Quality control methods for fruit extracts of Kigelia africana using high performance thin layer chromatography.

Kigelia africana is a tree native to Africa but also found in eastern and southern parts of India with reported anti-bacterial, anti-inflammatory, and immunomodulatory activities. Verbascoside, caffeic acid and ferulic acid are important markers for the quality control of the plant. Two different HPTLC methods were developed and validated; method - 1 for estimation of verbascoside and caffeic acid while method - 2 for estimation of caffeic acid and ferulic acid. Developed methods were applied to the methanolic fruit extract to determine the quantities of markers. Both methods were found to be linear, specific, precise, accurate, sensitive and robust. Results indicated that both methods can be used for quantitative determination of verbascoside, caffeic acid and ferulic acid in fruit extract. The developed methods may be utilised as a part of the quality control and standardisation for the raw material and extracts of Kigelia africana and can also aid to chromatographic fingerprinting of the plant.

Evaluation of efficacy of Bombax ceiba extract and its major constituent, mangiferin in streptozotocin (STZ)-induced diabetic rats.

OBJECTIVES: Based on the ethno-medicinal use of Bombax ceiba leaf, in the treatment of diabetes, the present study is aimed at evaluation of antidiabetic potential of leaf extract and its major constituent mangiferin. METHODS: Efficacy of hydroalcoholic extract of Bombax ceiba leaf (BCL, 200 and 400 mg/kg body wt.) and mangiferin (MF, 20 mg/kg body wt) was studied in streptozotocin (STZ)-induced diabetic rats and associated complications visually, retinopathy, cardiopathy and nephropathy. After 20 days, serum glucose, lipid profiles, glycol-hemoglobin % (HbA1c%), liver enzymes activity and glycogen content, and histopathology of the pancreas were corroborated. The study was coxswained for development of validated RP-HPLC method for the estimation of MF in BCL. RESULTS: The results demonstrated significant reductions in the levels of glucose (p<0.001), glycated hemoglobin (HbA1c%, p<0.001), cholesterol, triglycerides and low-density lipoproteins, and concurrent elevation of high density lipoproteins level in the groups administered BCL and MF relative to the controls. It significantly reversed most of the altered metabolic and oxidative stress parameters and histopathological changes. Mangiferin content in BCL was found to be 0.04%w/w. CONCLUSIONS: The anti-diabetic effects of BCL may be attributed to its ability to enhance insulin release, antioxidant and hypolipidemic potential.

In silico Screening of Natural Compounds as Potential Inhibitors of SARS-CoV-2 Main Protease and Spike RBD: Targets for COVID-19.

Historically, plants have been sought after as bio-factories for the production of diverse chemical compounds that offer a multitude of possibilities to cure diseases. To combat the current pandemic coronavirus disease 2019 (COVID-19), plant-based natural compounds are explored for their potential to inhibit the SARS-CoV-2 (severe acute respiratory syndrome coronavirus 2), the cause of COVID-19. The present study is aimed at the investigation of antiviral action of several groups of phytoconstituents against SARS-CoV-2 using a molecular docking approach to inhibit Main Protease (Mpro) (PDB code: 6LU7) and spike (S) glycoprotein receptor binding domain (RBD) to ACE2 (PDB code: 6M0J) of SARS-CoV-2. For binding affinity evaluation, the docking scores were calculated using the Extra Precision (XP) protocol of the Glide docking module of Maestro. CovDock was also used to investigate covalent docking. The OPLS3e force field was used in simulations. The docking score was calculated by preferring the conformation of the ligand that has the lowest binding free energy (best pose). The results are indicative of better potential of solanine, acetoside, and rutin, as Mpro and spike glycoprotein RBD dual inhibitors. Acetoside and curcumin were found to inhibit Mpro covalently. Curcumin also possessed all the physicochemical and pharmacokinetic parameters in the range. Thus, phytochemicals like solanine, acetoside, rutin, and curcumin hold potential to be developed as treatment options against COVID-19.

Anti-inflammatory and immunomodulatory activities of Inula cappa roots (Compositae).

Background The medicinal properties of Inula cappa, a perennial shrub, are ascribed to its roots. In this article, we study the anti-inflammatory and immunomodulatory activities of I. cappa root extracts based on its utilization in traditional medicine-treatment of rheumatoid arthritis, menoxenia, fever, jaundice and many others. Methods Anti-inflammatory and immunomodulatory activities of I. cappa extracts were studied. The anti-inflammatory activity was determined by carrageenan-induced rat paw edema and cotton pellet-induced granulation method while the immunomodulatory activity was estimated by phagocytic assay method, hemagglutinating antibody (HA) titer assay, delayed-type hypersensitivity assay method, plaque forming cell assay and determination of immunoprophylatic activity. Results The methanolic extract showed maximum reduction in the rat paw edema and showed significant inhibition of the cotton pellet-induced granulomas in rats. The methanolic extract also showed potential immunomodulatory activity in all the assays performed. Two sesquiterpenes, isoalantolactone and germacranolide were also isolated from the methanolic extract. Conclusions The present study supports the evidence that the roots of I. cappa can be used as a potent anti-inflammatory and immunomodulatory agent.

Development and Validation of a Rapid LC-MS/MS Method for Simultaneous Determination of Kaempferol and Quercetin in Thespesia populnea Extract.

In this study, a simple and rapid LC with tandem MS method was developed and validated for the simultaneous determination of kaempferol and quercetin in Thespesia populnea extract. The compounds were eluted using a Gemini C18 column (50 × 2.0 mm, 3 µm), with the mobile phase consisting of acetonitrile-0.3% formic acid in water at the flow rate of 0.400 mL/min. The assay exhibited a linear dynamic range of 25-2500 ng/mL for both kaempferol and quercetin. The values for intra- and interday precision and accuracy were well within the generally accepted criteria for analytical methods (<15%). Selectivity, linearity, LOD, LOQ, accuracy, and precision were evaluated for both analytes. The proposed method is accurate and sensitive and can be used for the routine quantification of kaempferol and quercetin in the herbal extract and in polyherbal formulations.

Validated high performance thin layer chromatography method for simultaneous determination of quercetin and gallic acid in Leea indica

ABSTRACT A sensitive and reliable high performance thin layer chromatography method has been developed for the simultaneous estimation of quercetin and gallic acid in Leea indica, Vitaceae. Ethyl acetate extract prepared from hydrolysed aqueous alcoholic extract (70%) was applied on silica gel G 60 F254 plate. The plate was developed using toluene-ethyl acetate-formic acid, 5:4:1 (v/v/v) as a mobile phase and detection and quantification were performed by densitometric scanning at 254 nm. The system was found to give well resolved bands for quercetin (Rf 0.63) and gallic acid (Rf 0.45) from other constituents present in the extract of L. indica. The correlation coefficient was found to be 0.991 and 0.999 with relative standard deviation, 0.97–1.23% and 0.1–1.13% for quercetin and gallic acid respectively in the developed method. The accuracy of the method was confirmed by conducting recovery studies at different levels using the standard addition method. The average recovery of quercetin and gallic acid was found close to 99% suggesting the accurateness of the method. The proposed validated high performance thin layer chromatographic method offers a new, sensitive, specific and precise gauge for quantification of quercetin and gallic acid in L. indica.

Evaluation of antiulcer potential of Mimusops hexandra in experimental gastro duodenal ulcers.

The study was aimed to investigate antiulcer effects of acetone extract and its different fractions Mimusops hexandra against experimental gastro-duodenal ulcers. 80% acetone extract of stem bark of Mimusops hexandra (Extract A, p.o.) and its different fractions namely diethyl ether (Extract A1, p.o.), ethyl acetate (Extract A2, p.o.) and aqueous (Extract A3, p.o.) were tested for the presence of preliminary phytoconstituents and were screened for their antiulcer potential in experimental animals using ethanol-HCl and aspirin-induced gastric damage at the dose of 500 mg kg-1p.o. Extract A2 being the most active fraction amongst all the fractions tested was also studied at different doses to find its ED50. Further, to establish the mechanism of action, Extract A2 was also tested for its effects in acetic acid-induced gastric ulcer models and cysteamine-induced duodenal ulcer. The effect was compared with cimetidine. Flavonoids (quercetin), procyanidins, saponins and triterpenoids were found to be present in bark. Oral administration of Extract A2 inhibited formation of gastric lesions induced by aspirin in a dose dependent manner. Elevated level of lipid peroxidation due to ethanol-HCl and aspirin induced gastric damage was significantly (p<0.05) reduced by the treatment with Extract A2. Further, Extract A2 at the dose of 100 mg kg-1 (p.o.) reduced extent of acetic acid induced gastric ulcer in experimental animals. Moreover, protection afforded by Extract A2 against cysteamine-induced duodenal lesions was evident from dose dependent decrease in ulcer index (p<0.05), score for intensity (p<0.05) and total lesion area (p<0.05), when compared with the control group. The antiulcer activity shown by Extract A2 in experimental gastro-duodenal ulcer could be attributed to decrease in gastric acid secretory activity along with strengthening of mucosal defensive mechanisms.

Hepatoprotective activity of punarnavashtak kwath, an Ayurvedic formulation, against CCl4-induced hepatotoxicity in rats and on the HepG2 cell line.

OBJECTIVE: Punarnavashtak kwath (PNK) is a classical Ayurvedic formulation, mentioned in Ayurvedic literature Bhaishajya Ratnavali, for hepatic disorders and asthma. This study investigated the hepatoprotective activity of PNK to validate the traditional use of this formulation. MATERIALS AND METHODS: PNK was prepared in the laboratory according to the method given in Ayurvedic literature. Phytochemical screening was performed to determine the presence of phytoconstituents. Hepatoprotective activity was evaluated against CCl(4)-induced hepatotoxicity in rats and by its effect on the HepG2 cell line. RESULTS: Preliminary phytochemical screening revealed the presence of alkaloids, tannins, flavonoids, saponins, and a bitter principle in PNK. Administration of PNK produced significant hepatoprotective effect as demonstrated by decreased levels of serum liver marker enzymes such as aspartate transaminase, serum alanine transaminase, serum alkaline phosphatase, and serum bilirubin and an increase in protein level. Thiopentone-induced sleeping time was also decreased in the PNK-treated animals compared with the CCl(4)-treated group. It also showed antioxidant activity by increase in activity of glutathione, superoxide dismutase, and catalase and by a decrease in thiobarbituric acid reactive substance level compared with the CCl(4)-treated group. Results of a histopathological study also support the hepatoprotective activity of PNK. Investigation carried out on the HepG2 cell line depicted significant increase in viability of cells exposed to PNK as compared with CCl(4)-treated cells. DISCUSSION AND CONCLUSION: It can be concluded that PNK protects hepatocytes from CCl(4)-induced liver damages due to its antioxidant effect on hepatocytes. An in vitro study on HepG2 cell lines also supports its protective effect.

Evaluation of anti-inflammatory and antioxidant activity of Premna integrifolia root.

Roots of Premna integrifolia Linn. Mant. (Verbanacea) are important rasayana (Adaptogenic) drugs and are considered to be useful in the treatment of variety of ailments. The present study was aimed at evaluating the for its anti-inflammatory and antioxidant activities. Pretreatment with a single dose of methanolic extract of P. integrifolia (PIM) (300 mg/kg b.w.) produced significant inhibition on carrageenan-induced rat hind paw edema, histamine induced wheal formation, and acetic acid-induced mouse vascular permeation. In a 7-day study, daily administration of PIM suppressed formalin induced paw edema and cotton pellet-induced rat granuloma formation. The extract also showed significant inhibition of cyclo-oxygenase (COX-I) activity on rat uterus and plasma membrane stabilization. Apart from this, the antioxidant activity (in vitro) of the extracts was evaluated using the anti radical, superoxide scavenging, erythrocyte membrane stability, anti lipid peroxidation, hydroxyl radical scavenging, nitric oxide scavenging and reducing power (ferric thiocynate method and ß-carotene bleaching test) assays. PIM showed significant anti-oxidant activity. In addition, PIM (300mg/kg b.w.) produced no observable sub acute toxicity in mice with in 15 days. The results scientifically demonstrated the antiinflammatory activity of P. integrifolia roots in various experimental models probably through their antihistaminic, antikinin, COX-inhibitory and antioxidant action. This therefore justifies the folkloric use of the plant.

Involvement of the PI3K/AKT pathway in the hypoglycemic effects of saponins from Helicteres isora.

AIM OF THE STUDY: Saponins from Helicteres isora have previously been shown to exert antidiabetic effects. The present study explored the underlying mechanisms in C2C12 skeletal muscle cells. MATERIALS AND METHODS: C2C12 cells were incubated with saponins and sapogenin followed by Western blotting and immunofluorescence analysis. RESULTS: Western blotting revealed that incubation with saponins (100 microg/ml) and sapogenin (100 microg/ml) induced the phosphorylation of the phosphatidylinositol-3-kinase (PI3K) as well as of the downstream targets protein kinase B/Akt (at Ser473) and glycogen synthase kinase GSK-3 alpha/beta (at Ser21/9) in a time-dependent manner. In contrast, no phosphorylation of the AMP-sensitive kinase AMPK (at Thr172) was observed. Within 48 h saponins/sapogenin treatment further increased the protein abundance of the insulin-sensitive glucose transporter Glut4. Confocal microscopy confirmed that saponins/sapogenin treatment stimulated Akt phosphorylation and revealed that the treatment was followed by translocation of Glut4 into the cell membrane of C2C12 muscle cells. CONCLUSIONS: Saponins and sapogenin activate the PI3K/Akt pathway thus leading to phosphorylation and inactivation of GSK-3 alpha/beta with subsequent stimulation of glycogen synthesis as well as increase of Glut4-dependent glucose transport across the cell membrane.

Effect of Manilkara hexandra (Roxb.) Dubard against experimentally-induced gastric ulcers.

Effects of the flavonoid rich fraction of the stem bark of Manilkara hexandra (Roxb.) Dubard, have been studied on ethanol, ethanol-indomethacin and pylorus ligated gastric ulcers in experimental animals. Oral administration of the ethyl acetate extract (extract A3) inhibited the formation of gastric lesions induced by ethanol in a dose dependent manner. The protective effect of extract A3 against ethanol induced gastric lesions was not abolished by pretreatment with indomethacin (10 mg kg(-1)). Further, extract A3 inhibited increase in vascular permeability due to ethanol administration. Extent of lipid peroxidation was significantly reduced in animals treated with extract. Extract A3 also inhibited the formation of gastric ulcers induced by pylorus ligation, when administered both orally and intraperitoneally. Moreover, pretreatment with extract A3 increased mucus production and glycoprotein content, which was evident from the rise in mucin activity and TC: PR ratio.

Study of Mimusops elengi bark in experimental gastric ulcers.

The present study was designed to investigate the effect of Mimusops elengi (Sapotaceae) against experimental gastric ulcers. The 50% alcoholic extract of Mimusops elengi (Ext E) and its different fractions namely ethyl acetate (Ext E1), n-butanol (Ext E2), methanol (Ext E3) and aqueous (Ext E4) were studied (p.o.) against ethanol-induced gastric damage. Ext E1 was also studied in ethanol-induced, pylorus-ligated and water-immersion plus stress-induced gastric ulcer models. Ranitidine HCl (80 mg kg(-1)) was used as a reference standard. In ethanol-induced gastric ulcer model, pantoprazole (20 mg kg(-1)) was also used as a reference standard. Ext E1 tested in mice up to the dose of 5000 mg kg(-1) (p.o.) did not produce any sign of toxicity. Ext E at the doses of 50, 100, 300 and 500 mg kg(-1) and its different fractions (100 mg kg(-1)) showed reduction in gastric ulceration (P < 0.05). Ext E1 at the doses of 10, 50 and 100 mg kg(-1) showed dose-dependent inhibition of gastric lesions against ethanol-induced gastric damage. In 19 h pylorus-ligated animals, Ext E1 at 50 and 100 mg kg(-1) doses showed significant reduction in ulcer index (P < 0.05). Significant reduction was also observed in total acidity, volume of gastric acid secretion, total acid output and pepsin activity (P < 0.05) when compared with the control group. Besides, Ext E1 showed increase in the mucosal glycoproteins that was evident from significant rise in total carbohydrates to protein ratio (TC:PR ratio) (P < 0.05), which is an indication of mucin activity. Ext E1 also showed protection against water-immersion plus stress-induced gastric lesions that was evident from dose-dependent decrease in ulcer index (P < 0.05), score for intensity (P < 0.05) and total lesion area (P < 0.05) when compared with the control group. It can be concluded from our study that Ext E1 possesses anti-ulcer activity against experimental gastric ulcers. The mechanism of anti-ulcer activity can be attributed to decrease in gastric acid secretory activity along with strengthening of mucosal defensive mechanisms.