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   Table of Contents - Current issue
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April 2021
Volume 17 | Issue 6 (Supplement)
Suppl 2
Page Nos. 123-291

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ORIGINAL ARTICLES  

Pharmacognosy and pharmacology of Calotropis gigantea for discovery of anticancer therapeutics Highly accessed article p. 123
Zahra Khosravi, Arun H S. Kumar
DOI:10.4103/pm.pm_166_21  
Background: Calotropis gigantea (CG) is a shrubby plant which is traditionally used for the treatment of diseases affecting the nervous system, digestive system, and skin. Several bioactive compounds from CG are isolated and investigated for their pharmacological properties. Methods: A systematic analysis of PubMed, Google Scholar, and PubChem database was performed, and the pharmacognosy and pharmacological properties of CG compounds were correlated. Results: Major phytochemicals such as those of the cardenolide, uscharin, and calotropin family were dominant in the CG plant. Phytochemicals found in the leaves, root, and latex of CG plant showed selective cytotoxicity, proapoptotic effects, and cell cycle arrest potential by modulating hypoxia-inducible factor 1 α, inhibitor of nuclear factor kappa-B kinase subunit beta (IKK-β), Notch, and Wnt signaling. Conclusion: The selective cytotoxicity against neuronal tumors, together with their neurogenesis/synaptogenesis potential and drug candidate like features merit the development of CG compounds as therapeutics for neuronal tumors.
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Evaluation of Acacia auriculiformis Benth. leaves for wound healing activity in type 2 diabetic rats p. 129
Naresh Kumar Rangra, Subir Samanta, Kishanta Kumar Pradhan
DOI:10.4103/pm.pm_496_20  
Background: Acacia auriculiformis Benth. is a perennial shrub which has been traditionally known to treat various medical complications such as sore eyes, allergy, rheumatism, sore eyes, and rashes. Objectives: The current study is designed to decipher a shred of clinical confirmation and biochemical support for wound-healing efficacy of methanolic leaves extract of A. auriculiformis. Materials and Methods: We prepared the leaves extract in different solvents, i.e., petroleum ether, chloroform, ethyl acetate, acetone, butanol, and methanol. The in vitro studies were carried out on the above leaf's extracts using α-glucosidase and α-amylase inhibition in vitro antidiabetic assays. Animal model of streptozotocin-induced diabetes was used in the 0-, 7-, and 15-day studies, respectively. The comparison study was carried out in diabetic wound control in respect of the period of epithelialization, %wound contraction, and hydroxyproline content in the excision wound model. However, the breaking strength parameter was used to calculate healing potential in the incision wound model. The bioactive methanol extract was subjected to LC-MS/MS analysis to characterize the phytoconstituents responsible for pharmacological activity. Results: The methanolic leaves extract showed the highest percentage inhibition of 94.259% and 95.259% in α-glucosidase and α-amylase in vitro antidiabetic assays, respectively. The high content of collagen fibers and stronger epithelial cells growth were observed in histopathological studies of hydrogel containing methanolic leaves extract as compared to the diabetic wound control and standard. Subsequently, for investigating the biological impact upon live cells, cytotoxicity study was tested in different cell lines (A549, HEK-293, and MCF-7). Cytotoxic results showed that greater than 75% of cells were visible in all the cell lines, which gives the confirmation of biocapability of the extract. The LC-MS/MS results revealed the presence of compounds such as β-sitosterol, lupeol, stigmasterol, and quercetin. Conclusion: A. auriculiformis is a potent medicinal plant that can be further utilized as a complementary and alternative therapy for the treatment of diabetes-induced wounds and the management of oxidative stress and diabetes.
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Ethanolic extract of Capparis decidua fruit ameliorates methotrexate-induced hepatotoxicity by suppressing oxidative stress and inflammation by modulating nuclear factor-kappa B signaling pathway p. 143
Preetha Santhakumar, Anitha Roy, M Karthik Ganesh, J Selvaraj, Lavanya Prathap, K Yuvaraj Babu
DOI:10.4103/pm.pm_402_20  
Background: Methotrexate is a stable derivative of aminopterin, a folate antimetabolite. Despite the widespread use of methotrexate, the clinical application of methotrexate is limited and restricted by the occurrence of hepatotoxicity. An important medicinal plant, Capparis decidua contains multiple pharmacological activities such as antioxidant, antiapoptotic, and anti-inflammatory effects. Objectives: This study investigated the protective effect of C. decidua against methotrexate-induced hepatotoxicity, focusing on its ability to attenuate oxidative stress and inflammatory nuclear factor-kappa B (NF-κB) signaling pathway which have not been studied earlier. Materials and Methods: Thirty-six female Wistar rats were randomly divided into six experimental groups. Rats were treated with C. decidua orally with doses of 250 mg/kg and 500 mg/kg for 14 consecutive days following a single dose of MTX injection (20 mg/kg, i. p). Serum levels of aspartate aminotransferase (AST), alanine aminotransferase (ALT), and alkaline phosphatase (ALP) were assessed. Malondialdehyde (MDA) levels, superoxide dismutase (SOD), and catalase (CAT) were assessed in hepatic tissue. mRNA expression of, NF-κB, tumor necrosis factor-alpha (TNF α), interleukin-1beta (IL-1 β), and IL 6 was carried using real-time polymerase chain reaction in hepatic tissue. Results: Results showed that methotrexate administration significantly increased AST, ALT, and ALP levels. It has also increased MDA levels indicating lipid peroxidation and decreased antioxidants levels of SOD and CAT levels. Treatment with C. decidua significantly diminished the hepatotoxic effects of methotrexate which was seen in a significant decrease in AST, ALT, ALP, MDA levels, significant increase in SOD and CAT levels. Methotrexate provoked hepatic NF-κB phosphorylation and increased mRNA abundance of TNF-α, IL-1 β, and IL-6 expression. Conclusion: These findings suggest that C. decidua prevents methotrexate hepatotoxicity through suppression of NF-κB signaling pathway, attenuating oxidative damage, inflammation, and cell death.
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Antioxidant and antiacetylcholinestrase studies of In vitro regenerated and transformed hairy roots of Ocimum sanctum (L.) p. 151
Annie Jessica Toppo, Sheela Chandra
DOI:10.4103/pm.pm_539_20  
Background: Ocimum sanctum (L.) is a well-known extensively employed Indian medicinal shrub in traditional and modern therapeutics. Hairy roots exhibit similar or superior capability to inhibit oxidants responsible for many disorders. Objectives: Agrobacterium-mediated transformation protocol has developed for O. sanctum which influences T-DNA delivery. The antioxidant, antiacetylcholinesterase (AChE) potential, and in vivo study of hairy root extract of O. sanctum has been evaluated. Materials and Methods: Sterilized O. sanctum shoot tips had grown in vitro by employing Murashige and Skoog basal media, Benzyl amino purine (BAP) (0.5 mg/L), and other additives. In vitro grown leaf explants were infected by Agrobacterium rhizogenes ATCC 15834 for various time durations. Polymerase chain reaction (PCR) employed to confirm the genetic assimilation of Agrobacterium. The hairy root extract's antioxidant potential has been tested by diphenyl picric hydroxyl (DPPH) and ferrous ion reducing assay (ferrous reduced antioxidant power [FRAP]) along with anti-AChE potential. In vivo study was done employing behavioral and biochemical estimation. Results: For in vitro culturing of O. sanctum, administration of BAP (0.5 mg/L) shows growth rate of 32.7% ±0.19% in 7 days. The Agrobacterium-infected explants show the good yield of hairy root in 1 h of infection duration. The PCR results show the genomic integration of transgene rol A and rol C. The methanolic extracts of hairy roots showed potential antioxidant activities for DPPH-free radical (66.15% ±0.34%) and FRAP assay (226.35 ± 0.17TE/gDW of the sample). These extracts effectively inhibit AChE (28.710 ± 0.26 μg/mL at 3 min incubation duration). In in vivo study, the pretreatment of OSM (200 and 400 mg/kg) exhibited significant increase of spatial and long term after administration of scopolamine. Conclusion: The obtained results exhibited a simple protocol for micropropagation and transgenic roots production with competently targeting AChE and related disorders.
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Bioprospecting of Lobelia nicotianifolia Roth. plant parts for antioxidant and cytotoxic activity and its phytoconstituents p. 162
Rupali Mukesh Kolap, Kailash D Datkhile, Saurabha Bhimrao Zimare
DOI:10.4103/pm.pm_398_20  
Background: Though the Lobelia nicotianifolia Roth. is ethnobotanically important plant of India and Sri Lanka its phytoconstituents, antioxidant, and anticancer potential was not yet reported. Objective: The objective of this study is to analyze the phytoconstituents of plant parts of L. nicotianifolia and to determine its antioxidant and cytotoxic potential. Materials and Methods: The plant parts of L. nicotianifolia were extracted with different solvents and qualitative analysis revealed the presence of different phytoconstituents. Total phenolic content (TPC) and total flavonoid content (TFC) were recorded in all plant parts. The extracts were subjected to the antioxidant assays and the potent methanolic extracts were used for cytotoxicity study and further characterized by Fourier–transform infrared spectroscopy and liquid chromatography with a high resolution mass spectrometer (LC–HRMS). Results: The qualitative analysis showed the presence of a wide array of phytoconstituents in L. nicotianifolia plant parts. A significantly higher TPC, TFC, and antioxidant activities were seen in methanolic stem extract. Stem extract showed maximum cytotoxicity against human breast adenocarcinoma (MCF–7) and human cervical adenocarcinoma (HeLa) cell lines whereas, root extract had higher cytotoxicity against human colon adenocarcinoma (HCT–15) cells. The results of cell viability indicated that the methanolic extracts of L. nicotianifolia plant parts exhibited a range of cytotoxic activity in a concentration and time dependent manner against selected cancer cell lines. The LC–HRMS showed the presence of cytotoxic compounds comparatively higher in stem. Conclusion: The study confirms the antioxidant and cytotoxic potential of L. nicotianifolia. To understand the detailed mechanism of cytotoxicity of L. nicotianifolia, it is necessary to study the molecular mechanism involved in this study.
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Vernonia anthelmintica (L.) willd extract alleviates cognitive deficits and neurodegeneration induced by infusion of amyloid beta (1–42) in rats p. 172
Swati Som, Justin Antony, S Palanisamy Dhanabal, Sivasankaran Ponnusankar
DOI:10.4103/pm.pm_518_20  
Background: There is a constant hunt for the development of new therapeutic alternative to address Alzheimer's disease (AD) due to its failing attempts in the evolution of new therapies and suboptimal results from the existing pharmacological interventions for the treatment of this severe neurodegenerative disease. Vernonia anthelmintica is an extensively used medicinal plant in Indian traditional medicine with wide range of therapeutic values. However, there is insufficient scientific documentation available for its protective effect against cognitive disorders. Objectives: The objective of the current investigation is to explore the neuroprotective activity of methanolic extract of V. anthelmintica (MEVA) in amyloid-beta (Aβ) (1–42) infused sporadic model of AD. Materials and Methods: Adult healthy male Wistar rats were treated orally with 250 mg/kg and 500 mg/kg of MEVA for 28 days, after a week from intracerebroventricular (i.c.v) infusion of Aβ (1–42) peptides followed by assessment of neurobehavioral deficits. Subsequently, animals were euthanized and brains were collected for estimation and quantification of neurochemical biomarkers including antioxidant enzymes, neurotransmitters, plaque load, and inflammatory mediators. Results: Dose-dependent reversal of cognitive impairment was observed upon MEVA treatment in amyloid intoxicated rats as corroborated by improved learning and memory and diminished oxidative stress, cholinergic hypofunction, and neuroinflammation induced by Aβ (1–42). Conclusion: Collectively, evidence-based data suggested the promising neurotherapeutic potential of V. anthelmintica and thereby can stand as a novel entity for curbing AD pathology.
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Effect of solvent extraction system on the antioxidant properties of seven wild edible mushrooms and identification of phenolic compounds by high-performance liquid chromatography p. 180
Ishika Bera, Bejoysekhar Datta, Kanad Das, Tapan Seal
DOI:10.4103/pm.pm_584_20  
Objectives: The main objective of this research was to analyze some selected indigenous wild mushrooms in Arunachal Pradesh, namely, Lactifluus pilosus, Lactifluus piperatus, Lactifluus indovolemus, Lactifluus crocatus, Lactarius hirtipes, Lactarius kesiyae and Lactarius viridinigrellus for their novel anti-oxidant components in two different solvent extracts. Materials and Methods: The antioxidant properties of water and methanol extracts of these mushrooms were determined by the total phenolics and flavonoid content, reducing power capacity, radical scavenging activity by 1,1-diphenyl-2-picrylhydrazyl and (2,2'-azino-bis [3–ethylbenzothiazoline–6–sulfonic acid]) radicals. The quantification of phenolics and flavonoids was carried out by high-performance liquid chromatography (HPLC) using Acclaim C18 column, Dionex Ultimate 3000 liquid chromatograph, and detection was carried out in photodiode array detector. Results: The result showed that the total phenolics and flavonoids of the different extracts of the investigated mushrooms ranged from 3.77 ± 0.57 to 65.88 ± 1.81 mg gallic acid equivalents/100 g dry extract (DE) and 1.98 ± 0.002–12.64 ± 0.08 mg rutin equivalent/100 g DE, respectively. Furthermore, the mushroom extracts exhibited good free radical scavenging capacity. The HPLC experiment of both extracts showed the presence of different phenolic acids and flavonoids such as protocatechuic acid, syringic acid, quercetin, and Kaempferol in various amounts. Conclusion: The wild mushrooms have been studied to analyze their antioxidant property. These mushrooms may be used as functional additives or can be incorporated into our food regime, representing an alternative source of food to prevent damage caused by oxidation in the human body.
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Hypoglycemic and anti-lipidemic properties of Cinnamomum zeylanicum (”Sri Wijaya” accession) water-soluble nutraceutical in streptozotocin-induced diabetic and healthy wistar rats p. 188
W A. Niroshani Madushika Wariyapperuma, Bimali Jayawardena, Mayuri G Thammitiyagodage, Ramani Karunakaran, W G. Sarath Sisira Kumara
DOI:10.4103/pm.pm_334_20  
Background: Hyperglycemia is a serious health crisis worldwide, and more than 90% of the Sri Lankan patients effected with the condition have diabetes mellitus type 2. Natural therapeutic agents can manage the progression of the disease. Objectives: To investigate the hypoglycemic and antilipidemic effects of Cinnamomum zeylanicum (”Sri Wijaya” accession) water-soluble nutraceuticals in diabetic-induced and healthy Wistar rats. Materials and Methods: The diabetic and nondiabetic Wistar rats were treated with Cinnamon pressured water, Cinnamon decoction, pretreated pressured water Cinnamon extract for 1 month. The results were compared with the group treated with the positive control, Acarbose untreated normal group. Blood glucose and other biochemical parameters were estimated using commercial test kits. Results: There was a significant difference in the fasting serum glucose, food consumption, and water consumption in rats with induced diabetes. The total cholesterol level was significantly decreased in the normal groups treated with Cinnamon extracts, compared with the untreated groups. There was a significant increase in high-density lipoprotein cholesterol levels in the normal group treated with pressured water Cinnamon extract and decoction when compared with the Acarbose-treated diabetic group. Alanine aminotransferase (ALT) levels were significantly higher in the diabetic group treated with Acarbose than in all Cinnamon-treated groups. However, no significant difference was shown in normal rat groups for aspartate aminotransferase and ALT. Conclusion: Cinnamon nutraceuticals have the potential to reduce hyperglycemia in diabetic rats. Cinnamon extracts may inhibit α-amylase and α-glucosidase enzymes in rat pancreatic tissues.
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Naringin, a natural flavonone glycoside attenuates N-nitrosodiethylamine- induced hepatocellular carcinoma in sprague-dawley rats p. 196
Reetuparna Acharya, Namita Mishra, Abhishek Kumar, Pritha Bose, Ashok Pattnaik, Kunal Mukhopadhyay, Priyashree Sunita, Shakti Prasad Pattanayak
DOI:10.4103/pm.pm_94_21  
Background: Hepatocellular Carcinoma (HCC) being proclaimed as the world's fifth common cancer shows a high mortality rate due to delayed diagnosis. Regular day-to-day activities expose us to promotive factors that potentiate further to risk of liver damage, cirrhosis, and carcinogenesis. Matrix metalloproteinases (MMPs), a proteolytic enzyme, involved in cancer invasion are considered as a prognostic biomarker for HCC. Naringin (NAR), a flavanone glycoside, is recognized to have therapeutic efficacy in HCC. Objectives: The present research work focuses on evaluation of chemotherapeutic efficiency, NAR against HCC in Sprague–Dawley rats. Materials and Methods: In vitro studies were primarily carried out to estimate the cell viability of NAR in HepG2 cells followed by further justifications through in vivo studies in N-nitrosodiethylamine developed HCC. The efficacy of NAR at doses 30mg/kg/day and 60mg/kg/day was assessed through estimation of liver marker enzymes, antioxidant levels, glycoproteins, and level of MMP-2 and 9 to confirm the hypothesis. Results: While in vitro results revealed pronounced potency of NAR in inhibiting HepG2 cell viability, NAR concurrently stabilized the serum levels of liver marker enzymes, antioxidant levels, and serum glycoproteins. It also significantly reduced the levels of MMPs in treatment groups which was confirmed by zymography. Conclusion: With the support of these results, it can be concluded that NAR attenuates HCC by controlling the alterations in morphological, biochemical, and histopathological parameters that make it a suitable candidate as a potential chemotherapeutic agent against HCC.
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Anti-inflammatory and analgesic potential of Amorphophallus commutatus var. wayanadensis and its inhibitory effect on inflammatory mediators in lipopolysaccharide-stimulated macrophages p. 205
Sreena Raj, Rama Jayaraj, KM Gothandam
DOI:10.4103/pm.pm_153_21  
Background: An edible tuber named Amorphophallus commutatus var. wayanadensis (ACW) is used by the local ethnic communities of Wayanad, India, for hemorrhoids while the health benefits remain unexplored to the scientific community. Objectives: Hence, our study was performed to screen the anti-inflammatory and analgesic activities of ACW. Materials and Methods: Screening for the in vitro anti-inflammatory potential on isolated peritoneal macrophages was performed using nitrite assay, nitroblue tetrazolium assay, tumor necrosis factor (TNF)-α production, and cyclooxygenase (COX) enzyme activity. Carrageenan- and formalin-induced paw edema assays were performed to determine anti-inflammatory potential, while tail immersion assays and acetic acid-induced writhing assays were carried out to explore antinociceptive activity in animal models. Results: Bio-molecular mechanistic investigation to evaluate the in-vitro anti-inflammatory activity confirmed the suppressive effect of MEAC on TNF-α, nitric oxide and COX-2 on lipopolysaccharide stimulated peritoneal macrophages, which benchmarked ACW as a potent medicinal plant against inflammation. Further, the efficacy of methanolic extract of A. commutatus var. wayanadensis (MEAC) as an anti-inflammatory agent in murine anti-inflammatory models was demonstrated by formalin- and carrageenan-induced paw edema assays. Administration of MEAC significantly increased the tail flicking latency in mice and also showed prominent reduction in the number of writhes induced by acetic acid, which establishes the real time application of ACW as an analgesic agent. Conclusion: The vital information regarding in vitro and in vivo action of MEAC provided scientific evidence for its traditional usage for hemorrhoids. Thus, this herbal medicine of ethnic tribes can be translated toward modern medicine for health maintenance and disease prevention.
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Combinatorial effect of Macrotyloma uniflorum (Lam.) Verdc. seed extract and vorinostat (suberoylanilide hydroxamic acid) in human leukemia cells p. 213
Azra Anjum, Humaira Farooqi, MZ Abdin, Ashok Kumar Tiwari
DOI:10.4103/pm.pm_70_21  
Background: Many histone deacetylase (HDAC) inhibitors have demonstrated preclinical efficacy as monotherapy or in combination with other anticancer drugs for both hematological and solid tumor malignancies. In clinical, however, the HDAC inhibitors have proven less effective as single agents. Thus, prompting the investigation of rational combination of HDAC inhibitors with other therapeutics. Objectives: This study aims to evaluate the potential of Macrotyloma uniflorum in facilitating the potency of histone deacetylase inhibitor, Vorinostat-suberoylanilide hydroxamic acid (SAHA). Materials and Methods: Powdered seeds of M. uniflorum were sequentially extracted with water, methanol, ethyl acetate, and hexane. The quantitative analysis of the different extracts was carried out. The cytotoxicity of M. uniflorum seed extracts was investigated on the leukemia cell line (HL-60). HeLa cells were used to study the histone deacetylase inhibitory activity of the extracts. Further, the effect of the combination of M. uniflorum seed extract and SAHA was studied. Results: The aqueous extract showed the highest amount of both phenolic (311.21 ± 2 mg gallic acid/g) and flavonoid content (89.96 ± 4.62 mg Quercetin/g extract). The combination showed more specificity toward class I HDAC isoforms among the different isoforms (class I, class IIa, class IIb, and class IV). Moreover, M. uniflorum alone showed no significant cytotoxicity on the HL-60 cell line. However, in combination with SAHA, the aqueous extract exhibited significant cytotoxic activity. Further, the results show that the extract of M. uniflorum has a significant effect on cell death and autophagy in leukemia. Conclusion: Mechanistic studies revealed that aqueous extract along with SAHA facilitated HDAC inhibitory activity, cell death, and autophagy.
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Inhibitory Roles of Nigella sativa seed extracts on in vitro glycation and aggregation p. 220
Umme Rubab, Dinesh Kumar, Mohammad Abul Farah, Khalid Mashay Al-Anazi, M Ajmal Ali, Ahmad Ali
DOI:10.4103/pm.pm_604_20  
Background: Glycation is the multi-step process wherein the carbonyl group of glucose reacts with the amino group of other nitrogen-containing compounds like proteins and DNA. This interaction leads to the generation of harmful metabolites commonly known as advanced glycation end products (AGEs). These AGEs have been implicated in various diseases such as cataracts, diabetes, and Alzheimer's. Objectives: The aim of the present study was to check the effect of Nigella sativa seed extracts on glycation and aggregation and for this purpose, two proteins (bovine serum albumin [BSA] and hemoglobin) and two sugars (glucose, and ribose) were selected. Materials and Methods: The amount of glycation mediated products was measured by established methods such as browning, nitroblue tetrazolium (NBT) assay, carbonyl content, and aggregation assays in the presence or absence of extracts (1 mg/ml). The effect of extracts was also checked on glycated DNA by agarose gel electrophoresis. Results: Analysis of results indicates that extracts (1 mg/ml) significantly prevented the generation of early products up to 30.57% (BSA-Rib) and 64.90% (Hb-Glu). The AGEs formation was also suppressed in different glycation systems up to 60.76% (BSA-Glu), 30.79% (BSA-Rib) and 10.74% (Hb-Glu) in the presence of extracts. There was also a decrease in the level of glycation-mediated protein aggregation. Seed extracts were able to prevent the glycoxidative damage of DNA. Conclusion: It can be concluded that black cumin seeds have antiglycating and antiaggregation potential as well as antioxidant properties. Further characterization will help in the identification of active compounds which can be used for the prevention of secondary complications of diabetes and aggregation.
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Effect of salinity stress on growth, water content, and guggulsterone production in callus cultures of Commiphora wightii (Arnott.) Bhandari p. 225
Soumi Datta, Jatin Katna, Dhiraj Shrivastav, Sarvepalli Badari Narayan, Shahid Umar, Jatavallabhula Lakshminarayana Sastry, Ranjan Mitra, Sayeed Ahmad
DOI:10.4103/pm.pm_514_20  
Objectives: A traditional herb Commiphora wightii (Arnott.) is well known for the management of different ailments, including hypolipidemic and hypocholesterolemic disorders. Overexploitation has resulted in enlisting it under endangered species list; thus, in vitro studies are desirable to circumvent further depletion of the wild population. The objective of our study is to analyze the effect of salinity stress on growth and guggulsterone content in callus cultures. Materials and Methods: The establishment of leaf-derived calli of C. wightii and the effect of salinity stress such as calcium chloride (CaCl2), sodium chloride (NaCl), potassium chloride, and ammonium sulfate at different concentrations and time periods on callus growth and steroid biosynthesis were studied. High-performance liquid chromatography and liquid chromatography-mass spectrometry/mass spectrometry-based spectrometric systems were carried to identify the guggulsterones. Results: The 50 mg/L CaCl2-induced callus was found to have the highest content of guggulsterones after 15 months of incubation as compared to 9 and 12 months and indicated a positive correlation between callus age and guggulsterone biosynthesis. Conclusion: Salinity stress played an important role in callus growth of C. wightii, while calcium-induced calli showed in vitro synthesis of guggulsterones, and this can be further explored for the synthesis of guggulsterones in vitro on large commercial scale using bioreactor, and thus helps to conserve the endangered species in its natural habitat.
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Isolation, characterization, and chromatographic estimation of esculin: A potential fluorescent marker from Launaea pinnatifida cass p. 233
Himanshu Makwana, Devang Pandya
DOI:10.4103/pm.pm_243_21  
Background: Launaea pinnatifida (Willd.) Sch. Bip ex Kunze (Asteraceae) is an Indian Ocean intertidal sand dune herb that is useful as traditional medicine. The plant is one of the species of traditional Indian Ayurvedic herb “Gojihva.” Objectives: The present study was performed to isolate, characterize, and estimate a potential marker compound from L. pinnatifida. Materials and Methods: The leaves of the plant were subjected to successive solvent extraction with the help of solvents of varying polarity. Based on bioactivity studies performed using these extracts, the most bioactive methanolic extract was further subjected to vacuum liquid chromatography using chloroform and methanol in varying proportions. A phytoconstituent was isolated from fraction chloroform: methanol (90:10), elucidated to be Esculin based on its analytical (FTIR, liquid chromatography–mass spectrometry, 1H nuclear magnetic resonance, and ultraviolet) data. For the quantitative analysis of esculin in L. pinnatifida leaves, simple and validated high-performance liquid chromatography (HPLC) and high-performance thin-layer chromatography (HPTLC) methods were developed. Results: The purity of isolated as well as purchased standard esculin was found to be 97.66% and 98.2%, respectively, by HPLC, whereas the yield of esculin in the methanolic extract of leaves was found to be 1.40% w/w by HPTLC. Conclusion: Esculin has been reported for the first time in this plant and can be considered a potent phytochemical marker. It can also be useful for the standardization and quality control of L. pinnatifida formulations.
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Moringa Oleifera leaf extract exerts antiproliferative effects and induces mitochondria mediated apoptosis within rat glioblastoma (c6) cells p. 240
Ahmed Alafnan, Talib Hussain, Abdulwahab Alamri, Farhan Alshammari, Afrasim Moin, KL Krishna
DOI:10.4103/pm.pm_145_21  
Background: Glioblastoma multiforme is a dreaded manifestation of brain tumors resulting in substantial mortality among affected individuals globally. Moringa oleifera (Mo) is well known from earlier times for its medicinal use in conventional medication for different ailments such as cancer. Objective: The present study aims to evaluate the antiproliferative efficacy of ethanolic Moleaf extract (Mo E t-OH) in mouse-derived glioblastoma C6 cells. Materials and Methods: MoEt-OH was prepared, and C6 cells were subjected to MoEt-OH treatment at a dosage of 100, 200, and 400 μg/ml and incubated for 24 h. Results: Postincubation, C6 cells exhibited a significant (P < 0.05) decline in their viability at 100 μg/ml, which further increased proportionally with increase in MoEt-OH concentration (P < 0.01; P < 0.001). MoEt-OH significantly enhanced the lipid peroxidation as assessed by measuring the increased levels of malondialdehyde at 100 μg/ml (P < 0.05), 200 μg/ml (P < 0.01), and 400 μg/ml (P < 0.001). MoEt-OH-mediated evaluation of glutathione levels also exhibited similar trends. Moreover, reactive oxygen species estimation revealed a substantial increase in oxidative stress posttreatment with MoEt-OH within C6 cells, even in a dose-dependent manner. MoEt-OH also instigated apoptosis with glioblastoma cells through enhanced nuclear condensation and fragmentation as qualitatively evaluated through Hoechst 33342 staining. The apoptosis within C6 cells post-MoEt-OH treatment was linked with enhanced expressional levels of caspase-9 and caspase-3 proportional to the MoEt-OH concentration. Conclusion: Thus, our preliminary study elucidated that MoEt-OH treatment results in antiproliferation within C6 cells by enhancing oxidative stress and instigating apoptosis by initiating nuclear fragmentation.
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In vitro hypoglycemic effects of molokhia leaves (Corchorus olitorius L.) p. 246
Faiyaz Ahmed
DOI:10.4103/pm.pm_525_20  
Background: Corchorus olitorius L., commonly known as molokhia in Arabic, is a leafy vegetable containing large amounts of mucilaginous polysaccharides and reported to exhibit antidiabetic diabetic activity in experimental diabetes. Although antidiabetic effect of molokhia is attributed to the presence of phenolic and non-phenolic compounds, the precise mechanism of action is not explored. Materials and Methods: The present study investigated the ability of molokhia leaves powder (MLP) to modulate starch digestion and glucose diffusion in vitro. Results: MLP at 2% and 4% levels bound significantly higher amount of glucose than wheat bran (2%) and also reduced diffusion of glucose through dialysis membrane as reflected by significantly higher glucose dialysis retardation index. In starch–α-amylase system, MLP (4%) showed complete inhibition of starch digestion and glucose diffusion similar to that of acarbose till 120 min. Further, glucose diffusion was retarded to an extent of 83.7% and 63.5% by MLP (4%) at 180 and 240 min, respectively. On the other hand, MLP (2%) retarded glucose diffusion to an extent of 96%, 65%, and 51% at 120, 180, and 240 min respectively. Furthermore, molokhia leaf extract significantly enhanced uptake of glucose by rat hemidiaphragm in vitro. Conclusion: These findings conclusively demonstrate that the antidiabetic effect of molokhia leaves is mediated through delaying starch digestion and physical adsorption of liberated glucose limiting its diffusion across intestinal lumen and enhancing glucose uptake in peripheral tissues.
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Cardioprotective effect of hydroalcoholic leaf extract of Jatropha mollissima on isoproterenol-induced myocardial infarction in rats p. 251
Muhammad Omer Iqbal, Imran Ahmad Khan, Ashira Manzoor, Shafia Arshad, Asad Saleem Sial, Eshwa Dar, Abdul Rashid Shaikh
DOI:10.4103/pm.pm_16_21  
We aim to explore various effects of doses of Jatropha mollissima against isoproterenol-induced myocardial infarction in the rat. Rats were divided into 6 groups (10 rats in each). Group-1 was normal to control, and Group-2 was considered intoxicated isoproterenol (ISP) (100 mg/kg, s. c.) Group-3 was treated with standard drug carvedilol, while Group-4, 5, and 6 were treatment groups and treated with J. mollissima extract at the doses of 400, 600, and 800 mg/kg, respectively. Preliminary phytochemistry, histopathological variation in the myocardium, antioxidant potential, and cardiac biomarkers (serum glutamic oxaloacetic transaminase, triglyceride, 2,4,6-trinitrotoluene, serum glutamic pyruvic transaminase, lactate dehydrogenase, and creatine kinase muscle–brain fraction), cardiac rate, electrocardiographing, and pressurization rate index were estimated. This study indicates that the extract proved to have cardioprotective potential and significantly reduced the cardiac biomarkers in a dose-dependent fashion because of flavonoid contents and antioxidant property. The histopathological analysis shows marked improvement in J. mollissima groups handled in comparison with ISP. The present evaluation suggests that J. mollissima has exceptional cardioprotective potential contrary to toxicity caused by isoproterenol. We recommend further studies at the molecular level to frame the exact mechanism of action.
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Combination effect of Spirulina fusiformis with rutin or chlorogenic acid in lipopolysaccharide-induced septic cardiac inflammation in experimental diabetic rat model p. 257
Aman Sharma, Sumeet Gupta, Sunil Sharma, Meenakshi Dhanawat, Kavita Munjal
DOI:10.4103/pm.pm_179_21  
Background: The connection between inflammation and hyperglycemia leads to diabetes mellitus and its complications. Nutraceuticals are natural sources which can be used as supplement food for the prevention of many disorders. Our aim was studied to the protective role of two antioxidants (Rutin and Chlorogenic acid) with Spirulina fusiformis (SP) on streptozotocin-induced diabetic rats and being challenged with lipopolysaccharide to accelerate the diabetic inflammation. Materials and Methods: Rats were distributed into eleven groups. Each group has 6 rats. Lipopolysaccharide was administered to induced inflammation in diabetes mellitus rats. Orally administered Spirulina fusiformis (500 and 1000 mg/kg), rutin and chlorogenic acid (30 and 60 mg/kg) individually or in combination for 28 days. Metformin (200 mg/kg) was used as a standard drug. Biochemical parameters, body weight, and blood pressure levels were measured at different intervals. Histological studies, Scanning Electronic Microscopy, and gene expression analysis Real-time polymerase chain reaction of NRF2 and inducible nitric oxide synthase gene were analyzed at the end of the experiment. Results: Decreased in blood glucose levels and lipid profile at different intervals were noted with combination drug therapy (Spirulina fusiformis with rutin and Spirulina fusiformis with Chlorogenic acid). Improvement in antioxidant enzymes and pro-inflammatory cytokines levels were decreased on the 45th day. Maximum diastolic blood pressure levels were decreased statistically on the 45th day. Proof of evidence was also supported with these results. Conclusion: Combination therapy was showed excellent results than individual therapy at all parameters.
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Plectranthus amboinicus (Spreng.) Semi-purified Fractions with Selective β-Glucuronidase Inhibition Elucidated with gas chromatography-mass spectrometry and in silico docking p. 268
Michael Russelle S. Alvarez, Junie Billones, Chun-Hung Lin, Francisco Heralde
DOI:10.4103/pm.pm_149_21  
Background: Plectranthus amboinicus (Spreng.) is an herb commonly used in folk medicine and food by several Asian countries. The bioactivities of this medicinal plant have so far not been linked to a specific enzyme target. Bacterial beta-glucuronidases (β-GUS) expressed by human gut microbiota affect xenobiotic processing by reactivating toxic substances (e.g., anticancer drugs, nonsteroidal anti-inflammatory drugs, and food carcinogens) in the gut lumen. Objectives: An approach to alleviating the toxic effects of these compounds is by inhibiting bacterial β-GUS. Materials and Methods: We determined the Escherichia coli β-GUS inhibitory activity of P. amboinicus leaves using a bioassay-guided purification approach. The P. amboinicus chloroform extract was purified using normal-phase column chromatography to produce several fractions. The fractions were screened for E. coli β-GUS inhibitory activity using the 4-Methylumbelliferyl glucuronide (4-MUG) assay. Fractions with high activity were assayed further to determine toxicity against E. coli and selectivity compared to human β-GUS. Highly-active and highly-selective fractions were further characterized using gas chromatography-mass spectrometry (GC-MS) and in silico docking to identify specific compounds. Results: Assay-guided purification of the crude chloroform extract with β-GUS inhibitory activity (IC50 = 57.8 μg/mL) yielded four fractions with high activity: Fraction-543W (IC50 = 16.24 μg/mL), Fraction-5231 (IC50 = 3.087 μg/mL), and Fraction-52335A (IC50 = 12.93 μg/mL). The crude extract and fractions exhibited high selectivity for E. coli β-GUS against human β-GUS (P < 0.0001, α =0.05). The antimicrobial assay of fractions showed no toxic effects on E. coli. GC-MS profiling of the active fractions identified the compounds present to be similar to essential oil extracts of P. amboinicus reported previously. Ranking of these compounds by in silico identified the compounds with high binding affinity: Phthalic acid, cyclobutyl tridecyl ester (-7.5 kcal/mol) from Fraction-543W, N-Benzyl-2-allyl-2-tosyl-4-penten-1-amine (-8.0 kcal/mol) from Fraction-5231, and Dehydroabietic Acid (-7.9 kcal/mol) from Fraction-52335A. By comparing to binding modes of reported inhibitors, we show that these compounds also interact with active site residues Tyr469 and Tyr472, and with several residues in the β-GUS bacterial loop. Conclusion: Herein, we identified highly-active and highly-selective E. coli β-GUS inhibitors from P. amboinicus leaf chloroform extracts, utilizing a bioassay-guided purification coupled by metabolomics and in silico docking approach. This is the first report on the potential of P. amboinicus as selective inhibitor of E. coli β-GUSs.
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Phenolics from the heartwood of Tecoma mollis as potential inhibitors of COVID-19 virus main protease and spike proteins: An In silico study p. 278
Lamya H Al-Wahaibi, Md Tabish Rehman, Muneera S M. Al-Saleem, Omer A Basudan, Ali A El-Gamal, Mohamed F AlAjmi, Enaam Y Backheet, Azza A Khalifa, Wael Mostafa Abdel-Mageed
DOI:10.4103/pm.pm_35_21  
Background: The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is an emerging novel coronavirus responsible for the viral pneumonia outbreak (coronavirus disease-19 [COVID-19]) that has impacted millions of people, causing a tremendous global public health concern and number of fatalities. The development of novel antiviral agents is considered an urgent research subject. Objectives: The objective of the study is to discover the phenolic constituents of the methanolic extract of Tecoma mollis Humb. and Bonpl. heartwood and to investigate their potential inhibitory action against SARS-CoV-2 protease and/or entry proteins. Materials and Methods: The heartwood of T. mollis was extracted by maceration with 70% EtOH until complete exhaustion. The extract was concentrated under reduced pressure, mixed with distilled H2O and defatted with CHCl3 to produce a CHCl3 fraction, and then subjected to solvent fractionation with n-butanol to produce an n-butanol fraction. The n-butanol fraction was subjected to a silica gel column using CHCl3–MeOH gradient mixtures followed by reversed-phase high-performance liquid chromatography. The isolated compounds were identified using one- and two-dimensional nuclear magnetic resonance as well as mass spectroscopy. Molecular docking studies have been implemented to identify the binding pattern between ligands and target enzymes, i.e. main protease (Mpro) and spike protein receptor-binding domain (RBD), and compared with the currently used COVID-19 inhibitors. Molecular dynamic simulations have been performed to evaluate the dynamics and stability of protein–ligand complexes. The obtained information is then correlated with the essential structural features, and finally the structure–activity relationship is suggested. Results: Fourteen phenolic glycosides were isolated from the methanolic extract of T. mollis Humb. and Bonpl. heartwood in addition to an iridoid, ixoside. The molecular docking study exhibited that the isolated compounds have a higher binding affinity toward the active site of Mpro and the angiotensin-converting enzyme-2 binding site of spike protein RBD. The phenylpropanoids have higher inhibitory action with higher binding energy toward SARS-CoV-2 Mpro protease as compared to spike protein RBD. Among all the isolated compounds, isoverbascoside (10) exhibited the most potent dual interaction with SARS-CoV-2 Mpro protease and spike protein RBD with high binding energy of − 8.8 and − 7.2 kcal/mol, respectively. This showed better potency than the currently used Mpro and spike–protein inhibitors. Conclusion: Our study is the first report on the potential inhibitory action of phenylpropanoids for SARS-CoV-2 protease and spike protein. It also correlates between the reported antiviral activities of some isolated compounds with their potential inhibitory action for COVID-19 viral proteins. Our results on T. mollis extract constituents could help in the discovery of a promising repurposable drug candidate that could contribute to the development of an effective therapy for COVID-19.
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Anti-cancer effects of Nepeta Deflersiana Extract (NDE) in estrogen positive and negative forms of breast cancer p. 287
Majed Saad Al Fayi
DOI:10.4103/pm.pm_464_20  
Aim: This study focused on efficacy of the ethanol extract gained from the aerial parts of Nepeta deflersiana (ND) on breast cancer (BC) cells. Materials and Methods: Shade-dried aerial parts of ND were extracted with ethanol using the Soxhlet extraction method (NDE). MTT assay was performed to assess the cytotoxicity of NDE in normal breast cells (MCF-10A), as well as estrogen receptor (ER) positive (MCF-7) and ER-negative (MDA-MB231) BC cells. Nuclear staining was performed using Hoechst/propidium iodide. Annexin V assays for apoptosis detection and cell cycle analysis were performed by flow cytometry. The effect of NDE on metastasis was analyzed by the transmigration assay. Western blot was used to detect the key signaling proteins. Results: NDE inhibited BC cell proliferation with GI50 values of 154.90 and 202.7 μg/ml for MCF-7 and MDA-MB231 cells, respectively. NDE was non-toxic to normal MCF 10A cells at concentrations up to 1000 μg/ml. NDE induced nuclear condensation/fragmentation in both types of BC cells near the GI50 doses values. An apparent increase in early and late apoptotic populations was confirmed by the flow cytometry. The extract also increased the sub-G0/G1 population in both types of BC cells. Dose-dependent increases in Caspase 3 and Bax levels were also recognized, as were NCE decreases in anti-apoptotic Bcl-2 levels in the same cell lines. Conclusion: NDE possessed excellent anticancer properties against ER-positive and negative BC cells, encouraging further research on the use of this natural product against BC.
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