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Jul-Sep 2020
Volume 16 | Issue 5 (Supplement)
Suppl 3
Page Nos. 443-583

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ORIGINAL ARTICLES  

Embelin from Embelia ribes ameliorates oxidative stress and inflammation in high-fat diet-fed obese C57BL/6 mice Highly accessed article p. 443

DOI:10.4103/pm.pm_77_20  
Background: A safe, efficacious, and economical drug for the treatment of obesity is the need of the time. Literature published in previous years referred to embelin as a potential investigational therapeutic agent to manage obesity. Objectives: This study was designed to isolate and characterize embelin from Embelia ribes and further to assess its role in alleviating oxidative stress and chronic inflammation in the high-fat diet (HFD) fed obese C57BL/6 mice. Materials and Methods: Embelin extracted from berries of E. ribes with n-hexane by soxhlet extraction was characterized using high-performance thin-layer chromatography, infrared and liquid chromatography-mass spectrometry analyses. The obesity was induced by feeding of HFD for 8 weeks. Embelin (50 mg/kg/day, p.o.) was administered from 5th to 8th weeks along with HFD. After 8 weeks, the body weight gain and body mass index were calculated. Then, animals were sacrificed; serum and tissues were collected to further assess the various biomarkers of oxidative stress and inflammation. Results: The presence of embelin was confirmed using the above-mentioned analytical techniques. Treatment with Embelin showed amelioration of obesity biomarkers along with the substantial decline in levels of protein expression of nuclear factor erythroid 2-related factor and nuclear factor kappa-B in liver tissue. Treatment with embelin also normalizes the liver tissue levels of thiobarbituric acid reactive substances, glutathione, superoxide dismutase, and catalase. The histopathological analysis of liver tissue showed significant prevention of necrotic and inflammatory changes in embelin treated HFD fed mice. Conclusion: The results of the study clearly indicated the potential of embelin in ameliorating obesity by alleviating oxidative stress and inflammation induced by HFD in C57BL/6 mice.
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Enhancement of chlorogenic content of the eggplant fruit with eggplant hydroxycinnamoyl CoA-quinate transferase gene via novel agroinfiltration protocol Highly accessed article p. 450
Prashant Kaushik, Pankaj Kumar, Shashi Kumar
DOI:10.4103/pm.pm_537_19  
Background: Eggplant (Solanum melongena L.) is rich in health-promoting phenolic acids, primarily in the chlorogenic acid. For the production of chlorogenic acid in the eggplant hydroxycinnamoyl CoA-quinate transferase (SmHQT), is a central enzyme that catalyzes the reaction to the chlorogenic acid production. Objective: The function of SmHQT is not well determined in the eggplant fruit, and the fruit agroinfiltration procedure is not standardized for eggplant. Materials and Methods: Here, the overexpression of SmHQT in the eggplant fruit´s flesh was studied using the agroinfiltration technique. In our gene construct, we also co-expressed the P19 protein for overexpression, and the results were validated with real-time quantitative reverse transcription-polymerase chain reaction (qRT-PCR) and high-performance liquid chromatography (HPLC). Results: Due to the overexpression of the SmHQT gene, higher chlorogenic content was exhibited by the eggplant fruits, which was further validated by HPLC. The chlorogenic acid content after following the agroinfiltration procedure was more two times in the agroinfiltrated fruit. To identify the optimal target for increasing chlorogenic pathway flux post-SmHQT activity, expression patterns were analyzed with qRT-PCR, and the results showed the changes in the expression level of the other chlorogenic acid pathway genes. Furthermore, the cis regulating elements and protein-protein interaction (PPI) analyses supported the HPLC results. Conclusion: Overall, here, insights into the eggplant chlorogenic content increment at the molecular level and the opportunities for the improvement of chlorogenic content as nutrition in crops are provided.
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Effect of plant part, extraction method, and harvest time over antioxidant yield of rubus coreanus Highly accessed article p. 455
Min Ju Lee, Ju Hee Nam, Jeong Ho Jeong, Il Rae Rho
DOI:10.4103/pm.pm_549_19  
Background: Immature fruit of Rubus coreanus, also known as “Bokbunja” in Korea, have been used as traditional medicinal plant in East-Asia. Although several studies have been conducted to the fruit composition of R. coreanus, research on the antioxidant composition of branch and leaves are limited. Objectives: We aimed to analyze the contents of five antioxidants in leaves and branches of R. coreanus using different extraction method. Materials and Methods: R. coreanus were cultivated at the research farm of the Gyeongsang National University. R. coreanus plants were harvested in May and July, and leaf and branch extracts were prepared using the ultrasonic bath and reflux extraction methods and analyzed using liquid chromatography-tandem mass spectrometry. Results: The ultrasonic bath extraction method extracted 7.1 and 1.5 mg/100 g gallic acid from leaves and branches, respectively, whereas the reflux extraction method yielded 12.4 and 16.5 mg/100 g, respectively. Thus, reflux extraction was superior to ultrasonic bath for both leaf and branch parts. In the extracts prepared by reflux extraction, contents of all five compounds were higher in leaves than in branches. In leaves extracted with the reflux extraction method, ellagic acid was the most abundant, followed by quercetin-3-O-glucoside (24.7 mg/100 g), gallic acid (12.4 mg/100 g), quercetin (7.3 mg/100 g), and kaempferol (1.5 mg/100 g). Contents of all compounds were higher in May (1.1 mg/g) than in July (0.7 mg/g). Conclusion: R. coreanus plant was identified to show antioxidant activity and to present abundantly five antioxidants not only in fruit but also in leaves and branches in May. Specially, quercetin was three-fold higher in leaves than in fruit juice.
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Chromatographic investigation of phytoconstituents in milk yam (Ipomoea digitata L.) tubers p. 462

DOI:10.4103/pm.pm_116_20  
Background: Milk yam commonly called Ksheervidhari is a perennial climber; its tubers are an integral crude drug in different Ayurvedic formulations as well as folkloric medicine. It is used as antidiabetic, restorative, carminative, expectorant, galactagogue, stomachic, and appetizer. Objective: The present study is done with the aim of identifying the maturity stage of milk yam tuber containing maximum concentration of umbelliferone – a coumarin present in the tubers. Materials and Methods: Umbelliferone present in immature tubers (6 months after planting), overmature tubers (36 months after planting), and tubers at optimum maturity (21 months after planting) was assessed using high-performance liquid chromatography (HPLC) and high-performance thin-layer chromatography (HPTLC) analysis. Results: HPLC and HPTLC techniques revealed the presence of umbelliferone and it was quantified to be higher in over mature tubers (0.44 and 0.42 μg/g, respectively), followed by optimally mature (0.31 and 0.22 μg/g, respectively) and immature tubers (0.20 and <0.11 μg/g, respectively). Conclusion: The study reveales that developmental stages affect umbelliferone concentration in milk yam tubers and it increased as the plant ages.
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Rasam (South Indian Spice Soup) - Attenuates the Mammary Tumor Induction Magnitude of 7,12-Dimethylbenz[a] anthracene in Sprague–Dawley Rats p. 467

DOI:10.4103/pm.pm_150_20  
Background: Recently, we have reported rasam (South Indian spice soup) for antiproliferative activity against Michigan Cancer Foundation-7 cell lines. Objectives: Breast cancer seems to be more common in the younger age group. Considering the deviation of the younger group from the traditional food habits as one of the many reasons of breast cancer incidence, rasam was investigated for its chemopreventive effect on mammary carcinoma. Materials and Methods: Rasam at 3 and 4 mL/kg dose was administered to female Spraque–Dawley rats for 30 days before 7,12-dimethylbenz[a] anthracene (DMBA) induction of mammary carcinoma. Tamoxifen was used as the standard drug. Body weight, thiobarbituric acid reactive substances (TBARSs), phase I enzymes, hexokinase (HK), aldolase (AD), glucose 6-phosphatase (G6P), fructose 1,6-bisphosphatase (F16P), citric acid cycle (CAC) enzymes, histopathology, tumor weight, tumor latency, and tumor incidence were studied. Results: In all the studied parameters such as body weight, TBARS, phase I enzymes, HK, AD, G6P, F16P, and CAC enzymes, rasam-pretreated groups showed better prevention than tamoxifen-treated groups. Histochemical findings also clearly confirm the chemopreventive effect of rasam. Particularly at the dose of 4 mL/kg, rasam was efficient in reducing the percentage of incidence of tumor, number of tumor, and tumor weight. Conclusion: The present study reveals that rasam attenuates the mammary tumor induction magnitude of DMBA in female rats.
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Herniarin, a natural coumarin loaded novel targeted plasmonic silver nanoparticles for light activated chemo-photothermal therapy in preclinical model of breast cancer p. 474

DOI:10.4103/pm.pm_223_20  
Background: The conventional chemotherapeutic approach for breast cancer involves use of free drug substances that succumb into considerable systemic toxicity. For the past few decades biologically active phytochemicals like coumarins have gained much interest against breast cancer. However, there are lacunas in direct use of these potent biomolecules that impede their pharmacological functions. On the other hand, recent years have also witnessed great advancements in stimuli actuated nanotherapeutics as combined treatment strategies in critical breast cancer cases. Objectives: In this study we formulated novel pentagonal plasmonic silver nanoparticles as folate receptor targeted light responsive nano-delivery platform for a potent coumarin derivative, Herniarin to improve its therapeutic proficiency. Materials and Methods: Primarily, we synthesized plasmonic silver based nanoscopic carrier for herniarin to facilitate its tumor targeted delivery via folate receptors. These formulated nanoparticles (Herniarin quenched, folic acid furnished, HER-FA-AgNPs) were subjected to series of physicochemical tests, followed by detailed in-vitro and in-vivo testing in breast cancer cell lines and DMBA induced breast cancer model in rats, respectively. Results: The pentagonal nanoparticles observed in transmission electron microscopy demonstrated strong plasmonic tunability in the NIR region while its key feature of transforming light into heat energy when exposed to laser (800 nm) allowed photothermal therapy to supplement the chemotherapeutic potential of Herniarin as illustrated in various in-vitro studies. The in-vivo studies including detailed cellular investigations of breast tissues through Field Emission Scanning Electron Microscope and histopathology reflected the improved outcome of combined therapeutic modalities (hyperthermia + chemotherapy) achieved by HER-FA-AgNPs. Conclusion: Thus, these novel nanoparticles may serve as an efficient as well as safe drug-disposing vehicle for combined and strategic therapy of breast cancer.
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Method development on reverse-phase high-performance liquid chromatography for the improvement of routine analysis of flavonoids on agricultural and food products p. 486
Paranthaman Ramakrishnan, Chakkaravarthi Kamalanathan, Vidyalakshmi Rajagopal
DOI:10.4103/pm.pm_422_19  
Objectives: The objective of this work was to investigate the effect of reversed-phase high-performance liquid chromatography conditions for the determination of flavonoids in the plant extract. Methods: The analytical conditions (total flow rate, mobile phase, colum temperature, and detection wavelength) optimized for gallic acid, rutin, and quercetin. The optimized method was compared with the control method and evaluated with the use of reference compounds. Results: The result shows that the maximum flavonoids recovery was in observed on modified chromatographic conditions, Colum: Octadecyl-silica C18column (250 mm × 4 mm, 5 μm) Mobile Phase solution water-acetic acid (25:1) and methanol. Mobile Phase solution B was a Mobile phase – 4th min - 20%, 10th min - 80%, and a flow rate of 0.5 mL/min. Colum temperature 35°C and detection wavelength was 260 nm. Conclusion: In this work, the proposed method will increase the analysis time but minimize the mobile solvents and have the potential of being useful for the routine analysis of flavonoids.
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Effect of Vitex pinnata L. leaf extract on estrogenic activity and lipid profile in ovariectomized rats p. 492

DOI:10.4103/pm.pm_443_19  
Background: The genus Vitex contains about 300 species distributed around the world. These genuses are used in the treatment of premenstrual syndrome. However, their estrogenic activity is not well understood. Objectives: To compare the estrogenic activity and lipid profile of ethanol extracts of leaves of Vitex pinnata L. with 17 β-estradiol in bilaterally ovariectomized (OVX) rats. Methods: Ethanol extracts were analyzed by gas chromatography-mass spectrometry (GC-MS). Bilaterally OVX rats were divided into five groups,(n = 6) receiving different treatments, consisting of a vehicle (1% Tween), ethanol extract of V. pinnata at three different doses (100, 500, 1000 mg/kg) and standard drug, 17 β-estradiol at a dose of 1 mg/kg. All groups were administered orally, daily for 14 days. Results: GC-MS data revealed that the major chemical constituents of the extract were 3, 7, 11, 15-Tetramethylhexadecen-2-en-1-ol, Gamma-Stigmasterol, 9,12,15-octadecatrienoic acid and n-hexadecanoic acid. V. pinnata extracts at 1000 mg/kg slightly increased uterine and vaginal weight and endometrial thickness. Doses of extract at 500 and 1000 mg/kg induced a significant (P < 0.05) decrease of triglycerides and total cholesterol in serum of OVX rats. Conclusion: V. pinnata leaf extract exhibits estrogenic activity and reduces levels of serum triglycerides and cholesterol. The understanding of such activity of V. pinnata leaf extract has benefits for postmenopausal women.
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In vitro evaluation of Hydrilla verticillata for anti-adipogenesis activity on 3T3 L1 cell lines p. 498

DOI:10.4103/pm.pm_234_20  
Background: Obesity is a metabolic disorder that has reached pandemic proportions worldwide. Phytol, a diterpene isoprenoid alcohol found as an integral part of chlorophyll of plants, algae, and guts of ruminant animals, is proven for its lipid-lowering activity. This compound is largely present in the Indian aquatic medicinal plant, Hydrilla verticillata. The research is focused to screen the anti-adipogenesis activity of H. verticillata extract on 3T3 L1 cell lines. Materials and Methods: The ethanolic extract of H. verticillata was prepared and characterized by gas chromatography–mass spectroscopy and Fourier transform infrared spectroscopy analysis. In vitro cytotoxic effect of H. verticillata extract on 3T3 L1 cell line was evaluated by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide test. The ethanolic solvent extract of this plant is screened for anti-adipogenesis activity by performing in vitro pancreatic lipase inhibition and Oil Red O staining assay. Results: Phytochemical analysis and characterization of the extract showed the presence of a large amount of phytol in H. verticillata. In vitro pancreatic assay of the crude extract showed maximum lipase inhibition activity at a minimal concentration of 125 μg/mL, similar to the standard anti-obesity drug, orlistat. The half-maximal inhibitory concentration (IC50) of the extract on 3T3 L1 cell line was 840.91 μg/mL. Cells treated with one-third IC50showed minimal lipid accumulation, which is determined by Oil Red O staining. Conclusion: This study confirms that the ethanolic extract of H. verticillata possesses anti-adipogenesis activity, which can be used to control obesity-related treatments.
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Pueraria candollei var. mirifica-Induced CYP1A1 and CYP1A2 expression in human choriocarcinoma bewo cells p. 506

DOI:10.4103/pm.pm_164_20  
Background: The human placenta metabolizes many endogenous substances, drugs, and xenobiotics. Cytochrome P450 family 1 (CYP1) is expressed in both early- and full-term placenta. The Thai medicinal plant Pueraria candollei var. mirifica (PM) is traditionally consumed for rejuvenation and has neuroprotective, anti-osteoporotic, and antioxidant activities. Objectives: The objective of this study was to compare the effects of PM and the CYP1A inducer β-naphthoflavone (BNF) on the expression of CYP1, aryl hydrocarbon receptor (AHR), AHR nuclear translocator (ARNT), and the transporter ABCG2. Materials and Methods: Human choriocarcinoma BeWo cells were treated with BNF (10 μM), ethanolic extract of PM (PM-EtOH), or column chromatographic fractions of PM-EtOH (F2, F4, and F6) at 1, 10, and 100 μg/mL for 24 h. The mRNA expression of target genes was determined using real-time quantitative polymerase chain reaction. The activity of ethoxyresorufin-O-deethylase (EROD), a marker for CYP1, was measured at the RNA harvesting time point. Results and Discussion: PM-EtOH, F2, and F4 significantly induced EROD activity and expression of CYP1A1 and CYP1A2 while CYP1B1 and AHR were slightly suppressed and ARNT was unchanged. ABCG2 was slightly induced by F2. Therefore, the expression of CYP1 in BeWo cells appears to be independent of the AHR/ARNT regulatory pathway. Conclusion: The use of PM-containing products at high quantities or for long periods during pregnancy is of concern due to likely herb–drug interactions and toxicological risks through activation of CYP1A1 and CYP1A2 transcription.
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A pentacyclic triterpene from Lippia origanoides H.B.K and its cytotoxic activity p. 513

DOI:10.4103/pm.pm_218_20  
Background: Lippia origanoides H. B. K. (Verbenaceae) is an aromatic small shrub appreciated in the traditional systems of medicine. L. origanoides essential oil is an ingredient of commercial poultry feed products and its postdistillation residual biomass is an interesting source of bioactive compounds. During our search for the valorization of this residual biomass, supercritical-CO2(SC-CO2)extraction afforded a mixture that was subjected to an investigation of phytochemicals and of cytotoxicity, which was not reported previously. Objectives: The current study was designed to investigate the phytochemicals from the steam-distilled residual biomass of thymol- and carvacrol-rich L. origanoides chemotypes and to evaluate their in vitro cytotoxic activity. Materials and Methods: Steam-distilled aerial parts of L. origanoides chemotypes were extracted with SC-CO2to obtain a greenish-yellow extract with a strong aromatic odor. The SC-CO2extract was subjected to column chromatography, and the isolate obtained was screened in vitro for cytotoxicity against human normal embryonic kidney 293, MRC-5, THP-1, and XP4PA cell lines, using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) assay. Results: A pentacyclic triterpene, friedelan-3-one (1) was isolated for the first time from L. origanoides chemotypes. The structure of the isolate was elucidated with spectroscopic (ultraviolet, infrared, mass spectra, and nuclear magnetic resonance) techniques. The in vitro cytotoxic activity of the isolated compound was determined. The results showed no significant results against the selected cell lines using the MTT assay. Conclusion: The main significance of the present study was to develop promising routes to utilize the residual biomass for value-addition. This is the first report of friedelan-3-one isolation from the genus Lippia.
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Extraction and volatile compounds profiling of the bioactive fraction of Monochoria hastata (L.) solms p. 517

DOI:10.4103/pm.pm_386_19  
Background: An aquatic macrophyte Monochoria hastata (L.) Solms has ethnomedicinal application in various ailments and experimentally is proved to have antibacterial property. Objectives: To evaluate the optimized extraction methods and solvents for antibacterial activity and their characterization by gas chromatography-mass spectrometry (GC-MS). Materials and Methods: Extractions from the aerial parts of the plant in solvents with different polarities and various techniques, viz., microwave, infusion, Soxhlet, and maceration, were used to evaluate the best antibacterial efficacy by agar well diffusion method. The phytochemical constituents present in the bioactive extract were analyzed using standard phytochemical screening methods and characterized by GC-MS analysis. Results: Ethyl acetate extract derived by the Soxhlet method showed the highest antibacterial activity against all the test bacterial strains, and Gram-positive strains were more susceptible than Gram-negative strains. The crude extracts showed antibacterial activity which ranged from 7.0 ± 0.3 to 16.5 ± 0.8 mm at 100 mg/ml. Fifty percent of methanol had the highest extractive value (21.3%) in the Soxhlet method. Phytochemical tests showed the presence of alkaloids, phenols, flavonoids, terpenoids, glycosides, and fats in the bioactive ethyl acetate extract. GC-MS analysis showed the presence of different fatty acids and their saturated esters as the principal components. The major compounds were tridecanoic acid, methyl ester, 2-hexyldecanoic acid, dodecanoic acid, and diethyl phthalate. Conclusion: The results suggest that the ethyl acetate extract of M. hastata (L.) Solms possesses some bioactive volatile compounds including fatty acids and their esters, which have antibacterial potentiality. This is the first report of such antibacterial assessment from M. hastata (L.) Solms.
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Effect of ethanolic extract of Carica papaya Leaves and their cytotoxicity and apoptotic potential in human ovarian cancer cell lines- PA-1 p. 524
Priyadurairaj , Palagati Rohith Kumar Reddy, Palaniyandi Thiruvanavukkarasu, Sindhu Rajesh, Suganya Karunakaran, Rajeswary Hari
DOI:10.4103/pm.pm_117_20  
Introduction: The aim of the present study is to investigate the efficacy of Carica papaya ethanolic extract for antiproliferative, cytotoxicity, apoptotic, cell cycle blockade and wound healing in the ovarian cancer PA-1 cell lines. Materials and Methods: PA-1 cells were treated without the sample (act as a control) and with the sample for the above assay as per the described protocol. Results: The end result antiproliferative showed that PA-1 cell viability decreases in a concentration dependent manner and the growth inhibitory effect (IC50) values are obtained at a concentration of 100 μg. The increase in number of apoptotic cell was observed and after treating with PA-1 cells the sample with double staining methods. G2/M phase of the cell cycle was significantly blocked by the test sample followed by the S phase in a negligible manner. And in vitro cell wound closure or contracture was not significant when compared the sample against control group. Conclusion: C. papaya ethanolic extract showed to possess excellent cytotoxic effect through inducing apoptosis especially causing arrest at the G2/M phase of the cell cycle.
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Germinated seeds of Hordeum vulgare target extrinsic pathway of apoptosis in triple-negative breast cancer cells p. 531

DOI:10.4103/pm.pm_123_20  
Background: The absence of receptors, lack of specific treatment regimen, and emergence of resistance against various currently available anticancer drugs have led to the development of lead molecules from botanicals for the mainstay treatment of triple-negative breast cancer (TNBC). Hordeum vulgare, commonly known as barley, has been reported to have many traditional uses, and the main alkaloid in its germinated seeds, hordenine, has been evaluated for many pharmacological properties. Objectives: The present study aimed to evaluate the anticancer activity of methanol extract of germinated seeds of H. vulgare (MGHV) against MDA-MB-231 TNBC cells, quantify hordenine in MGHV, and derive the probable target of action of the extract and hordenine. Materials and Methods: 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay, morphological evaluation, acridine orange/ethidium bromide (AO/EB) staining, Hoechst 33258 staining, JC-1 staining, comet assay, and Western blot were conducted to evaluate the anticancer effect. High-performance liquid chromatography (HPLC) analysis was conducted to detect and quantify the presence of hordenine in the extract. Stimulation of extrinsic pathway of apoptosis was evaluated by studying the interaction of hordenine with Caspase-8 using in silico methods. Results: MTT assay revealed significant concentration-dependent cytotoxicity. AO/EB staining exhibited yellow-green fluorescence indicative of early apoptosis. Hoechst 33258 staining showed nuclear marginalization and fragmentation. The results of JC-1 staining showed a combination of red and green fluorescence indicative of partial reduction in mitochondrial membrane potential. Comet assay revealed that the extract did not produce deoxyribo nucleic acid damage. Western blotting analysis did not show any change in the expression of Bcl-2, whereas significant upregulation of Caspase-8 by 2.8 folds was noticed indicative of extrinsic pathway of apoptosis. HPLC analysis detected the presence of hordenine as the major constituent in the extract. Morphological assessment of MDA-MB-231 cells treated with hordenine showed cytotoxic changes similar to that of the extract. In silico studies with hordenine also substantiated the results of Western blot. Conclusion: Thus, it was concluded that MGHV targeted extrinsic pathway of apoptosis due to the presence of hordenine and hence hordenine could evolve as a potent anticancer molecule against TNBC.
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DNA barcoding markers to identify intraspecies genetic variations in three ecotypes of Abrus Precatorius L. p. 540

DOI:10.4103/pm.pm_23_20  
Background: Abrus precatorius is also called as Gunja in Ayurveda. It is a commonly grown plant belonging to the family of Fabaceae. It is characterized under the Upavisha (semi-poisonous drugs) and used widely in various Ayurvedic formulations with great beneficial significance. It is used in the treatment of various diseases such as alopecia, edema, helminthiasis, skin diseases, itching, and urinary disorders after being passed through specific purification procedures. Objectives: The present study aims to compare three different varieties of A. precatorius which includes the species of white, black, and red which are used to study DNA barcoding marker and phylogenetic analysis. Materials and Methods: Whole genomic isolated DNA from three varieties (white, black, and red) of A. precatorius leaves and subjected to analysis of the polymerase chain reaction of rbcL, maturase K, and internal transcribed spacer 4,5 using 0.8% agarose gel electrophoresis. Results: The DNA barcoding markers and next-generation sequencing can identify the intraspecies genetic variations among these closely related plant varieties of A. precatorius of white, black, and red. Conclusion: The intraspecies genetic variations among these three varieties of A. precatorius white, black, and red are closely related with A. precatorius isolate TMP 144 Ribulose 1,5-bisphosphate carboxylase/oxygenase large subunit gene 99.81%.
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Anti-cancer activity of ethanolic leaf extract of Salvia officinalis against oral squamous carcinoma cells in vitro via caspase mediated mitochondrial apoptosis p. 546
Prasanna Rajagopalan, Shadma Wahab, Ayed A Dera, Harish C Chandramoorthy, Safia Irfan, Ayyub Ali Patel, Shahabe Saquib Abullias, Gaffar Sarwar Zaman, Irfan Ahmad
DOI:10.4103/pm.pm_90_20  
Aim: Need for novel agents that fight oral squamous cancer is on constant demand. The current study aims in the identification of active phytochemical ingredients from Salvia officinalis leaf extract (SOLE) and evaluation for anticancer properties in oral squamous carcinoma cells. Materials and Methods: Soxhlet method was used for SOLE extraction. Phytochemical tests and thin-layer chromatography (TLC) were performed for active compounds identification. Oral squamous cancer cells (SSC-15 and SSC-25) were used to assess anticancer efficacy. MTT analysis was utilized for the viability of cells. The utilization of flow cytometry was done to assess the changes in cell cycle and apoptosis induction. Western Blotting method was used to analyze the expressions of apoptotic protein. Results: Preliminary phytochemical screening showed the presence of sterols, flavonoids, and tannins. TLC study revealed the presence of rutin in SOLE. The extract inhibited cell proliferation of SSC-15 and SSC-25 cells with GI50values of 340.7 μg/ml and 287.7 μg/ml, respectively. SOLE inhibited the migration of these cells across the endothelial membrane and induced nuclear fragmentation in cancer cells. Analysis of the cell cycle revealed SOLE to increase the sub G0/G1population in SSC-15 and SSC-25 cells. SOLE increased both early and late phase apoptosis in both oral squamous cancer cell lines. Apoptotic markers such as caspase-3, Bax and P-53 were found to be dose-dependently increased with SOLE treatment in both the tested cell lines, while the anti-apoptotic Bcl-2 protein was decreased. Conclusion: In summary, SOLE demonstrated excellent anticancer and anti-migration efficacy in oral squamous cancer cells through caspase-mediated mitochondrial apoptosis. More than one compound might be responsible for the activity, which deserves further research.
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Binding of metronidazole to Enterococcus faecalis homoserine kinase: Binding studies, docking studies, and molecular dynamics simulation studies p. 553
Harpreet Singh, Satyajeet Das, Pramodkumar P Gupta, Sagar Batra, Richa Prakash, Vijay Kumar Srivastava, Anupam Jyoti, Vikas Gupta, SL Kothari, Sanket Kaushik
DOI:10.4103/pm.pm_99_20  
Background: Enterococcus faecalis (Ef) is an opportunistic virulent bacterial pathogen resistant to a diverse class of antibiotics by possession of a wide range of resistance mechanisms. Aim and Objective: Looking at the increasing number of infections caused by Ef, it is essential to develop alternative strategies to fight against Ef. In this regard, homoserine kinase (HSK) is an important enzyme of threonine biosynthesis pathway in Ef. Threonine is an essential amino acid, and HSK catalyzes the formation of O-phospho-L-homoserine production, which is an important step in threonine metabolism. Therefore, Enterococcus faecalis homoserine kinase (EfHSK) becomes an ideal target for antimicrobial drug development. Methodology: We report binding studies, docking studies, and molecular dynamics (MD) simulation studies of EfHSK. Fluorescence spectroscopy studies indicated the binding of metronidazole with EfHSK. Result: Docking studies further showed that amino acid residues such as Asn124 and Gly83 in the phosphate-binding loop of EfHSK play a vital role in the formation of H-bonds with the ligand metronidazole. The site where ligand was bound is a deep groove, which is regarded as the binding cavity of the protein. Docking studies were further confirmed by MD simulation studies. Conclusion: Metronidazole might be considered as a potential inhibitor of EfHSK, as this occupies the binding pocket and eventually can reduce the kinase activity of this enzyme.
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Effects of Artemisia judaica essential oil and ethanolic extract on experimentally-induced benign prostatic hyperplasia p. 561
Bahaa Al-Trad, Mazhar Al Zoubi, Mu'ath Migdady, Jamil Lahham, Alaa A A Aljabali, Malek Shehab, Sahar Alomari, Mahmoud A Al-Qudah, Janti Qar, Riyadh Muhaidat, Ahmad El-Oqlah, Murtaza M Tambuwala
DOI:10.4103/pm.pm_216_20  
Background: Recent studies have shown that the essential oil (EO) and the ethanolic extract (EE) from Artemisia judaica L., a Jordanian medicinal plant, exhibit a potent anti-angiogenic and anti-inflammatory activities. Angiogenesis and inflammation processes are known to play a role in benign prostatic hyperplasia (BPH), a disease associated with aging in men. Objectives: The present study aimed to address the effects of EO and EE on experimentally-induced BPH in rats. Materials and Methods: Four experimental groups were assigned with six rats in each group, including the corn oil vehicle as a control. The three other groups were induced to develop BPH by testosterone injection. The BPH rats were randomized into the BPH-untreated group, BPH-EO treated group (200 mg/kg/subcutaneously) and BPH-EE treated group (500 mg/kg/orally). Results: The prostate weight/body ratio and epithelial thickness showed a significant reduction in the EO and EE treated groups compared to the BPH untreated. In addition, mRNA expression levels of the proliferating marker (proliferating cell nuclear antigen), the angiogenesis marker (vascular endothelial growth factor-A) and interleukin-6; an inflammatory cytokine, were significantly down-regulated in the BPH groups that were treated with EE or EO. Conclusion: Our results indicated that in experimentally-induced BPH, EO and EE from A. judaica ameliorate BPH development by inhibiting prostatic cell proliferation, angiogenesis, and inflammation.
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Protective effect of Viburnum grandiflorum against ultraviolet-B radiation-induced cellular and molecular changes in human epidermal keratinocytes p. 566
Hanjun Liu, Haixiu Zhang, Minyan Dang, Yukiat Lin, Hui Yan
DOI:10.4103/pm.pm_397_19  
The aim of the present study was to evaluate the photoprotective potential of Viburnum grandiflorum (VG) against ultraviolet-B radiation-induced responses in HaCaT cells. The HaCaT cells were pretreated with VG prior ultraviolet-B (UVB)-radiation exposure and were further examined for lipid peroxidation, enzymatic antioxidant activity, % reactive oxygen species, DNA damage, mitochondrial membrane potential and for inflammatory, and apoptotic signaling markers such as tumor necrosis factor-alpha, nuclear factor kappa B, interleukin-1 (IL-1), IL-6, cyclooxygenase-2, p53, caspase-3/9, cytochrome-c, Bax, and Bcl-2. The VG pretreatment in UVB exposed cells shows significantly regulated both inflammatory as well as apoptotic signaling cascades. Our findings suggest that VG may be functional against UVB-induced photo-damages.
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Antioxidant activity and inhibitory effect on nitric oxide production of rang chuet (Thunbergia laurifolia Lindl.) leaf extracts in lipopolysaccharide-stimulated BV2 microglial cells p. 573
Nootchanat Mairuae, Poonlarp Cheepsunthorn, Benjaporn Buranrat
DOI:10.4103/pm.pm_44_20  
Background: Oxidative stress and neuroinflammation mediated by microglial activation play a significant role in the pathogenesis of neurodegenerative diseases. Therefore, negative regulators of microglial activation have been identified as potential therapeutic candidates for the treatment of such diseases. Objectives: The aim of this study was to investigate the effects of Thunbergia laurifolia Lindl. or Rang Chuet (RC) extracts on reactive oxygen species (ROS) and nitric oxide (NO) production in lipopolysaccharide (LPS)-activated BV2 microglia cells. Materials and Methods: BV2 cells were treated with LPS in the presence or absence of RC extract, and the levels of ROS and NO were measured using CM-H2DCFDA and Griess reagent assay, respectively. The nuclear levels of nuclear factor kappa B (NF-kB) p65 and the activation of Akt and the extracellular signal-regulated kinase 1/2 (ERK1/2) were detected using immunofluorescence and western blotting assay, respectively. Results: The treatment of BV2 cells with RC concentration-dependently suppressed the LPS-induced ROS production compared to the control group. RC treatment also significantly decreased LPS-induced NO production in a concentration-dependent manner. In addition, RC treatment also suppressed NF-kB p65 translocation and the activation of Akt and the ERK1/2 pathway in LPS-stimulated BV2 cells. Conclusion: Taken together, these results demonstrate that RC exerts anti-oxidative activity by suppressing ROS production and that it exerts antioxidative activity by suppressing NO in LPS-stimulated BV2 cells. The mechanisms by which RC suppresses NO and ROS production may occur through inhibition of Akt and the ERK1/2-mediated NF-κB pathway. The results suggest that RC may be useful in treating neurodegenerative diseases mediated by microglial cells.
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An integrated approach of network based system pharmacology approach and molecular docking to explore multiscale role of Pinus roxburghii and investigation into its mechanism p. 578
Ankur Omer, Poonam Singh
DOI:10.4103/0973-1296.301874  
Background: Pinus roxburghii extract is a multi composed mixture of constituent compounds known to be used as an alternative medicine to treat various diseases. However, due to complex nature the knowledge about its therapeutic range and mechanism of action is still in its infant state. By interpreting the behaviour pattern of complex regulatory networks during various states such as healthy, diseased etc the chances are increased for new target identification, and disease prediction. Objective: Therefore, it is necessary to exploit the regulatory network of Pinus plant to elucidate its mechanism of action and prospective roles. Materials and Methods: In the present work, chemical prediction, target prediction, network biology, virtual screening, binding affinity and molecular docking approaches have been integrated to unravel the multiple potentials of the Pinus plant. Results: The dataset contained constituent molecules and 121 targets that are mapped to drug-target and drug-target-pathway networks to elucidate the relationships amongst the constituent molecules and the targets. The HSD17B1 target with the central role in Pinus regulatory network was screened as the aspiring target; the predicted association between the target and the molecules was validated with the help of molecular docking to obtain Pinus plant constituents (Rutin, Cedeodarin and Isorhamnetin) to treat Cancer and Epilepsy. Conclusion: The Pinus plant shows potential to show anti-cancer activity although the experimental validation will definitely be required but the study shows the path to work in the similar area
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