases.
  3,666 122 4
Flavonoids derived from Abelmoschus esculentus attenuatesUV-B Induced cell damage in human dermal fibroblasts throughNrf2-ARE pathway
Juilee Patwardhan, Purvi Bhatt
May 2016, 12(46):129-138
DOI:10.4103/0973-1296.182175  PMID:27279697
Background: Ultraviolet-B (UV-B) radiation is a smaller fraction of the total radiation reaching the Earth but leads to extensive damage to the deoxyribonucleic acid (DNA) and other biomolecules through formation of free radicals altering redox homeostasis of the cell. Abelmoschus esculentus (okra) has been known in Ayurveda as antidiabetic, hypolipidemic, demulscent, antispasmodic, diuretic, purgative, etc. Objective: The aim of this study is to evaluate the protective effect of flavonoids from A. esculentus against UV-B-induced cell damage in human dermal fibroblasts. Materials and Methods: UV-B protective activity of ethyl acetate (EA) fraction of okra was studied against UV-B-induced cytotoxicity, antioxidant regulation, oxidative DNA damage, intracellular reactive oxygen species (ROS) generation, apoptotic morphological changes, and regulation of heme oxygenase-1 (HO-1) gene through nuclear factor E2-related factor 2-antioxidant response element (Nrf2-ARE) pathway. Results: Flavonoid-rich EA fraction depicted a significant antioxidant potential also showing presence of rutin. Pretreatment of cells with EA fraction (10–30 μg/ml) prevented UV-B-induced cytotoxicity, depletion of endogenous enzymatic antioxidants, oxidative DNA damage, intracellular ROS production, apoptotic changes, and overexpression of Nrf2 and HO-1. Conclusion: Our study demonstrated for the 1st time that EA fraction of okra may reduce oxidative stress through Nrf2-ARE pathway as well as through endogenous enzymatic antioxidant system. These results suggested that flavonoids from okra may be considered as potential UV-B protective agents and may also be formulated into herbal sunscreen for topical application. SUMMARY
  • Flavonoid.enriched ethyl acetate.(EA) fraction from A. esculentus protected against ultraviolet.B.(UV.B).induced oxidative DNA damage
  • EA fraction prevented UV-B-induced cytotoxicity, depletion of endogenous enzymatic antioxidants, and intracellular reactive oxygen species production
  • EA fraction could reduce oxidative stress through the Nrf2-.ARE Pathway
  • EA fraction was found to be nongenotoxic and prevented apoptotic changes.
  • Flavonoids from Abelmoschus esculentus protected from ultraviolet.B.induced damage
  • They were capable of reducing oxidative stress through Nrf2-.ARE Pathway
  • They are nongenotoxic and do not possess mutagenic potential
  • Flavonoids from A. esculentus can be studied and explored further for its topical application as sunscreen.
Abbreviations used:ABTS: 2,2'-azino-bis-(3-ethylbenzothiazoline -6-sulphonic acid), AO: Acridine orange, ANOVA: Analysis of variance, ARE: Antioxidant response elements, BSA: Bovine serum albumin, CAPE: Caffeic acid phenethyl ester, CAT: Catalase, DCFH-DA: 2',7'-dichlorofluorescein diacetate, DMEM: Dulbecco's modified eagle's medium, DMSO: dimethyl sulfoxide, DNA: Deoxyribonucleic acid, DPBS: Dulbecco's phosphate-buffered saline, DPPH: 2,2-diphenyl-1-picryl hydrazyl, ECL: Enhanced chemiluminescence, EDTA: Ethylenediaminetetraacetic acid, ELISA: Enzyme-linked immunosorbent assay, EtBr: Ethidium bromide, FBS: Fetal bovine serum, FE Fraction: Flavonoid-enriched fraction, FRAP: Ferric reducing antioxidant power, GPx: Glutathione peroxidase, GR: Glutathione reductase, GST: Glutathione-S-transferase, GSH: Reduced glutathione, GSSG: Oxidized glutathione, HDF: Human dermal fibroblast adult cells, HEPES: 4-(2-hydroxyethyl)-1-piperazineethanesulphonic acid, HRP: Horseradish peroxidase, HO-1: Heme oxygenase-1, HPTLC: High-performance thin layer chromatography, Keap-1: Kelch-like ECH-associated protein-1, MTT: 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide, NaCl: sodium chloride, NFDM: nonfat dry milk, Nrf2: Nuclear factor E2-related factor 2, NQO1: NAD (P) H: Quinine oxidoreductase 1, OH: Hydroxyl ions, PBST: Phosphate-buffered saline with 0.1% tween 20, PCR: Polymerase chain reaction, PMSF: Phenylmethanesulfonyl fluoride, Rf: Retention factor, ROS: Reactive oxygen species, rRNA: Ribosomal ribonucleic acid, SDS: Sodium dodecyl sulfate, SOD: Superoxide dismutase, TLC: Thin layer chromatography, TLC-DPPH: Thin layer chromatography-2,2-diphenyl-1-picryl hydrazyl, UV: Ultraviolet, UV-A: Ultraviolet-A, UV-B: Ultraviolet-B, UV-C: Ultraviolet-C, qPCR: Quantitative polymerase chain reaction
  3,402 126 2
Ketosteroid standardized Cissus quadrangularis L. extract and its anabolic activity: Time to look beyond ketosteroid?
Atul N Jadhav, Mohammed Rafiq, Rajendran Devanathan, Mohammed Azeemuddin, Suryakanth D Anturlikar, Akhil Ahmed, Ramchandran Sundaram, UV Babu, Rangesh Paramesh
May 2016, 12(46):213-217
DOI:10.4103/0973-1296.182177  PMID:27279709
Background: Cissus quadrangularis (CQ) L. reported to contain 3-ketosteroids and have bone health benefits. Aim: This study aimed at establishing the relationship between the ketosteroid content and anabolic as well as bone health-promoting activities of various Cissus extracts in well-established orchidectomized (ORX) rat model. Materials and Methods: Supercritical carbon dioxide, ethyl acetate, and aqueous extracts (AE) of CQ L. were prepared and standardized for ketosteroid content by two methods used in commerce. Moreover, ketosteroid standardized extracts of this plant were evaluated for anabolic activity in rats in well-established ORX rat model. Results: The increase in the absolute weight was appreciable in the CQ-AE treated group. Similarly, with respect to bone parameters, a similar trend was seen. The mean bone density, strength, and calcium content were found to be highest in the group treated with CQ-AE compared to groups treated with other extracts. This study reveals for the first time that 3-ketosteroids are not linked to the beneficial activities on bone and highlights the need for extensive characterization of biological active principles from CQ L. Conclusion: In light of the above estimation studies, we believe that current standardization of Cissus extraction “3-ketosteroids” is incorrect. We also did not find any report suggesting the presence of androgenic steroids in this plant and hence the characterization based on “3-ketosteroids” is scientifically incorrect. This study highlights the insufficient understanding of biological active principles from CQ L. and underlines the need for extensive bioactivity guided studies. SUMMARY
  • Cissus quadrangularis (CQ) L. reported to contain 3-ketosteroids and have bone health benefits
  • We did not find correlation between ketosteroid content obtained by conventional methods and its biological effect
  • Studies indicate that claims of ketosteroid content need not necessarily correlate to biological effects and hence warrants extensive phytochemical characterization of biological active principles from CQ L.
Abbreviations used: CQ: Cissus quadrangularis, ORX: Orchidectomized, AE: Aqueous extract, EE: Ethyl acetate extract, SFE: Supercritical fluid extract. Atul N. Jadhav
  3,375 128 -
Elicitation based enhancement of secondary metabolites in Rauwolfia serpentina and Solanum khasianum hairy root cultures
Mrinalini Srivastava, Swati Sharma, Pratibha Misra
April 2016, 12(46):315-320
DOI:10.4103/0973-1296.185726  PMID:27563218
Background: Rauwolfia serpentina and Solanum khasianum are well-known medicinally important plants contained important alkaloids in their different parts. Elicitation of these alkaloids is important because of associated pharmaceutical properties. Targeted metabolites were ajmaline and ajmalicine in R. serpentina; solasodine and α-solanine in S. khasianum. Objective: Enhancement of secondary metabolites through biotic and abiotic elicitors in hairy root cultures of R. serpentina and S. khasianum. Materials and Methods: In this report, hairy root cultures of these two plants were established through Agrobacterium rhizogenes mediated transformation by optimizing various parameters as age of explants, duration of preculture, and co-cultivation period. NaCl was used as abiotic elicitors in these two plants. Cellulase from Aspergillus niger was used as biotic elicitor in S. khasianum and mannan from Saccharomyces cerevisiae was used in R. serpentina. Results: First time we have reported the effect of biotic and abiotic elicitors on the production of important metabolites in hairy root cultures of these two plants. Ajmalicine production was stimulated up to 14.8-fold at 100 mM concentration of NaCl after 1 week of treatment. Ajmaline concentration was also increased 2.9-fold at 100 mg/l dose of mannan after 1 week. Solasodine content was enhanced up to 4.0-fold and 3.6-fold at 100 mM and 200 mM NaCl, respectively, after 6 days of treatments. Conclusion: This study explored the potential of the elicitation strategy in A. rhizogenes transformed cell cultures and this potential further used for commercial production of these pharmaceutically important secondary metabolites.
  3,231 221 -
Kinetics of inhibition of monoamine oxidase using curcumin and ellagic acid
Dharmendra Kumar Khatri, Archana Ramesh Juvekar
May 2016, 12(46):116-120
DOI:10.4103/0973-1296.182168  PMID:27279695
Background: Curcumin and ellagic are the natural polyphenols having a wide range of pharmacological actions. They have been reported to have their use in various neurological disorders. Objective: This study was aimed to evaluate the effect of curcumin and ellagic acid on the activity of monoamine oxidase (MAO), the enzyme responsible for metabolism of monoamine neurotransmitters which are pivotal for neuronal development and function. Materials and Methods: The in vitro effects of these selected polyphenols on MAO activities in mitochondria isolated from rat brains were examined. Brain mitochondria were assayed for MAO type-B (MAO-B) using benzylamine as substrates. Rat brain mitochondrial MAO preparation was used to study the kinetics of enzyme inhibition using double reciprocal Lineweaver–Burk plot. Results: MAO activity was inhibited by curcumin and ellagic acid; however, higher half maximal inhibitory concentrations of curcumin (500.46 nM) and ellagic acid (412.24 nM) were required compared to the known MAO-B inhibitor selegiline. It is observed that the curcumin and ellagic acid inhibit the MAO activity with both the competitive and noncompetitive type of inhibitions. Conclusions: Curcumin and ellagic acid can be considered a possible source of MAO inhibitor used in the treatment of Parkinson's and other neurological disorders. SUMMARY
  • Monoamine oxidase (MAO) is involved in a variety of neurological disorders including Parkinson's disease (PD)
  • Curcumin and ellagic acid inhibit the monoamine oxidase activity
  • Ellagic acid revealed more potent MAO type-B (MAO-B) inhibitory activity than curcumin
  • Kinetic studies of MAO inhibition using different concentrations of curcumin and ellagic acid were plotted as double reciprocal Lineweaver–Burk plot
  • The mode of inhibition of both compounds toward MAO-B is mixed (competitive and uncompetitive) type of inhibition with both the competitive and noncompetitive type of inhibitions.
Abbreviations used: MAO: Monoamine oxidase, IC50: Higher half maximal inhibitory concentrations, PD: Parkinson's disease, LB: Lewy bodies, SNpc: Substantia nigra pars compacta, ROS: Reactive oxygen species, SG: Selegiline, DMC: demethoxycurcumin, BDMC: Bisdemethoxycurcumin. Archana Ramesh Juvekar
  3,288 100 1
Application of ultra-performance liquid chromatography with time-of-flight mass spectrometry for the rapid analysis of constituents and metabolites from the extracts of Acanthopanax senticosus Harms leaf
Yingzhi Zhang, Aihua Zhang, Ying Zhang, Hui Sun, Xiangcai Meng, Guangli Yan, Xijun Wang
Apr-Jun 2016, 12(46):145-152
DOI:10.4103/0973-1296.177902  PMID:27076752
Acanthopanax senticosus (Rupr and Maxim) Harms (AS), a member of Araliaceae family, is a typical folk medicinal herb, which is widely distributed in the Northeastern part of China. Due to lack of this resource caused by the extensive use of its root, this work studied the chemical constituents of leaves of this plant with the purpose of looking for an alternative resource. In this work, a fast and optimized ultra-performance liquid chromatography method with quadrupole time-of-flight mass spectrometry (UPLC-QTOF-MS) has been developed for the analysis of constituents in leaves extracts. A total of 131 compounds were identified or tentatively characterized including triterpenoid saponins, phenols, flavonoids, lignans, coumarins, polysaccharides, and other compounds based on their fragmentation behaviors. Besides, a total of 21 metabolites were identified in serum in rats after oral administration, among which 12 prototypes and 9 metabolites through the metabolic pathways of reduction, methylation, sulfate conjugation, sulfoxide to thioether and deglycosylation. The coupling of UPLC-QTOF-MS led to the in-depth characterization of the leaves extracts of AS both in vitro and in vivo on the basis of retention time, mass accuracy, and tandem MS/MS spectra. It concluded that this analytical tool was very valuable in the study of complex compounds in medicinal herb.
  3,270 99 -
Quality analysis of chlorogenic acid and hyperoside in Crataegi fructus
Jin Bae Weon, Youn Sik Jung, Choong Je Ma
Apr-Jun 2016, 12(46):98-103
DOI:10.4103/0973-1296.177904  PMID:27076744
Background: Crataegi fructus is a herbal medicine for strong stomach, sterilization, and alcohol detoxification. Chlorogenic acid and hyperoside are the major compounds in Crataegi fructus. Objective: In this study, we established novel high-performance liquid chromatography (HPLC)-diode array detection analysis method of chlorogenic acid and hyperoside for quality control of Crataegi fructus. Materials and Methods: HPLC analysis was achieved on a reverse-phase C18 column (5 μm, 4.6 mm × 250 mm) using water and acetonitrile as mobile phase with gradient system. The method was validated for linearity, precision, and accuracy. About 31 batches of Crataegi fructus samples collected from Korea and China were analyzed by using HPLC fingerprint of developed HPLC method. Then, the contents of chlorogenic acid and hyperoside were compared for quality evaluation of Crataegi fructus. Results: The results have shown that the average contents (w/w %) of chlorogenic acid and hyperoside in Crataegi fructus collected from Korea were 0.0438% and 0.0416%, respectively, and the average contents (w/w %) of 0.0399% and 0.0325%, respectively. Conclusion: In conclusion, established HPLC analysis method was stable and could provide efficient quality evaluation for monitoring of commercial Crataegi fructus.
  3,005 89 -
Preparation and evaluation of andrographolide solid dispersion vectored by silicon dioxide
Dingkun Zhang, Junzhi Lin, Fang Zhang, Xue Han, Li Han, Ming Yang, Wenquan Zou
May 2016, 12(46):245-252
DOI:10.4103/0973-1296.182156  PMID:27279715
Background: Andrographolide (Andro) is a “natural antibiotic” as well as a typical insoluble drug. The purpose of this study was to investigate the feasibility of commercially available silica (SiO2) as a carrier of solid dispersion to enhance the dissolution of Andro. Materials and Methods: The solvent evaporation method was adopted, and a series of process parameters were studied to prepare a solid dispersion. Andro, SiO2, physical mixture, and solid dispersion were characterized with respect to particle size distribution, special surface area, pore volume, and scanning electron microscopy, Fourier transform infrared spectroscopy, and X-ray diffraction studies. Results: Single factor test suggested the best preparation of solid dispersion was the drug and carrier (SiO2B) ratio of 1:8, with tetrahydrofuran as the solvent, and a recovery temperature of 50°C. Compared to crude drug and mixture, solid dispersion was found to form a unique structure to disperse the drug and displayed superior performance in rapid dissolution. Conclusion: The present study signifies the commercially available SiO2is an excellent but cheap carrier to improve the dissolution of Andro. Our results provide a highly operability approach for improving the dissolution of insoluble natural products and are beneficial for the clinical effects improvement. SUMMARY
  • The potential of commercially available silica as a carrier for enhancing the insoluble drug dissolution was investigated
  • Factors affecting the dissolution of solid dispersion were investigated
  • Solid dispersion formed a unique structure to disperse the drug and release drug rapidly
  • Commercially available silica is an excellent but cheap carrier to improve the dissolution of Andro.
Abbreviation used: Andro: Andrographolide, BCS: Biopharmaceutics Classification System, SDS: Tetrahydrofuran and Sodium dodecyl sulfate, HPLC: High Performance Liquid Chromatography, SEM: Scanning Electron Microscope, BET: Brumauer–Emmett–Teller, FTIR: Fourier Transform Infrared Spectroscopy, XRD: X-ray Diffraction.
  2,955 116 2
Evaluation of xanthine oxidase inhibitory potential and In vivo hypouricemic activity of Dimocarpus longan lour. extracts
Shi-Yuan Sheu, Yuan-Tsung Fu, Wen-Dar Huang, Yung-Ann Chen, Yi-Chih Lei, Chun-Hsu Yao, Feng-Lin Hsu, Tzong-Fu Kuo
May 2016, 12(46):206-212
DOI:10.4103/0973-1296.182176  PMID:27279708
Background: Longan is a fruit tree known to contain many phenolic components, which are capable of protecting people from oxidative damage through an anti-inflammatory mechanism. It may be also worthwhile to study the effect on lowering uric acid activity. Materials and Methods: This study investigates the lowering of uric acid using longan extracts, including flowers, pericarps, seeds, leaves, and twigs, on potassium-oxonate-induced hyperuricemia mice and its inhibitory actions against xanthine oxidase (XO) activities. Results: The findings revealed that ethyl acetate fraction of longan extracts exhibited strong XO-inhibitory activity, and the flower extracts (IC50 = 115.8 μg/mL) revealed more potent XO-inhibitory activity to those of pericarps (118.9 μg/mL), twigs (125.3 μg/mL), seeds (262.5 μg/mL), and leaves (331.1 μg/mL) in vitro. In addition, different dosages of longan extract (50–100 mg/kg) were administered to hyperuricemic mice. The lowering effect of longan extracts on uric acid at 75 mg/kg markedly reduced plasma uric acid levels in decreasing order: Flowers (80%) > seeds (72%) > pericarps (64%) > twigs (59%) > leaves (41%), compared with allopurinol (89%). Finally, 10 isolated phytochemicals from longan flowers were then examined in vitro. The results indicated that proanthocyanidin A2 and acetonylgeraniin A significantly inhibited XO activity in vitro. This is the first report providing new insights into the urate-reducing effect of phenolic dimer and hydrolyzable tannin, which can be developed to potential hypouricemic agents. SUMMARY
  • Longan flower extracts possess more potent XO-.inhibitory activity than pericarps, twigs, seeds, and leaves in vitro
  • The lowering effect of longan flowers and seeds extracts markedly reduced plasma uric acid levels as compared to allopurinol in vivo
  • The extract proanthocyanidin A2 and acetonylgeraniin A were demonstrated potent XO inhibitory activity in vitro.
Abbreviations used: PO: Potassium-oxonate, XO: xanthine oxidase, HE: n-hexane, EA: ethyl acetate, i.p.: intraperitoneal, PBS: phosphate-buffered saline, AP: allopurinol, PUA: plasma uric acid.
  2,935 114 -
Studies on the red sea sponge Haliclona sp. for its chemical and cytotoxic properties
Shaza Mohamed Al-Massarani, Ali Ali El-Gamal, Mansour Sulaiman Al-Said, Maged S Abdel-Kader, Abdelkader E Ashour, Ashok Kumar, Wael M Abdel-Mageed, Adnan Jathlan Al-Rehaily, Hazem A Ghabbour, Hoong-Kun Fun
Apr-Jun 2016, 12(46):114-119
DOI:10.4103/0973-1296.177906  PMID:27076747
Background: A great number of novel compounds with rich chemical diversity and significant bioactivity have been reported from Red Sea sponges. Objective: To isolate, identify, and evaluate the cytotoxic activity of the chemical constituents of a sponge belonging to genus Haliclona collected from the Eastern coast of the Red Sea. Materials and Methods: The total ethanolic extract of the titled sponge was subjected to intensive chromatographic fractionation and purification guided by cytotoxic bioassay toward various cancer cell lines. The structures of the isolated compounds were elucidated using spectroscopic techniques including one-dimension and two-dimension nuclear magnetic resonance, mass spectrometry, ultraviolet, and infrared data, as well as comparison with the reported spectral data for the known compounds. X-ray single-crystal structure determination was performed to determine the absolute configuration of compound 4. The screening of antiproliferative activity of the compounds was carried on three tumor cell lines, namely the human cervical cancer (HeLa), human hepatocellular carcinoma (HepG2), and human medulloblastoma (Daoy) cells using MTT assay. Results: This investigation resulted in the isolation of a new indole alkaloid, 1-(1H-indol-3-yloxy) propan-2-ol (1), with the previously synthesized pyrrolidine alkaloid, (2R, 3S, 4R, 5R) pyrrolidine-(1-hydroxyethyl)-3,4-diol hydrochloride (4), isolated here from a natural source for the first time. In addition, six known compounds tetillapyrone (2), nortetillapyrone (3), 2-methyl maleimide-5-oxime (5), maleimide-5-oxime (6), 5-(hydroxymethyl) dihydrofuran-2 (3H)-one (7), and ergosta-5,24 (28)-dien-3-ol (8) were also identified. Most of the isolated compounds exhibited weak cytotoxic activity against HepG-2, Daoy, and HeLa cancer cell lines. Conclusion: This is the first report of the occurrence of the indole and pyrrolidine alkaloids, 1-(1H-indol-2-yloxy) propan-2-ol (1), and the - (1-hydroxyethyl)-3,4-diol hydrochloride (4), in the Red Sea Haliclona sp.
  2,827 132 1
Effect of procyanidin-rich extract from natural cocoa powder on cellular viability, cell cycle progression, and chemoresistance in human epithelial ovarian carcinoma cell lines
Shruti Taparia, Aparna Khanna
May 2016, 12(46):109-115
DOI:10.4103/0973-1296.182164  PMID:27279694
Background: Over the last 400 years, cocoa and chocolate have been described as having potential medicinal value, being consumed as a beverage or eaten as food. Concentration–dependant, antiproliferation, and cytotoxic effects of some of their polyphenolic constituents have been demonstrated against various cancers. Such an effect remains to be demonstrated in ovarian cancer Objective: To investigate the effect of cocoa procyanidins against ovarian cancer in vitro using OAW42 and OVCAR3 cell lines. Materials and Methods: Cocoa procyanidins were extracted and enriched from non alkalized cocoa powder. The polyphenolic content and antioxidant activity were determined. Effect on cell viability was determined after the treatment with ≤1000 μg/mL cocoa procyanidin-rich extract on OAW42 and OVCAR3 and normal human dermal fibroblasts. Similarly, chemosensitization effect was determined by pretreating cancer cell lines with extract followed by doxorubicin hydrochloride treatment. The effect of treatment on cell cycle and P-glycoprotein (P-gp) expression was determined using flow cytometry. Results: The cocoa extract showed high polyphenolic content and antioxidant activity. Treatment with extract caused cytotoxicity and chemosensitization in OAW42 and OVCAR3 cell lines. Normal dermal fibroblasts showed an increase in cell viability post treatment with extract. Treatment with extract affected the cell cycle and an increasing percentage of cells in hypodiploid sub-G1/G0phase was observed. Treatment of OVCAR3 with the extract caused reduction of P-gp expression. Conclusion: Cocoa procyanidins were found to be selectively cytotoxic against epithelial ovarian cancer, interfered with the normal cell cycle and sensitized cells to subsequent chemotherapeutic treatment. Chemosensitization was found to be associated with P-gp reduction in OVCAR3 cells. SUMMARY
  • Among the naturally occurring flavonoids, procyanidins have been shown to be effective against cancers
  • Non alkalized cocoa powder is one of the richest sources of procyanidins
  • Cocoa procyanidin.rich extract (CPRE) caused cytotoxicity and chemosensitization in ovarian carcinoma cell lines OAW42 and OVCAR3
  • CPRE affected normal cell cycle progression.
  • CPRE also downregulated P.glycoprotein, which mediates chemoresistance in multidrug.resistant OVCAR3 cell line.
Abbreviation used: P-gp : P-glycoprotein, CPRE: Cocoa procyanidin rich extract, DMAC: 4-dimethylaminocinnamaldehyde, DPPH: Diphenylpicrylhydrazyl, ABTS: 2,2'-azinobis-(3-ethylbenzothiazoline-6-sulfonic acid), PI: Propidium iodide, FITC: Fluorescein isothiocyanate, MTT: 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide, TLC: Thin layer chromatography, HPTLC: High-performance thin layer chromatography. Aparna Khanna
  2,823 101 -
Protective effects of silymarin, alone or in combination with chlorogenic acid and/or melatonin, against carbon tetrachloride-induced hepatotoxicity
Nouf Al-Rasheed, Laila Faddah, Nawal Al-Rasheed, Yieldez A Bassiouni, Iman H Hasan, Ayman M Mahmoud, Raeesa A Mohamad, Hazar I Yacoub
April 2016, 12(46):337-345
DOI:10.4103/0973-1296.185765  PMID:27563222
Objective: The aim of this study was to evaluate the hepatoprotective effects of silymarin (SIL), alone and combined with chlorogenic acid (CA) and/or melatonin (ME), using a rat model of carbon tetrachloride (CCl4)-induced injury. Materials and Methods: Hepatotoxicity was induced by a single dose of CCl4 (1 ml/kg, IP). One day after, rats were received SIL (200 mg/kg) alone or in combination with CA (60 mg/kg) and/or ME (20 mg/kg) for 21 days. Results: SIL significantly decreased serum alanine aminotransferase, inflammatory cytokines, and vascular endothelial growth factor levels. Histological alterations, fibrogenesis, oxidative DNA damage, inflammatory mediators, and caspase-3 activity were significantly attenuated in SIL treated CCl4-intoxicated rats. On the other hand, cytochrome P450 2E1 activity showed a significant decrease in the liver of CCl4-intoxicated rats, an effect that was reversed following treatment with SIL. All beneficial effects of SIL were markedly potentiated when combined with CA and/or ME. Conclusions: These data indicate that SIL, alone and combined with CA and/or ME, protected the liver against CCl4-induced hepatotoxicity via attenuating inflammation, oxidative DNA damage, apoptosis, and fibrotic changes. The significantly intensified hepatoprotective effects of SIL when combined with both CA and ME suggest a possible synergism. These synergistic effects need to be further confirmed using detailed studies.
  2,647 166 -
Identification of medicinal Mugua origin by near infrared spectroscopy combined with partial least-squares discriminant analysis
Bangxing Han, Huasheng Peng, Hui Yan
Apr-Jun 2016, 12(46):93-97
DOI:10.4103/0973-1296.177907  PMID:27076743
Background: Mugua is a common Chinese herbal medicine. There are three main medicinal origin places in China, Xuancheng City Anhui Province, Qijiang District Chongqing City, Yichang City, Hubei Province, and suitable for food origin places Linyi City Shandong Province. Objective: To construct a qualitative analytical method to identify the origin of medicinal Mugua by near infrared spectroscopy (NIRS). Materials and Methods: Partial least squares discriminant analysis (PLSDA) model was established after the Mugua derived from five different origins were preprocessed by the original spectrum. Moreover, the hierarchical cluster analysis was performed. Results: The result showed that PLSDA model was established. According to the relationship of the origins-related important score and wavenumber, and K-mean cluster analysis, the Muguas derived from different origins were effectively identified. Conclusion: NIRS technology can quickly and accurately identify the origin of Mugua, provide a new method and technology for the identification of Chinese medicinal materials.
  2,710 63 -
In vitro cytotoxicity and anti-herpes simplex virus type 1 activity of hydroethanolic extract, fractions, and isolated compounds from stem bark of Schinus terebinthifolius Raddi
Samara Requena Nocchi, Gislaine Franco de Moura-Costa, Claudio Roberto Novello, Juliana Rodrigues, Renata Longhini, João Carlos Palazzo de Mello, Benedito Prado Dias Filho, Celso Vataru Nakamura, Tânia Ueda-Nakamura
Apr-Jun 2016, 12(46):160-164
DOI:10.4103/0973-1296.177903  PMID:27076754
Background: Herpes simplex virus type 1 (HSV-1) is associated with orofacial infections and is transmitted by direct contact with infected secretions. Several efforts have been expended in the search for drugs to the treatment for herpes. Schinus terebinthifolius is used in several illnesses and among them, for the topical treatment of skin wounds, especially wounds of mucous membranes, whether infected or not. Objective: To evaluate the cytotoxicity and anti-HSV-1 activity of the crude hydroethanolic extract (CHE) from the stem bark of S. terebinthifolius, as well as its fractions and isolated compounds. Materials and Methods: The CHE was subjected to bioguided fractionation. The anti-HSV-1 activity and the cytotoxicity of the CHE, its fractions, and isolated compounds were evaluated in vitro by SRB method. A preliminar investigation of the action of CHE in the virus–host interaction was conducted by the same assay. Results: CHE presented flavan-3-ols and showed anti-HSV-1 activity, better than its fractions and isolated compounds. The class of substances found in CHE can bind to proteins to form unstable complexes and enveloped viruses, as HSV-1 may be vulnerable to this action. Our results suggest that the CHE interfered with virion envelope structures, masking viral receptors that are necessary for adsorption or entry into host cells. Conclusion: The plant investigated exhibited potential for future development treatment against HSV-1, but further tests are necessary, especially to elucidate the mechanism of action of CHE, as well as preclinical and clinical studies to confirm its safety and efficacy.
  2,674 81 1
Chromatographic and spectrophotometric analysis of phenolic compounds from fruits of Libidibia ferrea Martius
Magda R. A. Ferreira, Mônica T. M. Fernandes, Wliana A. V. da Silva, Isabelle C. F. Bezerra, Tatiane P de Souza, Maria F Pimentel, Luiz A. L. Soares
May 2016, 12(46):285-291
DOI:10.4103/0973-1296.182165  PMID:27279721
Background: Libidibia ferrea (Mart. ex Tul.) L.P. Queiroz (Fabaceae) is a tree which is native to Brazil, widely known as “Jucá,” where its herbal derivatives are used in folk medicine with several therapeutic properties. The constituents, which have already been described in the fruit, are mainly hydrolysable tannins (gallic acid [GA] and ellagic acid emailpage.asp). Objective: The aim of this study was to investigate the phenolic variability in the fruit of L. ferrea by ultraviolet/visible (UV/VIS) and chromatographic methods (high-performance liquid chromatography [HPLC]/high-performance thin layer chromatography [HPTLC]). Materials and Methods: Several samples were collected from different regions of Brazil and the qualitative (fingerprints by HPTLC and HPLC) and quantitative analysis (UV/VIS and HPLC) of polyphenols were performed. Results: The HPTLC and HPLC profiles allowed separation and identification of both major analytical markers: EA and GA. The chemical profiles were similar in a number of spots or peaks for the samples, but some differences could be observed in the intensity or area of the analytical markers for HPTLC or HPLC, respectively. Regarding the quantitative analysis, the polyphenolic content by UV/VIS ranged from 13.99 to 37.86 g% expressed as GA or from 10.75 to 29.09 g% expressed as EA. The contents of EA and GA by liquid chromatography-reversed phase (LC-RP) method ranged from 0.57 to 2.68 g% and from 0.54 to 3.23 g%, respectively. Conclusion: The chemical profiles obtained by HPTLC or HPLC, as well as the quantitative analysis by spectrophotometry or LC-RP method, were suitable for discrimination of each herbal sample and can be used as tools for the comparative analysis of the fruits from L. ferrea. SUMMARY
  • The polyphenols of fruits of Libidibia ferrea can be quantified by UV/VIS and HPLC
  • The HPLC method was able to detect the gallic and ellagic acids in several samples of fruits of Libidibia ferrea
  • The phenolic profiles of fruits from Libidibia ferrea by HPTLC and HPLC were reproductible.
Abbreviations used: HPTLC: high performance thin layer chromatography, HPLC: high performance liquid chromatography, UV-Vis: spectrophotometry
  2,637 87 1
Assessment of the polyphenolic content, free radical scavenging, anti-inflammatory, and antimicrobial activities of acetone and aqueous extracts of Lippia javanica (Burm.F.) spreng
Abiola M Asowata-Ayodele, Gloria A Otunola, Anthony J Afolayan
April 2016, 12(46):353-362
DOI:10.4103/0973-1296.185770  PMID:27563225
Background: Lippia javanica (Burm.F.) Spreng is one of the spice plants commonly found in almost every part of South Africa. Apart from its culinary uses, it is also traditionally used as an insect repellant and infusion for fever, flu, kidney stone treatment, cough, common cold, and chest pain. Materials and Methods: The antioxidant activities of the aqueous and acetone extracts were determined by measuring their effects against 1,1-Diphenyl-2-picryl-hydrazyl, 2,2'azino-bis (3-ethylbenzthiazoline-6-sulfonic acid), nitric oxide, phosphomolybdate, lipid peroxidation, hydrogen peroxide, and reducing power. The antimicrobial activities were evaluated against four bacterial (two Gram-positive, two Gram-negative) strains and 9 fungal pathogens using the agar well diffusion and microdilution methods. Anti-inflammatory activity was assessed by determining the inhibition against protein denaturation and membrane stabilizing effects. Objective: The polyphenolic content, free radical scavenging, anti-inflammatory, and antimicrobial activities of the aqueous and acetone extracts of the plant were evaluated. Results: A significantly high total phenolic content and free radical scavenging activities were observed in the acetone extracts of the plants. The study also revealed a concentration-dependent inhibition of protein denaturation and membrane stabilization effects by both the aqueous and acetone extracts at the concentrations studied. The ability of L. javanica extracts to inhibit protein denaturation and maintain membrane stability could be responsible for its folkloric use. The overall antimicrobial activity indicates that both extracts were active against the bacterial strains but the acetone extract exhibited the most potent antifungal activity higher than even the reference drugs. Conclusion: Overall, the acetone extract of L. javanica exhibited a more pronounced antioxidant, anti-inflammatory, and antimicrobial effects than the aqueous extract.
  2,546 146 1
In vitro screening and evaluation of 37 traditional chinese medicines for their potential to activate peroxisome proliferator-activated receptors-γ
Die Gao, Yonglan Zhang, Fengqing Yang, Yexin Lin, Qihui Zhang, Zhining Xia
Apr-Jun 2016, 12(46):120-127
DOI:10.4103/0973-1296.177909  PMID:27076748
Background: Peroxisome proliferator-activated receptors (PPAR)-γ is widely used as an attractive target for the treatment of type 2 diabetes mellitus. Thiazolidinediones, the agonists of PPARγ, has been popularly utilized as insulin sensitizers in the therapy of type 2 diabetes whereas numerous severe side-effects may also occur concomitantly. Objective: The PPARγ activation activity of different polar extracts, including petroleum ether, ethyl acetate, n-butanol, residual of ethanol, the precipitate part of water and the supernatant of water extracts, from 37 traditional Chinese medicines were systematically evaluated. Materials and Methods: HeLa cells were transiently co-transfected with the re-constructed plasmids of GAL4-PPARγ-ligand binding domain and pGL4.35. The activation of PPARγ by different polarity extracts were evaluated based on the PPARγ transactivation assay and rosiglitazone was used as positive control. Results: Seven medicines (root bark of Lycium barbarum, Anoectochilus sroxburghii, the rhizome of Phragmites australis, Pterocephalus hookeri, Polygonatum sibiricum, fruit of Gleditsia sinensis, and Epimedium brevicornu) were able to significantly activate PPARγ. Conclusion: As seven medicines were able to activate PPARγ, the anti-diabetic activity of them is likely to be mediated by this nuclear receptor.
  2,573 103 -
Discovery of novel human epidermal growth factor receptor-2 inhibitors by structure-based virtual screening
Zheng Shi, Tian Yu, Rong Sun, Shan Wang, Xiao-Qian Chen, Li-Jia Cheng, Rong Liu
Apr-Jun 2016, 12(46):139-144
DOI:10.4103/0973-1296.177912  PMID:27076751
Background: Human epidermal growth factor receptor-2 (HER2) is a trans-membrane receptor like protein, and aberrant signaling of HER2 is implicated in many human cancers, such as ovarian cancer, gastric cancer, and prostate cancer, most notably breast cancer. Moreover, it has been in the spotlight in the recent years as a promising new target for therapy of breast cancer. Objective: Since virtual screening has become an integral part of the drug discovery process, it is of great significant to identify novel HER2 inhibitors by structure-based virtual screening. Materials and Methods: In this study, we carried out a series of elegant bioinformatics approaches, such as virtual screening and molecular dynamics (MD) simulations to identify HER2 inhibitors from Food and Drug Administration-approved small molecule drug as potential “new use” drugs. Results: Molecular docking identified top 10 potential drugs which showed spectrum affinity to HER2. Moreover, MD simulations suggested that ZINC08214629 (Nonoxynol-9) and ZINC03830276 (Benzonatate) might exert potential inhibitory effects against HER2-targeted anti-breast cancer therapeutics. Conclusion: Together, our findings may provide successful application of virtual screening studies in the lead discovery process, and suggest that our discovered small molecules could be effective HER2 inhibitor candidates for further study.
  2,513 110 -
Epicatechin plus treadmill exercise are neuroprotective against moderate-stage amyloid precursor protein/presenilin 1 mice
Zhiyuan Zhang, Hao Wu, Houcai Huang
May 2016, 12(46):139-146
DOI:10.4103/0973-1296.182174  PMID:27279698
Background: Epidemiological evidence suggests that exercise and dietary polyphenols are beneficial in reducing Alzheimer's disease (AD) risk. Materials and Methods: In the present study, 8 months old amyloid precursor protein/presenilin 1 (APP/PS1) mice (a moderate pathology phase) were given the green tea catechin (-)-epicatechin delivered orally in the drinking water (50 mg/kg daily), along with treadmill exercise for 4 months, in order to investigate whether the combination can ameliorate the cognitive loss and delay the progression of AD in APP/PS1 transgenic (Tg) mice. Results: At termination, untreated-Tg mice showed elevated soluble amyloid-β (Aβ1–40) and Aβ1–42levels and deficits in spatial learning and memory, compared with their wild-type littermates. The combined intervention protected against cognitive deficits in the Morris water maze, lowered soluble Aβ1–40and Aβ1–42levels in the hippocampus as well as reducing brain oxidative stress. In addition, brain-derived neurotrophic factor proteins wee elevated and Akt/GSK-3/cAMP response element-binding protein signaling was activated in the combination group. Conclusions: Dietary polyphenol plus exercise may exert beneficial effects on brain health and slow the progression of moderate- or mid-stages of AD. SUMMARY
  • Amyloid precursor protein/presenilin 1 transgenic mice showed elevated soluble amyloid-β (Aβ1–40) and Aβ1–42 levels and deficits in spatial learning and memory, compared with their wild.type littermates
  • Oral administration of epicatechin, combined with treadmill exercise for 4-months, could protect against cognitive deficits, and lowered soluble Aβ1–40) and Aβ1–42 levels as well as reducing brain oxidative stress
  • Brain-derived neurotrophic factor proteins were elevated, and Akt/GSK-3/cAMP response element binding protein signaling was activated in the combination group
  • Dietary polyphenol plus exercise might exert beneficial effects on brain health and slow the progression of moderate- or mid-stages of Alzheimer's disease.
Abbreviations used:AD: Alzheimer's disease, Tg: APP/PS1 transgenic, BDNF: Brain-derived neurotrophic factor, Aβ: Amyloid-β, APP: Amyloid precursor protein, PS1: Presenilin 1, nTg: Wild-type littermates, IACUC: Institutional Animal Care and Use Committee, GSSG: Glutathione oxidized form, GSH: Glutathione reductase, SOD: Superoxide dismutase, CAT: Catalase, LPO: Lipoperoxidation, CREB: cAMP response element binding protein. Houcai Huang
  2,513 98 2
Efficiency of matK, rbcL, trnH-psbA, and trnL-F (cpDNA) to molecularly authenticate Philippine ethnomedicinal Apocynaceae through DNA barcoding
Vincent Louie Domingo Cabelin, Grecebio Jonathan Duran Alejandro
April 2016, 12(46):384-388
DOI:10.4103/0973-1296.185780  PMID:27563229
Background: The Philippines is home to some ethnomedicinal Apocynaceae that has been used to cure common ailments. They are perceived to be safe, but misidentification can lead to substitution and adulteration. Morphological characters are primarily utilized to identify these species but a new method utilizing molecular characters called DNA barcoding has emerged. In this study, the efficiency of matK, rbcL, trnH-psbA, and trnL-F to molecularly authenticate selected Apocynaceae species were tested. Materials and Methods: Genomic DNA from silica-dried leaf samples were isolated and used as a template for generating DNA barcodes. Pair-wise sequence divergence using Kimura-2-Parameter was used to analyze inter-specific and intraspecific variations among the barcodes, whereas basic local alignment search tool (BLAST) and neighbor-joining (NJ) analyses were employed to examine discrimination success. Results: The results show that matK is the best barcode for Apocynaceae as it has the highest amplification and sequencing success together with rbcL while having high inter-specific and low intra-specific divergence relative to the other candidate barcodes. Furthermore, matK provided the highest discrimination both in BLAST and NJ analyses. Conclusion: This study proposes the use of matK as the principal barcode for Apocynaceae.
  2,472 136 -
Pharmacognostic screening of Piper trichostachyon fruits and its comparative analysis with Piper nigrum using chromatographic techniques
Vinayak Upadhya, Sandeep R Pai, Gireesh M Ankad, Harsha V Hegde
May 2016, 12(46):152-158
DOI:10.4103/0973-1296.182172  PMID:27279700
Background: Piper trichostachyon is a wild, endemic Piper species from Western Ghats of India. The folklore healers of Belagavi region use this plant, similar to Piper nigrum. Aims: The present study investigates the comparison between P. nigrum and P. trichostachyon using pharmacognostic parameters. Materials and Methods: Pharmacognostic evaluation was carried out in terms of morphological, microscopic characters, and phytochemical analysis using standard methods. Comparative physicochemical analysis between P. trichostachyon and P. nigrum was also carried out through estimation of micro-macro nutrients, high-performance thin layer chromatography (HPTLC) investigation and using piperine as a marker compound for reversed phase-ultra flow liquid chromatographic (RP-UFLC) technique. Results: P. trichostachyon grows in the forests, and the fruits are morphologically similar to P. nigrum fruits, so the name in Kannada “Kaadu Kalu menasu” (wild/forest black pepper). The microscopy revealed the presence of stone cells, starch grains, oil cells and globules, beaker cells, and yellowish brown pigment layer, parenchymatous cells. The presence of alkaloids, oil, and tannins were observed in P. trichostachyon fruits. The HPTLC studies visibly indicated differences among two species with 12 peaks and varied banding pattern. RP-UFLC results showed less amount of piperine in P. trichostachyon (0.05 ± 0.002 mg/g) than in P. nigrum (16.14 ± 0.807 mg/g). Conclusion: The study reports on pharmacognostic parameters of P. trichostachyon for the 1st time and will be useful for the identification and authentication. The comparative HPTLC and RP-UFLC studies resolve the differentiation impasse among two species. However, further biological efficacy studies are required to establish its use in traditional medicine. SUMMARY
  • Piper trichostachyon grows in the forests, and the fruits are morphologically similar to Piper nigrum fruits
  • The microscopy of P. trichostachyon revealed the presence of stone cells, starch grains, oil cells and globules, beaker cells and yellowish brown pigment layer, parenchymatous cells
  • The high-performance thin layer chromatography studies visibly indicated differences among two species with varied banding pattern
  • Reversed phase-ultra flow liquid chromatographic results showed less amount of piperine in P. trichostachyon than in P. nigrum.
Abbreviation used: HPTLC: High Performance Thin Layer Chromatography, RP-UFLC: Reversed phase-ultra fl ow liquid chromatographic analysis, DST: Length of line, Maj: Length of large half axis for ellipse RDS - radius for circle, Rf: Retention Factor, TS: Transverse Section, TLC: Thin Layer Chromatography.
  2,493 83 -
Cardioprotective and antioxidant influence of aqueous extracts from Sesamum indicum seeds on oxidative stress induced by cadmium in wistar rats
Babatunji Emmanuel Oyinloye, Basiru Olaitan Ajiboye, Oluwafemi Adeleke Ojo, Sarah Onyenibe Nwozo, Abidemi Paul Kappo
May 2016, 12(46):170-174
DOI:10.4103/0973-1296.182155  PMID:27279703
Background: Oxidative stress has been implicated in the pathogenesis of several acute and chronic diseases of the heart as a result of indiscriminate exposure to cardiotoxic heavy metals. The study reported here was designed to evaluate the possible ameliorative effect of aqueous extracts from Sesamum indicum (SI) seeds on oxidative stress induced by cadmium (Cd) in Wistar rats. Materials and Methods: Daily administration of Cd (200 mg/L Cd as CdCl2) in the animals' main drinking water for 21 days led to oxidative stress. Thereafter, the ameliorative effects were assessed by measuring biochemical parameters such as extent of lipid peroxidation (LPO), lipid profile, and enzymatic and nonenzymatic antioxidants, as well as serum aminotransferase activities. Results: Treatment with SI extract elicited notable reduction in serum total cholesterol, triglyceride, and low-density lipoprotein cholesterol levels as well as concomitant increase in high-density lipoprotein cholesterol. SI extract also reversed the elevations witnessed in serum aminotransferase activities, LPO level, and ameliorated enzymatic and nonenzymatic antioxidant status in the heart of Cd-exposed rats. Conclusion: Thus, SI appears to be an attractive candidate with potential for the novel treatment of cardiotoxicity and management of oxidative stress arising from Cd exposure. SUMMARY
  • Cadmium (200 mg/L) exposure in drinking water caused pronounced oxidative stress and cardiac tissue damage in animal model
  • Aqueous extract of Sesamum indicum (SI) seeds at a dose of 200 or 400 mg/kg body weight exhibited a significant reversal effect in all biochemical parameters measured such as extent of lipid peroxidation, lipid profile, and enzymatic and nonenzymatic antioxidants, as well as serum aminotransferase activities
  • Aqueous extract of SI seeds possess antioxidant and cardioprotective potential in a dose-dependent manner, thus conferring protection against oxidative stress induced by cadmium.
Abbreviation used: SI: Sesamum indicum, Cd: Cadmium, CdCl2: Cadmium chloride, LPO: Lipid peroxidation, TBA: Thiobarbituric acid, ALT: Alanine aminotransferase, AST: Aspartate aminotransferase, ALP: Alkaline phosphatise, TC: Total cholesterol, TG: Triglyceride, HDL-C: High-density lipoprotein cholesterol, LDL-C: Low-density lipoprotein cholesterol, SD: Standard deviation, GSH: Glutathione, SOD: Superoxide dismutase, CAT: Catalase, GST: Glutathione-S-transferase, GPx: Glutathione peroxidise.
  2,472 81 1
In vitro metabolism of sodium 9-dehydro-17-hydro- andrographolide-19-yl sulfate in rat liver s9 by liquid chromatography–mass spectrometry method
Dongkun Zheng, Jun Shao, Weikang Chen, Yuehua Luo
May 2016, 12(46):102-108
DOI:10.4103/0973-1296.182194  PMID:27279693
Background: Sodium 9-dehydro-17-hydro-andrographolide-19-yl sulfate (DHAS) is the active ingredient of Xiyanping injection, a traditional Chinese medicine in clinical use. However, there has been no report about the metabolic rate and metabolites of DHAS in vitro. Materials and Methods: In this article, DHAS was incubated with rat liver S9, and liquid chromatography/mass spectrometry (LC/MS) was used for the metabolism study. The residual concentrations of substrate were determined by ultra-high-performance liquid chromatography-electrospray ionization–tandem mass spectrometry method for the metabolic rate study of DHAS in liver S9. Metabolites were identified by the (UPLC-TOF-MSE) Ultra-performance liquid chromatography/quadrupole time-of-flight mass spectrometry method. Results: The calibration curves of DHAS were linear over the concentration range from 0.75 μM to 75.22 μM with correlation coefficients >0.99. The lower limit of quantification was 0.150 μM for DHAS. The determination recoveries of DHAS were in the range of 84.9–90.6%. The t½and CLintof DHAS in rat liver S9 were 98.6 ± 2.1 min and 3.5 ± 0.1 mL/min/g, respectively. Five metabolites were preliminarily identified based on the high resolution mass spectrum data in comparison with related references. These metabolites were mainly the products of dehydration and hydrogenation of DHAS. Conclusion: The present in vitro metabolic study of DHAS provided valuable information about the metabolic rate and potential metabolites of DHAS, which are important for future in vivo metabolism studies of DHAS and the discovery of more active andrographolide derivatives. SUMMARY
  • In this paper, sodium 9-dehydro-17-hydro-andrographolide-19-yl sulfate (DHAS) metabolism in vitro has been investigated with rat liver S9 using liquid chromatography-mass spectrometry (LC-MS). The result of quantitative analysis showed that DHAS had a long t1/2, which indicated its high metabolic stability. Five metabolites of DHAS were identified in the incubation system based on the high resolution mass spectrum data in comparison with related references, particularly dehydrated and hydrogenated products. The results would provide certain references to screen out more active andrographolide derivative for pre-clinically.
Abbreviations used: MRM: Multiple reaction monitoring, DHAS: Sodium 9-dehydro-17-hydro-andrographolide-19-yl sulfate, IS: Internal standard.
  2,410 84 -
Isolation and structure determination of 24-methylenecycloartanyl ferulate from Indian rice bran and its quantitative analysis
Amol P Muthal, Supada R Rojatkar, Subhash Laxmanrao Bodhankar
April 2016, 12(46):307-314
DOI:10.4103/0973-1296.185722  PMID:27563217
Background: γ-oryzanol is a major bioactive constituent in rice. Most of the literature reports isolation of 24-methylenecycloartanyl ferulate (24-mCAF) from rice bran oil (RBO) of other than Indian variety. Current research has successfully applied high-performance thin layer chromatography (HPTLC) method for isolation of 24-mCAF from Indian variety (Indrayani) of RBO. Materials and Methods: HPTLC method was developed for standard γ-oryzanol using tinidazole as an internal standard. The proposed HPTLC method was optimized and validated as per the guidelines stated by the International Conference on Harmonization Q2 R1 recommendations. The mobile phase composed of toluene:ethyl acetate:methanol (15.0:1.7:3.3, (v/v/v) was selected because well-resolved peaks were obtained. The optimum wavelength chosen for detection and quantitation was 317 nm. Results: The retention factors for tinidazole, 24-mCAF, and CAF were found to be 0.27 ± 0.02, 0.72 ± 0.02, and 0.79 ± 0.02, respectively. The percent content of 24-mCAF in ethanol fraction was found to be 1.02%. The 24-mCAF was isolated from RBO using HPTLC method. Conclusion: The characterization data of 1D, 2D spectral analysis confirm that the isolated compound 1 is 24-mCAF.
  2,306 123 -
The attenuation of Scutellariae radix extract on oxidative stress for colon injury in lipopolysaccharide-induced RAW264.7 cell and 2,4,6-trinitrobenzene sulfonic acid-induced ulcerative colitis rats
Yu Jin, Jun Yang, Lianjie Lin, Yan Lin, Changqing Zheng
Apr-Jun 2016, 12(46):153-159
DOI:10.4103/0973-1296.177913  PMID:27076753
Background: Oxidative stress (OS) has been regarded as one of the major pathogeneses of ulcerative colitis (UC) through damaging colon. It has been shown that Scutellariae radix (SR) extract has a beneficial effect for the prevention and treatment of UC. Objective: The aim of this study was to investigate whether SR had a potential capacity on oxidant damage for colon injury both in vivo and in vitro. Materials and Methods: The 2,4,6-trinitrobenzene sulfonic acid (TNBS) was used to induce UC rats model while 1 μg/ml lipopolysaccharide (LPS) was for RAW264.7 cell damage. Disease activity index (DAI) was determined to response the severity of colitis. The myeloperoxidase (MPO) activity in rat colon was also estimated. The 2,2'-azino-bis-3-ethylbenzthiazoline-6-sulfonic acid assay was performed to evaluate the total antioxidant capacity of SR. Furthermore, the activity of glutathione peroxidase (GSH-PX), catalase (CAT), superoxide dismutase (SOD), and lipid peroxidation malondialdehyde (MDA) in cell supernatant and rat serum were detected by appropriate kits. In addition, an immunohistochemical assay was applied to examine transforming growth factor beta 1 (TGF-β1) protein expression in colon tissue. Results: The treatment with SR could significantly increase the activity of GSH-PX, CAT, and SOD associated with OS in LPS-induced RAW264.7 cell damage and TNBS-induced UC rats. However, the level of MDA was markedly reduced both in vitro and in vivo. Furthermore, SR significantly decreased DAI and reversed the increased MPO activity. Thus, SR could decrease the severity of acute TNBS-induced colitis in rats. Immunohistochemical assay showed that SR significantly downregulated TGF-β1 protein expression in colon tissue. Conclusion: Our data provided evidence to support this fact that SR attenuated OS in LPS-induced RAW264.7 cell and also in TNBS-induced UC rats. Thus, SR may be an interesting candidate drug for the management of UC.
  2,335 80 2
Phytochemicals from Tradescantia albiflora Kunth extracts reduce serum uric acid levels in oxonate-induced rats
Wen-Ling Wang, Shi-Yuan Sheu, Wen-Dar Huang, Ya-Ling Chuang, Han-Chun Tseng, Tzann-Shun Hwang, Yuan-Tsung Fu, Yueh-Hsiung Kuo, Chun-Hsu Yao, Tzong-Fu Kuo
May 2016, 12(46):223-227
DOI:10.4103/0973-1296.182171  PMID:27279711
Background: Tradescantia albiflora (TA) Kunth (Commelinaceae) has been used for treating gout and hyperuricemia as folklore remedies in Taiwan. Therefore, it is worthwhile to study the effect of TA extracts on lowering uric acid activity. The hypouricemic effects of TA extracts on potassium oxonate (PO)-induced acute hyperuricemia were investigated for the first time. Materials and Methods: All treatments at the same volume (1 ml) were orally administered to the abdominal cavity of PO-induced hyperuricemic rats. One milliliter of TA extract in n-hexane (HE), ethyl acetate (EA), n-butanol (BuOH), and water fractions has 0.28, 0.21, 0.28, and 1.03 mg TA, respectively; and the plasma uric acid (PUA) level was measured for a consecutive 4 h after administration. Results: All four fractions' extracts derived from TA were observed to significantly reduce PUAcompared with the PO group. The EA-soluble fraction (TA-EA) exhibited the best xanthine oxidase (XO) inhibitory activity. Following column chromatography, 12 phytochemicals were isolated and identified from the EA fraction. The IC50values of isolated phytochemicals indicated that bracteanolide A (AR11) showed the remarkable XO inhibitory effect (IC50value of 76.4 μg/ml). These findings showed that the in vivo hypouricemic effect in hyperuricemic rats was consistent with in vitro XO inhibitory activity, indicating that TA extracts and derived phytochemicals could be potential candidates as hypouricemic agents. SUMMARY
  • Tradescantia albiflora extracts possess in vivo hypouricemic action in hyperuricemic rats
  • T. albiflora extracts exhibited strong inhibitory activity against xanthine oxidase (XO)
  • Butenolide may play an important role in XO inhibition
  • The extract bracteanolide A was demonstrated potent XO inhibitory activity in vitro.
Abbreviations used: TA: Tradescantia albiflora, PO: potassium oxonate, HE: n-hexane, EA: ethyl acetate, BuOH: n-butanol, PUA: plasma uric acid, XO: xanthine oxidase, MeOH: methanol, IP: intraperitoneal
  2,325 56 -
Chemopreventive agents from Physalis minima function as michael reaction acceptors
Ruizhi Men, Ning Li, Chihong Ding, Yingzhan Tang, Yachao Xing, Wanjing Ding, Zhongjun Ma
May 2016, 12(46):231-236
DOI:10.4103/0973-1296.182153  PMID:27279713
Background: The fruits of some varieties of genus Physalis have been used as delicious fruits and functional food in the Northeast of China. Materials and Methods: To reveal the functional material basis, we performed bioactivity-guided phytochemical research and chemopreventive effect assay of the constituents from Physalis minima. Results: It was demonstrated that the ethyl acetate extract of P. minima L. (EEPM) had potential quinone reductase (QR) inducing activity with induction ratio (IR, QR induction activity) value of 1.47 ± 0.24, and glutathione binding property as potential Michael reaction acceptors (with an α, β-unsaturated ketone moiety). Furthermore, bioactivity-guided phytochemical research led eight compounds (1–8), which were elucidated as 3-isopropyl-5-acetoxycyclohexene-2-one-1 (1), isophysalin B (2), physalin G (3), physalin D (4), physalin I (5), physordinose B (6), stigmasterol-3-O-β-D-glucopyranoside (7) and 5α-6β-dihydroxyphysalin R (8) on the basis of nuclear magnetic resonance spectroscopy analyses and HRESIMS. Then, isophysalin B (2) and physordinose B (6) showed significant QR inducing activity with IR value of 2.80 ± 0.19 and 2.38 ± 0.46, respectively. SUMMARY
  • An ultra.performance liquid chromatographic method with glutathione as the substrate was used to detect the Michael reaction acceptors in extracts of Physalis minima.(EPM)
  • We investigated the chemical constituents of EPM guided by biological activity method
  • Isophysalin B.(1) and physordinose B.(6) showed strong quinone reductase inducing activity with induction ratio values of 2.80±0.19 and 2.38±0.4
  • Abbreviations used: EPM: Extracts of Physalis minima, EEPM: Ethyl acetate extract of Physalis minima L., GSH: Glutathione, MRAs: Michael reaction acceptors, QR: Quinone reductase.
  • This study generated useful information for consumers and many encourage researchers to utilize edible fruits from Physalis as a source of phytochemicals.
  2,278 73 1
Neuroprotective properties of compounds extracted from Dianthus superbus L. against glutamate-induced cell death in HT22 cells
Bo-Ra Yun, Hye Jin Yang, Jin Bae Weon, Jiwoo Lee, Min Rye Eom, Choong Je Ma
Apr-Jun 2016, 12(46):109-113
DOI:10.4103/0973-1296.177905  PMID:27076746
Background: Dianthus superbus L. has been used in Chinese herbal medicine as a diuretic and anti-inflammatory agent. Objective: In this study, we isolated ten bioactive compounds from D. superbus and evaluated their neuroprotective activity against glutamate-induced cell death in the hippocampal neuronal HT22 cells. Materials and Methods: New compound, (E)-methyl-4-hydroxy-4-(8a-methyl-3-oxodecahydronaphthalen-4a-yl) (1) and, nine known compounds, diosmetin-7-O (2'',6''-di-O-α-L-rhamnopyranosyl)-β-D-glucopyranoside (2), 4-hydroxy-3-methoxy-pentyl ester benzenepropanoic acid (3), vanillic acid (4), 4-hydroxy-benzeneacetic acid (5), 4-methoxybenzeneacetic acid (6), (E)-4-methoxycinnamic acid (7), 3-methoxy-4-hydroxyphenylethanol (8), hydroferulic acid (9), and methyl hydroferulate (10), were isolated by bioactivity-guided separation. Structures of the isolated compounds were identified on the basis of 1H nuclear magnetic resonance (NMR), 13C NMR, and two-dimensional NMR spectra, while their neuroprotective properties were evaluated by performing the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. Results: D. superbus extract had a neuroprotective effect and isolated 10 compounds. Among the compounds, compounds 5 and 6 effectively protected HT22 cells against glutamate toxicity. Conclusion: In conclusion, the extract of D. superbus and compounds isolated from it exhibited neuroprotective properties, suggesting therapeutic potential for applications in neurotoxic diseases.
  2,276 72 -
Catha edulis extract induces H9c2 cell apoptosis by increasing reactive oxygen species generation and activation of mitochondrial proteins
Syam Mohan, Siddig Ibrahim Abdelwahab, Yahya Hasan Hobani, Suvitha Syam, Adel S Al-Zubairi, Rashad Al-sanousi, Magbool Essa Oraiby
April 2016, 12(46):321-326
DOI:10.4103/0973-1296.185732  PMID:27563219
Background: Catha edulis (Khat) is an evergreen shrub or small tree, traditionally used by various peoples of the Arabian Peninsula and Africa as an integral component of the socioeconomic traditions. It is believed that the psychostimulant nature and toxic nature of khat is primarily due to the presence of cathinone and cathine respectively. Studies have shown that khat chewing is closely associated with cardiac complications, especially myocardial infarction. Hence in this study, we exposed cathine-rich khat extract in a cardiomyoblast H9c2 (2-1) cell line to check the cell death mechanism. Materials and Methods: Extraction of Catha edulis leaves was done and the presence of cathine was confirmed with LC-MS-MS. The anti-proliferative activity was assayed using MTT and apoptosis detection by acridine orange/propidium iodide assay. The expression of Bcl-2 and Bax protein and caspase-3/7 expression were analyzed. The level of reactive oxygen species generation was also evaluated. Results: The khat extract showed an IC50 value of 86.5 μg/ml at 48 hours treatment. We have observed significant early apoptosis events by intervened acridine orange within the fragmented DNA with bright green fluorescence upon treatment. The Bcl-2 expression in the treatment with IC50 concentration of khat extract for 24, 48 and 72 hours of incubation significantly decreased with increase in bax level. The increased activation of caspase-3/7 was significantly observed upon treatment together with significant increase of ROS was detected at 24 and 48 hours treatment. Conclusion: Collectively, our results provide insight into the mechanisms by which Catha edulis leaves mediate cell death in cardiomyocytes.
  2,233 75 -
A new sucrase enzyme inhibitor from Azadirachta indica
Mohamed I. S. Abdelhady, Usama Shaheen, Ammar Bader, Mahmoud A Youns
April 2016, 12(46):293-296
DOI:10.4103/0973-1296.185705  PMID:27563214
Background: Sucrase enzyme inhibitor considered as an oral anti-diabetic therapy that delays the absorption of eaten carbohydrates, reducing the postprandial glucose and insulin peaks to reach normoglycemia. Materials and Methods: Chromatographic fractionation of the hydroalcoholic extract of leaves of Azadirachta indica growing in KSA, followed by in-vitro assay of sucrase enzyme inhibition activity. Results: This investigation led to the isolation of a new remarkable sucrase enzyme inhibitor; 4`-methyl Quercetin-7-O-β-D-glucuronopyranoside (1) alongside with four known compounds; 2,3-hexahydroxydiphenoyl-(α/β)-D-4C1 -glucopyranose (2), Avicularin (3), Castalagin (4) and Quercetin-3-O-glucoside (5). The structure of the new compound (1) was elucidated on the basis of its spectral data, including ESI-MS, UV, 1H NMR, 13C NMR, 1H- 1H COSY, HSQC, NOESY and HMBC. Conclusion: Under the assay conditions, hydroalcoholic extract of A. indica and compounds 1-5 exhibited significant sucrase enzyme inhibitory activity.
  2,164 140 -
DNA-based simultaneous identification of three Terminalia species targeting adulteration
Sonal Sharma, Neeta Shrivastava
April 2016, 12(46):379-383
DOI:10.4103/0973-1296.185776  PMID:27563228
Background: Various parts of three Terminalia species, namely, Terminalia arjuna (stem bark), Terminalia bellirica (fruit), and Terminalia chebula (fruit) are widely known for their therapeutic principles and other commercial values. However, stem bark of T. bellirica and T. chebula along with Terminalia tomentosa are reported as adulterants of T. arjuna. Correct botanical identification is very critical for safe and effective herbal drugs. DNA-based identification approaches are advancing the conventional methods and sometime proved more beneficial. Objective: The purpose of the study was to develop polymerase chain reaction (PCR) method using internal transcribed spacer (ITS) region to ascertain the identity of T. arjuna herbal material as well as detection of mixing of other three Terminalia species. Materials and Methods: DNA from stem barks samples were isolated and subjected to ITS region amplification and sequencing. Sequences were compared for polymorphic nucleotides determination to develop species-specific primers. Final primers were selected on the basis of in silico analysis and experimentally validated. PCR assays for botanical identification of Terminalia species were developed. Sensitivity testing and assay validation were also performed. Results: The PCR assays developed for Terminalia species were resulted in definite amplicons of the corresponding species. No cross-reactivity of the primers was detected. Sensitivity was found enough to amplify as low as 2 ng of DNA. Mixing of DNA in various concentrations for validation also proved the sensitivity of assay to detect original botanicals in the mixture. The developed methods proved very specific and sensitive to authenticate Arjuna bark to develop evidence-based herbal medicines.
  2,153 147 -
The Inhibitory Effect of Rhein on Proliferation of High Glucose-induced Mesangial Cell Through Cell Cycle Regulation and Induction of Cell Apoptosis
Shouzhu Xu, Yanying Lv, Jing Zhao, Junping Wang, Guangjian Wang, Siwang Wang
May 2016, 12(46):257-263
DOI:10.4103/0973-1296.182158  PMID:27279717
Objectives: Increased mesangial cell proliferation and accumulation of extracellular matrix (ECM) are the major pathological features of early-stage diabetic nephropathy. This study was sought to investigate the inhibitory effects of rhein (RH) on high glucose (HG)-cultured mesangial cells. Specially, we focus on the analysis of proliferation rate, cell cycle regulation, apoptosis, and the expression of collagen IV and laminin. Materials and Methods: The established rat renal mesangial cell (RMC) line was cultured in medium with different concentrations of glucose (5.6 mM or 25 mM) and RH (40 μM, 20 μM, and 10 μM). Pro-treated cells were collected at 12 h, 24 h, and 48 h for cell proliferation analysis and after 24 h for the experiments of flow cytometry, transmission electron microscope, real-time polymerase chain reaction, and Western blotting. Results: Our data shows HG can promote the proliferation of RMCs and RH has an inhibitory effect on HG-induced RMC proliferation and expression of ECM. Based on our data, we hypothesize this inhibitory effect might be a result of cell cycle regulation and the induction of cellular apoptosis. Conclusion: RH can inhibit cellular proliferation and downregulate the expression of ECM under the circumstance of HG. The mechanism of growth suppression may be due to cell cycle arrest at G1phase, induction of cell apoptosis, and upregulation of apoptotic mediators bax and caspase-3. SUMMARY
  • Rhein (RH) has an inhibitory effect on high glucose-induced rat mesangial cells proliferation
  • RH has an inhibitory effect on the expression of extracellular matrix
  • RH has a growth-suppression effect
  • RH can upregulate the expression of apoptotic mediators bax and caspase-3
  • All above shows RH is one of the main active ingredient in Shenkang injection.
Abbreviations used: RH: Rhein, ECM: Extracellular matrix, DN: Diabetic nephropathy, RMC: Renal mesangial cell, SKI: Shenkang injection, MTT: 3-(4,5-dimethylthiazol–2-yl)-2,5-diphenyltetrazolium bromide Siwang Wang
  2,190 92 -
Intrathecal injection of resveratrol attenuates burn injury pain by activating spinal sirtuin 1
Wei Cheng, Jin-Feng Wang, Cong-Xian Yang, Liang Wu, Qin Yin, He Liu, Zhi-Jian Fu
May 2016, 12(46):201-205
DOI:10.4103/0973-1296.182167  PMID:27279707
Objective: The present study sought to detect spinal sirtuin 1 (SIRT1) and acetylation of histone H3 (Ac-H3) expression in rats with burn injury pain (BIP model). Procedures and Results: A BIP model was first established. BIP rats showed lower paw withdrawal threshold (PWT) from day 1, which persisted for 21 days following the burn injury. Spinal SIRT1/Ac-H3 expression increased following burn injury. The intrathecal use of resveratrol increased PWT and SIRT1 expression but induced down-regulation of Ac-H3 expression. We first demonstrated that the inhibition of SIRT1 significantly induced mechanical allodynia in naïve rats. The preinjection of SIRT1 inhibitor partly antagonized the analgesic effects of resveratrol in BIP rats. Conclusion: Inhibition of SIRT1 produces pain facilitation in the naïve rats. The expression of spinal SIRT1 increased after burn injury in the BIP model. The activation of spinal SIRT1 might mediate the resveratrol-induced analgesic effects. SUMMARY
  • Burn injury resulted in pain facilitation
  • Resveratrol attenuates pain facilitation induced by burn injury
  • Intrathecal injection of resveratrol attenuates burn injury pain by increasing spinal sirtuin 1 (SIRT1) expression
  • Inhibition of SIRT1 by selisistat, an SIRT1 inhibitor attenuated analgesic effects of resveratrol.
Abbreviations used: SIRT1: Sirtuin 1, Ac-H3: Acetylation of histone H3, SD: Sprague-Dawley, EX527: Selisistat, an SIRT1 inhibitor, BIP: Burn injury pain, DMSO: Dimethyl sulfoxide, PWTs: Paw withdrawal thresholds Zhi-Jian Fu
  2,137 68 -
The antioxidant potential of Azadirachta indica ameliorates cardioprotection following diabetic mellitus-induced microangiopathy
Naveen Kumar Gupta, Nidhi Srivastva, Parvesh Bubber, Sanjeev Puri
April 2016, 12(46):371-378
DOI:10.4103/0973-1296.185772  PMID:27563227
Background: Cardiac complications associated with diabetes mellitus have become major cause of concern. Antidiabetic drugs, with varied mode of action, are although available, apprehensions exist for their limited action or side effects upon prolonged use. Efforts are therefore inclined toward finding other alternatives. The present study was, thus, undertaken to evaluate the cardioprotective effect of Azadirachta indica(AI) on microangiopathic changes in rat model of diabetes. Materials and Methods: Diabetes was induced in male rats by single intraperitoneal injection of streptozotocin (60 mg/kg body weight). Seven days after glucose levels are stabilized, aqueous leaf extract of AI (ALE) (600 mg/kg1 body weight) was administered orally to diabetic animals every day for 7 days. Results: High blood glucose characterizing diabetes in these animals was found to show increased lipid peroxidation (LPO), altered antioxidant biomarkers together with microangiopathic alterations. The treatment of diabetic rats with ALE reduced the levels of blood glucose, LPO, and restored the activities of antioxidant enzyme. Light and transmission electron microscopic analysis revealed reduced necrotic areas and inflammation in tissue architecture of ALE treated heart in comparison to untreated diabetic group. Conclusion: AI provides cardioprotection by ameliorating oxidative stress in rat model of diabetic mellitus.
  2,050 144 -
Optimization extraction process of polysaccharides from Suillus granulatus and their antioxidant and immunological activities In vitro
Feng Zhou, Song Yan, Shuang Chen, Liying Gong, Tingting Su, Zhanyong Wang
May 2016, 12(46):277-284
DOI:10.4103/0973-1296.182161  PMID:27279720
Background: Suillus granulatus is an edible and medicinal fungus in China. S. granulatus polysaccharide (SGP) was considered as the main bioactivity compounds in S. granulatus. Therefore, the extraction of SGP and their antioxidant activities were studied in this work. Materials and Methods: Fruiting bodies of S. granulatus were purchased from a local market (Fushun, China). Response surface methodology was adopted to optimize the extraction conditions of SGP. The antioxidant and immunological activities in vitro were also assayed. Results: The extraction of SGP was optimized by a Box–Behnken design. The optimal conditions for the extraction of polysaccharides were as follows: Pre-extraction time, 2 h; extraction temperature, 94°C; ratio of water to raw material, 25; and extraction frequency, 2. Under these conditions, the experimental yield of polysaccharides was 5.38% ±0.15%, which agreed with the predicted yield. The antioxidant assay in vitro showed that SGPs had relatively high scavenging ability for hydroxyl radicals and higher scavenging ability for 1,1-diphenyl-2-picrylhydrazyl radical. However, the scavenging ability of SGPs for superoxide anion radical and reducing power was relatively low. The polysaccharides also significantly increased splenocyte proliferation in vitro. Conclusion: SGP possessed good antioxidant and immunological activities in vitro and explored as a novel natural antioxidant or functional food. SUMMARY
  • The predictive model of Suillus granulatus polysaccharide (SGP) extraction is adequate for the extraction process
  • SGP possessed a good antioxidant activity in vitro
  • Lymphocyte proliferation in vitro was significantly increased by SGP
  • Pictorial abstract (in MS Powerpoint Format) is submitted as a separated file in the online submission system.
Abbreviation used: SGP: Suillus granulatus polysaccharides, RSM: Response surface methodology, BBD: Box–Behnken design, Vc: Ascorbic acid, DPPH: 1,1-diphenyl-2-picrylhydrazyl, MTT: 3-(4,5- dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide, ConA: Concanavalin A, LPS: lipopolysaccharide, RPMI-1640: Roswell Park Memorial Institute-1640. Zhanyong Wang
  2,115 73 -
Pyrostegia venusta (Ker Gawl.) miers crude extract and fractions: prevention of dental biofilm formation and immunomodulatory capacity
Mayara Brito de Sousa, José Otávio Carrera Silva, Wagner Luiz Ramos Barbosa, Erika da Silva Valério, Andriele da Mata Lima, Marlon Heggdorne de Araújo, Michelle Frazã Muzitano, Celso Vataru Nakamura, Joã Carlos Palazzo de Mello, Francisco Martins Teixeira
May 2016, 12(46):218-222
DOI:10.4103/0973-1296.182150  PMID:27279710
Background: Caries and periodontal diseases remain as important diseases in the Brazilian population. One important pathogen associated with this situation is Streptococcus mutans and other important factor is this pathogen's ability to adhere firmly to the tooth surface leading to dental biofilm formation and caries development. Objectives: Determine the antibacterial and other biological activities of P. venusta related to its potential to be used in the treatment of caries and periodontal disease. Methods: The growth inhibition by P. venusta of Streptococcus mutans, S. mitis, S. oralis and Candida albicans was determined using the broth microdilution method. In addition, the effect of the samples in adherence and reducing production of acids by S. mutans, and germ-tube formation of C. albicans was analysed. The Nitric Oxide (NO) production and cytotoxicity of P. venusta to peripheral blood mononuclear cells (PBMC) and RAW 264.7 Cell Line Murine Macrophage from Blood were assessed. Results: The crude extract (CE) and ethyl-acetate (AF) and n-butanol (BF) fractions showed antibacterial activity. The ethyl-acetate (AF) fraction showed the highest inhibition percentage against the adherence of S. mutans and C. albicans cells without budding, beyond NO production inhibition. There was not any cytotoxicity in the murine macrophages RAW 264.7 cells. Conclusion: Our results suggest that P. venusta presents potential to be used as a preliminary source of compounds that can provide helpful activity when used in prophylaxis or treatment of caries or periodontal disease. SUMMARY
  • Biological activities of Pyrostegia venusta and its potential for use in formulations for the prevention of oral diseases.
Abbreviations used: NO: Nitric oxide, PBMC: Peripheral blood mononuclear cells, CE: Crude extract, AF: Ethyl-acetate fraction, BF: n-butanol fraction, HF: Hexane fraction, WF: Water fraction, MIC: Minimum inhibitory concentration, MBC: Minimum bactericidal concentration, ATCC: American Type Culture Collection, CFU: Colony-forming units, BHI: Brain heart infusion, RPMI: Roswell Park Memorial Institute, MOPS: 3-(N-morpholino)propanesulfonic acid, DMEM: Dulbecco's modified Eagle's médium, LPS: Lipopolysacharide, MTT: 3-(4,5-Dimethylthiazol-2-yl)-2,5-Diphenyltetrazolium Bromide, OD: Optical density, AC: Acteoside, VB: β-OH-Verbascoside, TB: Trypan blue Francisco Martins Teixeira
  2,111 65 -
Screening of Chonemorpha fragrans bioactive extracts for cytotoxicity potential and inhibition studies of key enzymes involved in replication
Pradnya Prakash Kedari, Nutan Padmanabh Malpathak
April 2016, 12(46):297-302
DOI:10.4103/0973-1296.185708  PMID:27563215
Background: Chonemorpha fragrans (Moon) Alston, a liana belonging to family Apocynaceae, is used in traditional medicinal systems for the treatment of various ailments. It is an unexplored medicinal plant with respect to its anticancer potential. Objective: Cytotoxicity of sequential as well as crude extracts of in vivo plant parts (leaves, bark, and roots), in vitro cultures, and callus were compared. Materials and Methods: 3-(4,5- dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) cell proliferation assay was used to compare the extracts of various in vivo plant parts (leaves, bark, and roots) along with in vitro culture systems (in vitro plantlets, callus). Furthermore, the extracts were used to evaluate inhibition of key enzymes involved in replication, i.e. topoisomerase (Topo) I and II, DNA polymerase, to check the probable mechanism of action for this cytotoxicity. Results: MTT assay showed that the chloroform extract of callus has potent anticancer potential. The plant has a promising anticancer activity against human colon epithelium, lung carcinoma, and epidermoidal carcinoma cell lines. It was found to possess Topo as well as DNA polymerase inhibitory activity. Conclusion: The results have pointed toward pharmaceutical importance of this plant. This study is the first report of exploring the antiproliferative potential as well as inhibition studies of key enzymes involved in replication, which was useful to point out probable mechanism of action for extracts of C. fragrans.
  2,032 137 -
A new octadecenoic acid derivative from Caesalpinia gilliesii flowers with potent hepatoprotective activity
Samir M Osman, Alaadin E El-Haddad, Mohamed A El-Raey, Soad M Abd El-Khalik, Mahmoud A Koheil, Michael Wink
April 2016, 12(46):332-336
DOI:10.4103/0973-1296.185752  PMID:27563221
Background: Caesalpinia gilliesii Hook is an ornamental shrub with showy yellow flowers. It was used in folk medicine due to its contents of different classes of secondary metabolites. In our previous study, dichloromethane extract of C. gilliesii flowers showed a good antioxidant activity. Aim of the Study: Isolation and identification of bioactive hepatoprotective compounds from C. gilliesii flowers dichloromethane fraction. Materials and Methods: The hepatoprotective activity of dichloromethane fraction and isolated compounds were studied in CCl4-intoxicated rat liver slices by measuring liver injury markers (alanine aminotransferase, aspartate aminotransferase and glutathione [GSH]). All compounds were structurally elucidated on the basis of electron ionization-mass spectrometry, one- and two-dimensional nuclear magnetic resonance. Results: A new 12,13,16-trihydroxy-14(Z)-octadecenoic acid was identified in addition to the known β-sitosterol-3-O-butyl, daucosterol, isorhamnetin, isorhamnetin-3-O-rhamnoside, luteolin-7,4'-dimethyl ether, genistein-5-methyl ether, luteolin-7-O-rhamnoside, isovanillic acid, and p-methoxybenzoic acid. Dichloromethane fraction and isorhamnetin were able to significantly protect the liver against intoxication. Moreover, the dichloromethane fraction and the isolated phytosterols induced GSH above the normal level. Conclusion: The hepatoprotective activity of C. gilliesii may be attributed to its high content of phytosterols and phenolic compounds.
  2,021 138 -
Total alkaloids of Sophora alopecuroides inhibit growth and induce apoptosis in human cervical tumor hela cells In vitro
Jian-Guang Li, Xiao-Yi Yang, Wei Huang
May 2016, 12(46):253-256
DOI:10.4103/0973-1296.182157  PMID:27279716
Background: Uygur females of Xinjiang have the higher incidence of cervical tumor in the country. Alkaloids are the major active ingredients in Sophora alopecuroides, and its antitumor effect was recognized by the medical profession. Xinjiang is the main site of S. alopecuroides production in China so these plants are abundant in the region. Studies on the antitumor properties of total alkaloids of S. alopecuroides (TASA) can take full use of the traditional folk medicine in antitumor unique utility. Objectives: To explore the effects of TASA on proliferation and apoptosis of human cervical tumor HeLa cells in vitro. Materials and Methods: TASA was extracted, purified, and each monomer component was analyzed by high-performance liquid chromatography. The effect of TASA at different concentrations on the survival of HeLa cells was determined after 24 h using the Cell Counting Kit-8. In addition, cells were photographed using an inverted microscope to document morphological changes. The effect of TASA on apoptotic rate of HeLa cells was assessed by flow cytometry. Results: Monomers of TASA were found to be sophoridine, matrine, and sophocarpine. On treatment with 8.75 mg/ml of TASA, more than 50% of HeLa cells died, and cell death rate increased further with longer incubation. The apoptotic rates of HeLa cells in the experimental groups were 16.0% and 33.3% at concentrations of 6.25 mg/ml and 12.50 mg/ml, respectively. Conclusion: TASA can induce apoptosis in cervical tumor HeLa cells, and it has obvious inhibitory effects on cell growth. SUMMARY
  • Total alkaloids of Sophora alopecuroides (TASA) exhibits anti-human cervical tumor properties
  • Monomer component of TASA was analyzed by high-performance liquid chromatography, and its main effect component are sophoridine, matrine, and sophocarpine
  • TASA inhibits growth and induces apoptosis in HeLa cells.
Abbreviations used: TASA: Total alkaloids of S. alopecuroides, CCK-8: Cell Counting Kit-8, FBS: Fetal bovine serum, PBS: Phosphate buffered saline, DMEM: Dulbecco's modified Eagle medium
  2,063 74 -
The anticholinesterase properties of plants from the northeast of brazil selected by an ethnopharmacological study for disorders relating to the nervous system
Valerium Thijan Nobre de Almeida e Castro, Tadeu Jose da Silva Peixoto Sobrinho, Allan Jonathan Chernichiarro Corrêa, Thiago Antonio de Sousa Araújo, Terezinha Gonçalves Da Silva, Elba Lucia Cavalcanti de Amorim
May 2016, 12(46):195-200
DOI:10.4103/0973-1296.182166  PMID:27279706
Background: Various factors may trigger Alzheimer's disease and the cholinergic hypothesis, which is one of the most widely accepted, argues damage to the brain nuclei, may reduce the production of the choline acetyltransferase enzyme, and cause a decline in the synthesis of acetylcholine (ACh). Studies have thus focused on discovering molecules that are capable of inhibiting the action of cholinesterase enzymes that degrade ACh, thereby preventing the evolution of the disease. Objective: The aim of the present study is to assess the anticholinesterase properties of extracts of medicinal plants in a semi-arid region of Northeast of Brazil. Materials and Methods: The species were selected by way of an ethnobotanical study and were collected if there were some indications that they are related to the nervous system. The plant samples were extracted using hexane, ethyl acetate, and methanol. Anticholinesterase activity in vitro was assessed by way of bioautography in thin layer chromatography and microassays in 96-well plates. Results: Twenty-three species of plant were collected, and 75 extracts were analyzed. The bioautography revealed that 26.7% of the samples showed inhibitory activity against the acetylcholinesterase (AChE) enzyme. After the test for false positives, 8% of the samples were found to inhibit AChE. Thirty samples were analyzed by microassay (500 μg/mL), on which 86.7% showed moderate to powerful anticholinesterase activity. Conclusion: Of the extracts tested, Citrus limonum, Ricinus communis, and Senna occidentalis stand out as was the most promising in terms of anticholinesterase activity and may serve as a guide for the discovery and development of new substances for the treatment of AD. SUMMARY
  • The bioautography revealed that 26.7% of the samples showed inhibitory activity against the acetylcholinesterase enzyme
  • Samples were analyzed by microassay (500 μg/mL), upon which 86.7% showed moderate to powerful anticholinesterase activity
  • Citrus limonum, Ricinus communis, and Senna occidentalis stand out as being the most promising in terms of anticholinesterase activity
  • C. limonum, R. communis, and S. occidentalis may serve as a guide for the discovery and development of new substances for the treatment of Alzheimer's disease.
Abbreviations used: AChE: Acetilcolinesterase Valerium Thijan Nobre de Almeida e Castro
  2,062 73 -
A study on the chemical compositions of the Yinqiaosan (Lonicerae and Forsythiae powder) at different time of later-decoction by gas chromatography mass spectrometry
Yachun Shu, Yajun Chen, Kunming Qin, Xiao Liu, Baochang Cai
Apr-Jun 2016, 12(46):134-138
DOI:10.4103/0973-1296.177911  PMID:27076750
Background: Yinqiaosan (Lonicerae and Forsythiae Powder), as a famous prescription of Dr. Wu Jutong in Qing dynasty of China, has the effects of diaphoresis cooling, fire-purging, and detoxicaton. It is mainly used in the treatment of influenza, hand-foot-mouth disease, esophagitis, pneumonia, acute tonsillitis, mumps, and other viral infections. It is one of the widely used traditional Chinese medicine prescriptions with proven curative effects in clinical use. Objective: To research the material basis of Yinqiaosan decoction when decocting mint, herba schizonepetae in different length of later-decoction time, to find the influence on volatile components of Yinqiaosan decoction decocted later in different length of time, to lay the foundation to further clarify the after-decoction mechanism of Yinqiaosan, and the specification of Yinqiaosan decoction process. Materials and Methods: Gas chromatography mass spectrometry method is used to analyze the volatile components of Yinqiaosan decoction samples decocted for 0, 3, 5, 8, and 10 min. Results: Later-decocting mint and herba schizonepetae at different time when decocting Yinqiaosan had a significant influence on the volatile components of the solution. 54 different chemical components were identified: 25 were identified when later-decocting the sample for 3 min; 13 were identified when later-decocting the sample for 5 min; 11 were identified when later-decocting the sample for 8 min; 7 were identified when later-decocting the sample for 10 min; and 26 were identified when later-decocting the sample for 0 min. There were more volatile components in the sample after-decocted for 3 min. A total of 54 different chemical components were identified in different later-decocting solution samples. These components form the basis of the Yinqiaosan drug effect. Conclusions: The length of later-decoction time of mint and herba schizonepetae was confirmed to be 3 min when decocting Yinqiaosan.
  2,056 65 -
Quantitative analysis and comparison of four major flavonol glycosides in the leaves of Toona sinensis (A. Juss.) roemer (chinese toon) from various origins by high-performance liquid chromatography-diode array detector and hierarchical clustering analysis
Xiaoxiang Sun, Liting Zhang, Yaqi Cao, Qinying Gu, Huan Yang, James P Tam
May 2016, 12(46):270-276
DOI:10.4103/0973-1296.182160  PMID:27279719
Background: Toona sinensis (A. Juss.) Roemer is an endemic species of Toona genus native to Asian area. Its dried leaves are applied in the treatment of many diseases; however, few investigations have been reported for the quantitative analysis and comparison of major bioactive flavonol glycosides in the leaves harvested from various origins. Objective: To quantitatively analyze four major flavonol glycosides including rutinoside, quercetin-3-O-β-D-glucoside, quercetin-3-O-α-L-rhamnoside, and kaempferol-3-O-α-L-rhamnoside in the leaves from different production sites and classify them according to the content of these glycosides. Materials and Methods: A high-performance liquid chromatography-diode array detector (HPLC-DAD) method for their simultaneous determination was developed and validated for linearity, precision, accuracy, stability, and repeatability. Moreover, the method established was then employed to explore the difference in the content of these four glycosides in raw materials. Finally, a hierarchical clustering analysis was performed to classify 11 voucher specimens. Results: The separation was performed on a Waters XBridge Shield RP18 column (150 mm × 4.6 mm, 3.5 μm) kept at 35°C, and acetonitrile and H2O containing 0.30% trifluoroacetic acid as mobile phase was driven at 1.0 mL/min during the analysis. Ten microliters of solution were injected and 254 nm was selected to monitor the separation. A strong linear relationship between the peak area and concentration of four analytes was observed. And, the method was also validated to be repeatable, stable, precise, and accurate. Conclusion: An efficient and reliable HPLC-DAD method was established and applied in the assays for the samples from 11 origins successfully. Moreover, the content of those flavonol glycosides varied much among different batches, and the flavonoids could be considered as biomarkers to control the quality of Chinese Toon. SUMMARY
  • Four major flavonol glycosides in the leaves of Toona sinensis were determined by HPLC-.DAD and their contents were compared among various origins by HCA.
Abbreviations used: HPLC-DAD: High-performance liquid chromatography-diode array detector, HCA: Hierarchical clustering analysis, MS: Mass spectrometry, RSD: Relative standard deviation. Huan Yang, achieved his Ph. D. degree in 2008 from Nanjing University of Chinese Medicine, China. Currently, he is an Associate Professor at the Department of Chinese Materia Medica in the School of Pharmacy, Jiangsu University, China. His major research interest is in the area of chemical constituents of Chinese Medicine. In past years, Dr. Yang has been working on both plant sourced and animal derived Chinese Medicines, and engaged in finding active substances as well as pharmacological mechanism of these natural products. Huan Yang
  2,027 78 -
Purified essential oil from Ocimum sanctum Linn. triggers the apoptotic mechanism in human breast cancer cells
Thamilvaani Manaharan, Ramaraj Thirugnanasampandan, Rajarajeswaran Jayakumar, MS Kanthimathi, Gunasekar Ramya, Madhusudhanan Gogul Ramnath
April 2016, 12(46):327-331
DOI:10.4103/0973-1296.185738  PMID:27563220
Background: Essential oil of Ocimum sanctum Linn. exhibited various pharmacological activities including antifungal and antimicrobial activities. In this study, we analyzed the anticancer and apoptosis mechanisms of Ocimum sanctum essential oil (OSEO). Objective: To trigger the apoptosis mechanism in human breast cancer cells using OSEO. Materials and Methods: OSEO was extracted using hydrodistillation of the leaves. Cell proliferation was determined using different concentrations of OSEO. Apoptosis studies were carried out in human breast cancer cells using propidium iodide (PI) and Hoechst staining. Results: We found that OSEO inhibited proliferation (IC50 = 170 μg/ml) of Michigan cancer foundation-7 (MCF-7) cells in a dose-dependent manner. The OSEO also induced apoptosis as evidenced by the increasing number of PI-stained apoptotic nucleic of MCF-7 cells. Flow cytometry analysis revealed that treatment with OSEO (50–500 μg/ml) increased the apoptotic cells population (16–84%) dose dependently compared to the control. OSEO has the ability to up-regulate the apoptotic genes p53 and Bid and as well as elevates the ratio of Bax/Bcl-2. Conclusion: Our findings indicate that OSEO has the ability as proapoptotic inducer and it could be developed as an anticancer agent.
  2,003 80 -
Life span and motility effects of ethanolic extracts from Sophora moorcroftiana seeds on Caenorhabditis elegans
Xin Li, Junxian Han, Rongyan Zhu, Rongrong Cui, Xingming Ma, Kaizhong Dong
May 2016, 12(46):228-230
DOI:10.4103/0973-1296.182152  PMID:27279712
Background: Sophora moorcroftiana is an endemic shrub species with a great value in folk medicine in Tibet, China. In this study, relatively little is known about whether S. moorcroftiana is beneficial in animals' nervous system and life span or not. Materials and Methods: To address this question, under survival normal temperature (25°C), S. moorcroftiana seeds were extracted with 95% ethanol, and Caenorhabditis elegans were exposed to three different extract concentrations (100 mg/L, 200 mg/L, and 400 mg/mL) from S. moorcroftiana seeds. Results: The 95% ethanolic extracts from S. moorcroftiana seeds could increase life span and slow aging-related increase in C. elegans and could not obviously influence the motility of C. elegans. Conclusion: Given these results by our experiment for life span and motility with 95% ethanolic extracts from S. moorcroftiana seeds in C. elegans, the question whether S. moorcroftiana acts as an anti-aging substance in vivo arises. SUMMARY
  • The 95% ethanolic extracts from S. moorcroftiana seeds have no effect on the life span in C. elegans when extract concentrations from S. moorcroftiana seeds <400 mg/L
  • The 400 mg/L 95% ethanolic extracts from S. moorcroftiana seeds could increase life span in C. elegans
  • The 95% ethanolic extracts from S. moorcroftiana seeds could not obviously influence the motility in C. elegans.
Abbreviation used: S. moorcroftiana: Sophora moorcroftiana; C. elegan: Caenorhabditis elegan; E. coli OP50: Escherichia coli OP50; DMSO: Dimethyl sulfoxide.
  1,962 87 -
High performance liquid chromatography-mass spectrometry analysis of high antioxidant australian fruits with antiproliferative activity against cancer cells
Joseph Sirdaarta, Anton Maen, Paran Rayan, Ben Matthews, Ian Edwin Cock
May 2016, 12(46):181-194
DOI:10.4103/0973-1296.182178  PMID:27279705
Background: High antioxidant capacities have been linked to the treatment and prevention of several cancers. Recent reports have identified several native Australian fruits with high antioxidant capacities. Despite this, several of these species are yet to be tested for anticancer activity. Materials and Methods: Solvent extracts prepared from high antioxidant native Australian fruits were analyzed for antioxidant capacity by the di (phenyl)-(2,4,6-trinitrophenyl) iminoazanium free radical scavenging assay. Antiproliferative activities against CaCo2 and HeLa cancer cells were determined by a multicellular tumor spheroid-based cell proliferation assay. Toxicity was determined by Artemia franciscana bioassay. Results: Methanolic extracts of all plant species displayed high antioxidant contents (equivalent to approximately 7–16 mg of vitamin C per gram of fruit extracted). Most aqueous extracts also contained relatively high antioxidant capacities. In contrast, the ethyl acetate, chloroform, and hexane extracts of most species (except lemon aspen and bush tomato) had lower antioxidant contents (below 1.5 mg of vitamin C equivalents per gram of plant material extracted). The antioxidant contents correlated with the ability of the extracts to inhibit proliferation of CaCo2 and HeLa cancer cell lines. The high antioxidant methanolic extracts of all species were potent inhibitors of cell proliferation. The methanolic lemon aspen extract was particularly effective, with IC50 values of 480 and 769 μg/mL against HeLa and CaCo2 cells, respectively. In contrast, the lower antioxidant ethyl acetate and hexane extracts (except the lemon aspen ethyl acetate extract) generally did not inhibit cancer cell proliferation or inhibited to only a minor degree. Indeed, most of the ethyl acetate and hexane extracts induced potent cell proliferation. The native tamarind ethyl acetate extract displayed low-moderate toxicity in the A. franciscana bioassay (LC50 values below 1000 μg/mL). All other extracts were nontoxic. A total of 145 unique mass signals were detected in the lemon aspen methanolic and aqueous extracts by nonbiased high-performance liquid chromatography-mass spectrometry analysis. Of these, 20 compounds were identified as being of particular interest due to their reported antioxidant and/or anticancer activities. Conclusions: The lack of toxicity and antiproliferative activity of the high antioxidant plant extracts against HeLa and CaCo2 cancer cell lines indicates their potential in the treatment and prevention of some cancers. SUMMARY
  • Australian fruit extracts with high antioxidant contents were potent inhibitors of CaCo2 and HeLa carcinoma cell proliferation
  • Methanolic lemon aspen extract was particularly potent, with IC50 values of 480 μg/mL (HeLa) and 769 μg/mL (CaCo2)
  • High.performance liquid chromatography.mass spectrometry.quadrupole time.of.flight analysis highlighted and putatively identified 20 compounds in the antiproliferative lemon aspen extracts
  • In contrast, lower antioxidant content extracts stimulated carcinoma cell proliferation
  • All extracts with antiproliferative activity were nontoxic in the Artemia nauplii assay.
Abbreviations used: DPPH: di (phenyl)- (2,4,6-trinitrophenyl) iminoazanium, HPLC: High-performance liquid chromatography, IC50: The concentration required to inhibit by 50%, LC50: The concentration required to achieve 50% mortality, MS: Mass spectrometry. Ian Edwin Cock
  1,990 58 -
Cocculus hirsutus: Molecular docking to identify suitable targets for hepatocellular carcinoma by in silico technique
B Samuel Thavamani, Molly Mathew, SP Dhanabal
April 2016, 12(46):350-352
DOI:10.4103/0973-1296.185769  PMID:27563224
Background: Protein–ligand interaction plays a major role in identification of the possible mechanism by which a ligand can bind with the target and exerts the pharmacological action. Objective: The aim is to identify the best candidate of Cocculus hirsutus which binds with the hepatocellular carcinoma (HCC) targets by docking studies. Materials and Methods: The reported phytoconstituents such as coclaurine, hirsutine, cohirsine, cohirsinine, lirioresinol, cohirsitinine, haiderine, jamtinine, isotrilobine, shaheenine, jamtine, and cocsoline present in the plant, C. hirsutus were docked with the HCC targets such as Aurora kinase, c-Kit, fibroblast growth factor, nuclear factor kappa B (NF-kB), B-cell lymphoma-extra large, and vascular endothelial growth factor (VEGF) using in silico technique with the software Grid-Based Ligand Docking with Energies. Results: Haiderine, shaheenine, and coclaurine had good interaction with Aurora kinase with the glide score and glide energy of − 7.632, −7.620, −7.464; and − 56.536, −55.203, −52,822, respectively. Coclaurine, lirioresinol, and haiderine possess good binding with c-Kit with the glide score and glide energy of − 8.572, −6.640, −6.478; and − 56.527, −57.138, −20,522, respectively. Lirioresinol, hirsutine, and coclaurine exhibit good binding with c-Kit with the glide score and glide energy of − 5.702, −5.694, −5.678; and − 48.666, −35.778, −41,673, respectively. Similarly, coclaurine, haiderine, and hisutine had good interaction with NF-kB. Haiderine, jamtinine, and coclaurine had good binding with VEGF receptors (VEGFR) and coclaurine, lirioresinol, and haiderine exhibit good bonding with VEGFR. Conclusion: Coclaurine, haiderine, and lirioresinol exibited good hydrogen bonding interactions and binding energy with the select targets. Hence, these compounds have to be taken up for experimental work against hepatocellular carcinoma.
  1,920 119 -
Potency of Massoia bark in combating immunosuppressed-related infection
Triana Hertiani, Sylvia Utami Tunjung Pratiwi, Agustinus Yuswanto, Prisci Permanasari
April 2016, 12(46):363-370
DOI:10.4103/0973-1296.185771  PMID:27563226
Background: As part of our search for new potential natural resources to eradicate infection, we have revealed the prominent potency of massoia bark (Massoia aromatica Becc, Lauraceae) in combating immunosuppressed-related infection. Materials and Methods: The extract was prepared by macerating the pulverized dried bark in ethanol 95%, followed by solvent evaporation. The oil was extracted from the dried bark by steam-hydrodistillation of which preparative thin-layer chromatography was performed on the oil to isolate the active constituent, C-10 massoia lactone (ML). Anti-biofilm assay against Candida albicans was conducted on polystyrene 96 wells microtiter plates, followed by a confocal laser scanning microscope observation to get three-dimensional profiles of the affected biofilms. Effects on the hyphae development inoculated on RPMI-1640 agar plates were observed for 7 days. Influences of samples on mice macrophage phagocytosis were examined by an in vitro technique. Samples concentration tested were in the range of 2.0–0.0625 mg/mL and done in triplicate. Results: Massoia bark extracts (oil and solid phase) and ML exhibited promising activities as anti-biofilm against C. albicans at IC50 0.074% v/v, 271 μg/mL and 0.026 μg/mL, respectively. The ML did not inhibit the hyphae development at the concentration tested; however, the extracts showed inhibition at 62.5 μg/mL. Macrophage phagocytosis stimulation was correlated to the ML content. Conclusion: Massoia bark is potential to be developed as anti-infective in immunosuppressed condition of which the C10 ML (C10H16O2) plays a major role in exerting activity.
  1,880 136 1
New abietane diterpenes from Euphorbia pseudocactus berger (Euphorbiaceae) and their antimicrobial activity
Azza Ramy Abdel-Monem, Enas Hussein Abdelrahman
April 2016, 12(46):346-349
DOI:10.4103/0973-1296.185768  PMID:27563223
Background: Euphorbia is the largest genus in Euphorbiaceae. Terpenoids were isolated from most species of this genus. Objective: Since no previous study was reported about Euphorbia pseudocactus Berger, we started here a phytochemical investigation on this species to isolate and identify its terpenoid constituents and to estimate the antimicrobial activity of the isolated compounds. Materials and Methods: The n-hexane fraction of the ethanolic extract of E. pseudocactus Berger was chromatographed on silica gel columns, the structures of the isolated compounds (1–5) were identified based on their MS, 1 D, and 2 D NMR spectral data. The antimicrobial activity of the n-hexane fraction and the isolated compounds (1–4) was investigated using diffusion plate method against Gram-positive (Staphylococcus aureus [12600] and Bacillus subtilis [6051]) and Gram-negative (Pseudomonas aeruginosa [10145] and Escherichia coli [11775]) bacteria, yeast (Candida albicans [7102]), and fungi (Aspergillus flavus). Results: Two triterpenes (glut-5-en-3 β-ol [1] and olean-12,15-diene-3 β-ol [2]) and three abietane diterpene (3-hydoxy-19-cyclopropenoyloxy-abietane [3], ent-abieta-9,12,14-triene-12,16-olide [4], and 12,19-dihydroxy-abieta-5-ene [5]) were isolated. Compound 1 exhibited no antibacterial activity against the tested bacteria, compound 2 and n-hexane fraction exhibited weak activity, whereas compounds 3 and 4 showed moderate activity. All samples showed no activity against the tested yeast and fungi. Discussion and Conclusion: Five compounds were isolated for the 1st time from E. pseudocactus Berger, three of them (3–5) are new natural compounds. As the major isolated compound (1) exhibited no antimicrobial activity, the observed activity of the n-hexane fraction is mainly due to its diterpenoid constituents.
  1,877 137 -
Reversed phase high-performance liquid chromatographic ultra-violet (photo diode array) quantification of oleanolic acid and its isomer ursolic acid for phytochemical comparison and pharmacological evaluation of four Leucas species used in ayurveda
Pushpendra Kumar Shukla, Ankita Misra, Sharad Srivastava, Ajay K. S. Rawat
May 2016, 12(46):159-164
DOI:10.4103/0973-1296.182173  PMID:27279701
Content: Different Leucas species are well known as “Dronpushpi,” a well-known herb of Ayurveda, used in the treatment of various ailments. Objective: Evaluation of four industrially important Leucas species for their in vitro antidiabetic potential and radical scavenging effect along with high-performance liquid chromatographic quantification of the bioactive triterpenes. Materials and Methods: The quantification of triterpenes was carried out on C-18 column with acetonitrile and water (90:10) as the solvent system at a detection wavelength of 210 nm. In vitro antidiabetic activity was evaluated by α-amylase inhibition assay based on starch–iodine and 3,5 dinitrosalicylic acid (DNS) method. Antioxidant activity was calculated by five different models, namely total phenolic and total flavonoid content, free radical scavenging activity by 1-1-diphenyl-2-pic-rylhydrazyl (DPPH), ferric-reducing power assay, and the total antioxidant capacity. Results: Maximum concentration of oleanolic acid was found in Leucas cristata, followed by Leucas mollissima, Leucas Aspera, and Leucas biflora. Ursolic acid was highest in L. mollissima and then in L. biflora, L. cristata, and L. aspera, respectively. In in vitro antidiabetic activity, IC50of L. aspera (1.56 ± 0.01 mg/ml) and L. mollissima (0.75 ± 0.005 mg/ml) were found to be highest in DNS and iodine starch assay. IC50in DPPH assay ranges from 0.6 ± 0.011 to 1.68 ± 0.011 mg/ml. Antioxidant capacity follows the order; L. aspera > L. mollissima > L. biflora > L. cristata. Conclusion: Promising activities were observed in targeted species, thus L. mollissima, L. biflora, and L. cristata can be used alternatively as a substitute to L. aspera. SUMMARY
  • Physicochemical parameters are within the limit as per the Ayurvedic Pharmacopoeia of India
  • Maximum concentration of oleanolic acid was found in Leucas cristata; however, ursolic acid was highest in Leucas mollissima
  • In vitro antidiabetic activity of Leucas aspera and L. mollissima was found to
  • be heighest as compared to other species. However, antioxidant capacity is almost similar in targeted species.
  • Promising activities were observed in all the species, thus L. mollissima, Leucas biflora, and L. cristata can be used alternatively as a substitute to L. aspera.
  1,918 69 -
Minimization of the risk of diabetic microangiopathy in rats by Nigella sativa
Juraiporn Somboonwong, Mariem Yusuksawad, Somboon Keelawat, Sirima Thongruay, Ubon Poumsuk
May 2016, 12(46):175-180
DOI:10.4103/0973-1296.182169  PMID:27279704
Background: Microangiopathy is a chronic diabetic complication resulting from metabolic derangements, oxidative stress, and increased pro-inflammatory cytokine production. Nigella sativa Linn. is used as an herbal medicine that exerts hypoglycemic, antilipidemic, anti-inflammatory, and antioxidant effects. Objective: To examine the effects of N. sativa extract on cutaneous microvascular changes in diabetic rats. Materials and Methods: Sprague-Dawley rats were randomly assigned into the following four groups: Untreated and N. sativa-treated normal controls and untreated and N. sativa-treated rats with streptozotocin-induced diabetes. A cold-pressed N. sativa extract was then orally administered (1000 mg/kg/day). After 8 weeks of treatment, the glucose, glycosylated hemoglobin (HbA1c), tumor necrosis factor-alpha (TNF-α), insulin levels, and lipid profile were determined in cardiac blood. Dermal capillary wall thickness was measured in tail skin sections stained with periodic acid-Schiff. Endothelial apoptosis was morphologically evaluated by hematoxylin and eosin staining. Results: Diabetes significantly reduced the circulating insulin and low-density lipoprotein levels and caused elevations in the glucose, HbA1c, and triglyceride levels, accompanied by a slight increase in total cholesterol levels and no change in the high-density lipoprotein and TNF-α levels. Capillary basement membrane thickening and a decreased capillary luminal diameter despite no evidence of endothelial cell apoptosis were also observed. N. sativa treatment of diabetic rats reduced the mean HbA1cconcentration by 1.4%, enlarged the capillary lumens, and tended to attenuate dermal capillary basement membrane thickening without affecting the lipid profile or TNF-α level. Conclusion: Our results indicate that N. sativa may be used to minimize the risk of diabetic microangiopathy, potentially due in part to its glycemic control activity. SUMMARY
  • Diabetes causes dermal capillary basement membrane thickening and a decreased capillary luminal diameter
  • Nigella sativa treatment of diabetic rats enlarged the capillary lumens and tended to attenuate dermal capillary basement membrane thickening
  • N. sativa treatment of diabetic rats reduced the mean glycosylated hemoglobin concentration by 1.4%, which exceeds the necessary reduction previously described to decrease the risk of diabetic microangiopathy, without affecting the lipid profile or tumor necrosis factor-alpha level
  • N. sativa improves rat diabetic microangiopathy, potentially due in part to its glycemic control activity.
Abbreviations used: H and E: Hematoxylin and eosin, HbA1c: Glycosylated hemoglobin, HDL-C: High-density lipoprotein cholesterol, LDL-C: Low-density lipoprotein cholesterol, PAS: Periodic acid-Schiff, STZ: Streptozotocin, TNF-α: Tumor necrosis factor-alpha. Juraiporn Somboonwong
  1,874 68 -
DNA barcoding identification of kadsurae caulis and spatholobi caulis based on internal transcribed spacer 2 region and secondary structure prediction
Xiaoxue Yu, Zhiyong Xie, Junwei Wu, Junfei Tao, Xinjun Xu
May 2016, 12(46):165-169
DOI:10.4103/0973-1296.182162  PMID:27279702
Background: Kadsurae Caulis and Spatholobi Caulis have very similar Chinese names. Their commodities were hard to distinguish because their stems were very alike after dried and processed. These two herbal drugs were often mixed in clinical use. Objective: Authenticity assurance is crucial for quality control of herbal drugs. Therefore, it is essential to establish a method for identifying the two herbs. Materials and Methods: In this paper, we used the DNA barcoding technology, based on the internal transcribed spacer 2 (ITS2) regions, to differentiate Kadsurae Caulis and Spatholobi Caulis. Results: The ITS2 of these two herbs were very different. They were successfully differentiated using the DNA barcoding technique. Conclusions: DNA barcoding was a promising and reliable tool for the identification of medicinal plants. It can be a powerful complementary method for traditional authentication. SUMMARY
  • The internal transcribed spacer 2 (ITS2) regions between Kadsurae Caulis and Spatholobi Caulis varied considerably, totally 139 variable sites
  • Sample 1 was not Kadsurae Caulis as it labeled, but it should be Spatholobi Caulis in fact based on ITS2 region
  • The secondary structure can also separate Kadsurae Caulis and Spatholobi Caulis effectively
  • DNA barcoding provided an accurate and strong prove to identify these two herbs.
Abbreviations used: CTAB: hexadecyltrimethylammonium bromide, DNA: deoxyribonucleic acid, ITS2:internal transcribed spacer 2, PCR: polymerase chain reaction
  1,857 78 1
Protective effect of high molecular weight protein sub-fraction of Calotropis procera Latex in Monoarthritic Rats
Priyanka Chaudhary, Marcio V Ramos, Mirele da Silveira Vasconcelos, Vijay L Kumar
May 2016, 12(46):147-151
DOI:10.4103/0973-1296.182151  PMID:27279699
Background: Proteins present in the latex of Calotropis procera have been shown to produce anti-inflammatory effect and to afford protection in various disease models. Objectives: To determine the efficacy of high molecular weight protein sub-fraction (LPPI) of latex of C. procera in ameliorating joint inflammation and hyperalgesia in a preclinical model of arthritis. Materials and Methods: Monoarthritis was induced in rats by intra-articular injection of Freund's complete adjuvant (FCA) and the effect of two doses of LPPI (5 and 25 mg/kg) and diclofenac (5 mg/kg) was evaluated on joint swelling, stair climbing ability, motility, and dorsal flexion pain on day 3. The rats were sacrificed on day 3 to measure tissue levels of reduced glutathione (GSH) and thiobarbituric acid reactive substances (TBARS). Evaluation of joint histology was also made. Results: Intra-articular injection of FCA produced joint swelling and difficulty in stair climbing ability, motility, and pain on flexion of the joint as revealed by scores obtained for these functional parameters. LPPIproduced a dose-dependent decrease in joint swelling and improved joint functions. Arthritic rats also revealed altered oxidative homeostasis where joint tissue GSH levels were decreased and TBARS levels were increased as compared to normal rats. The levels of these oxidative stress markers were near normal in arthritic rats treated with LPPI. Moreover, treatment with LPPIalso maintained the structural integrity of the joint. The protective effect of LPPIwas comparable to the standard anti-inflammatory drug, diclofenac. Conclusion: The findings of the present study show that LPPIfraction comprising high molecular weight proteins could be used for the alleviation of arthritic symptoms. SUMMARY
  • High molecular weight protein sub.fraction of latex of Calotropis procera.(LPPI) reduced joint swelling and hyperalgesia in arthritic rats
  • LPPI produced a significant improvement in stair climbing ability and motility in arthritic rats
  • LPPI normalized the levels of oxidative stress markers in the arthritic joints
  • Treatment with LPPI reduced neutrophil influx and edema in the arthritic joints
Abbreviations used:FCA: Freund's complete adjuvant, GSH: Glutathione, TBARS: Thiobarbituric acid reactive substances, TBA: Thiobarbituric acid, MDA: Malondialdehyde, LPPI: Latex protein fraction PI.
  1,824 59 -
Simultaneous determination of eight bioactive compounds in Dianthus superbus by high-performance liquid chromatography
Bo-Ra Yun, Hye Jin Yang, Jin Bae Weon, Jiwoo Lee, Min Rye Eom, Choong Je Ma
May 2016, 12(46):264-269
DOI:10.4103/0973-1296.182159  PMID:27279718
Background: Dianthus superbus, one of traditional herbal medicine, is widely used to treat urethritis, carbuncles and carcinoma. Objective: A simultaneous determination method was established for controlling the quality of D. superbus using the eight compounds, (E)-methyl-4-hydroxy-4-(8a-methyl-3-oxodecahydronaphthalen-4a-yl) (1), diosmetin-7-O(2'',6''-di-O-α-L-rhamnopyranosyl)-β-D-glucopyranoside (2), vanillic acid (3), 4-hydroxyphenyl acetic acid (4), 4-methoxyphenyl acetic acid (5), (E)-4-methoxycinnamic acid (6), 3-methoxy-4-hydroxyphenylethanol (7), and methyl hydroferulate (8) isolated from D. superbus. Materials and Methods: This analysis method was developed using high performance liquid chromatography coupled with diode array detector with a Shishedo C18column at a column temperature of 3°C. The mobile phase was composed of 0.1% trifluoroacetic acid in water and acetonitrile. The flow rate was 1 ml/min and detection wavelength was set at 205 nm and 280 nm. Validation was performed in order to demonstrate selectivity, accuracy and precision of the method. Results: The calibration curves showed good linearity (R2 > 0.99). The limits of detection and limits of quantification were within the ranges 0.0159–0.6205 μg/ml and 0.3210–1.8802 μg/ml, respectively. Moreover, the relative standard deviations of intra- and inter-day precision were both <2.98%. The overall recoveries were in the range of 96.23–109.87%. Quantitative analysis of eight compounds in 12 D. superbus samples (D-1–D-12) from various regions were analyzed and compared by developed method. Conclusion: As a result, this established method was accurate and sensitive for the quality evaluation of eight compounds isolated from D. superbus and may provide a new basis for quality control of D. superbus. SUMMARY
  • A simultaneous determination method of eight compounds in Dianthus superbus was established by high performance liquid chromatography.diode array detector
  • Developed analysis method is validated with linearity, precious and accuracy
  • The newly established method was successfully evaluated contents of eight compounds in 12 D..superbus samples.(D.1.D.12) from various regions and compared.
Abbreviations used: HPLC: High performance liquid chromatography, LOD: Limits of detection, LOQ: Limits of quantification, RSD: Relative standard deviation.
  1,788 65 -
Storing of extracts in polypropylene microcentrifuge tubes yields contaminant peak during ultra-flow liquid chromatographic analysis
Parthraj R Kshirsagar, Harsha Hegde, Sandeep R Pai
April 2016, 12(46):303-306
DOI:10.4103/0973-1296.185719  PMID:27563216
Background and Aim: This study was designed to understand the effect of storage in polypropylene microcentrifuge tubes and glass vials during ultra-flow liquid chromatographic (UFLC) analysis. Materials and Methods: One ml of methanol was placed in polypropylene microcentrifuge tubes (PP material, Autoclavable) and glass vials (Borosilicate) separately for 1, 2, 4, 8, 10, 20, 40, and 80 days intervals stored at −4°C. Results: Contaminant peak was detected in methanol stored in polypropylene microcentrifuge tubes using UFLC analysis. The contaminant peak detected was prominent, sharp detectable at 9.176 ± 0.138 min on a Waters 250–4.6 mm, 4 μ, Nova-Pak C18 column with mobile phase consisting of methanol:water (70:30). Conclusion: It was evident from the study that long-term storage of biological samples prepared using methanol in polypropylene microcentrifuge tubes produce contaminant peak. Further, this may mislead in future reporting an unnatural compound by researchers.
  1,521 117 -