Rutin from Begonia roxburghii modulates iNOS and Sep A activity in treatment of Shigella flexneri induced diarrhoea in rats: An in vitro, in vivo and computational analysis.

In developing countries, diarrhoea is a major issue of concern, where consistent use of antibiotics has resulted in several side effects along with development of resistance among pathogens against these antibiotics. Since natural products are becoming the treatment of choice, therefore present investigation involves mechanistic evaluation of antidiarrhoeal potential of Begonia roxburghii and its marker rutin against Shigella flexneri (SF) induced diarrhoea in rats following in vitro, in vivo and in silico protocols. The roots of the plant are used as vegetable in the North East India and are also used traditionally in treating diarrhoea. Phytochemically standardized ethanolic extract of B. roxburghii (EBR) roots and its marker rutin were first subjected to in vitro antibacterial evaluation against SF. Diarrhoea was induced in rats using suspension of SF and various diarrhoeagenic parameters were examined after first, third and fifth day of treatment at 100, 200 and 300 mg/kg, p.o. with EBR and 50 mg/kg, p.o. with rutin respectively. Additionally, density of SF in stools, stool water content, haematological and biochemical parameters, cytokine profiling, ion concentration, histopathology and Na+/K+-ATPase activity were also performed. Molecular docking and dynamics simulation studies of ligand rutin was studied against secreted extracellular protein A (Sep A, PDB: 5J44) from SF and Inducible nitric oxide synthase (iNOS, PDB: 1DD7) followed by network pharmacology. EBR and rutin demonstrated a potent antibacterial activity against SF and also showed significant recovery from diarrhoea (EBR: 81.29 ± 0.91% and rutin: 75.27 ± 0.89%) in rats after five days of treatment. EBR and rutin also showed significant decline in SF density in stools, decreased cytokine expression, potential antioxidant activity, cellular proliferative nature and recovered ion loss due to enhanced Na+/K+-ATPase activity, which was also supported by histopathology. Rutin showed a very high docking score of -11.61 and -9.98 kcal/mol against iNOS and Sep A respectively and their stable complex was also confirmed through dynamics, while network pharmacology suggested that, rutin is quite capable of modulating the pathways of iNOS and Sep A. Thus, we may presume that rutin played a key role in the observed antidiarrhoeal activity of B. roxburghii against SF induced diarrhoea.

Quercetin a major biomarker of Psidium guajava L. inhibits SepA protease activity of Shigella flexneri in treatment of infectious diarrhoea.

The leaves of the plant Psidium guajava L. (Myrtaceae) has been traditionally used in treatment of various gastrointestinal disorders including diarrhoea and have also been reported for its potent antidiarrhoeal activity on various chemical induced diarrhoea models. The objective of our present study was to evaluate the potency of the leaf extract of the plant Psidium guajava (PGE) along with its major biomarker quercetin against Shigella flexneri-induced sub chronic model of infectious diarrhoea. PGE at 100, 200 and 400 mg/kg, p.o. and quercetin at 50 mg/kg, p.o. were administered to Shigella flexneri-induced diarrhoeal rats for five days and various behavioural parameters were evaluated on 1st, 3rd and 5th day of treatment. This was followed by assessment of stool water content, density of Shigella flexneri in stools and blood parameters examination. After treatment, colon and small intestine of rats was dissected and subjected to biochemical estimations, cytokine profiling, antioxidant evaluations, ion concentration determination, Na+/K+-ATPase activity and histopathology. Molecular docking studies on crystal structure of Secreted Extracellular Protein A (SepA) from Shigella flexneri with biomarker quercetin was also performed. PGE at 200 mg/kg followed by quercetin depicted maximum antidiarrhoeal potential, which was confirmed through diarrhoea score and % protection, while PGE at 400 mg/kg showed similar effect to PGE 200 mg/kg thus, the later may have ceiling effect. PGE and quercetin also significantly reduced the density of Shigella flexneri in stools, water content of stools and restored the alterations observed in blood parameters, antioxidant status and pro-inflammatory cytokines (IL-6 and TNF-α) expression. These parameters contributed in normalization of electrolyte balance, reactivation of Na+/K+-ATPase activity and repairing of epithelial tissue damage, confirmed through histopathology. Docking simulation studies revealed the role of quercetin in inactivating the protease activity of SepA, a protein secreted by Shigella, which disrupts epithelial barrier integrity during infection and also manages its signal production. Thus, the overall results confirmed the role of quercetin as a major biomarker for the observed antidiarrhoeal potential of P. guajava against Shigella flexneri induced infectious diarrhoea.

Corrigendum to "Wound healing potential of Acacia catechu in streptozotocin-induced diabetic mice using in vivo and in silico approach" [J Tradit Compl Med 13 (5) (2023) 489-499].

[This corrects the article DOI: 10.1016/j.jtcme.2023.05.001.].

Wound healing potential of Acacia catechu in streptozotocin-induced diabetic mice using in vivo and in silico approach.

Background and aim: Acacia catechu Wild. (Fabaceae) barks are traditionally used in the treatment of diabetes and wounds. Therefore, the objective of the present study was to evaluate the wound healing potential of the alcoholic extract of A. catechu (EAC) in streptozotocin-induced diabetic mice. Experimental procedures: EAC was first subjected to phytochemical estimations and standardization using (-) epicatechin as marker with the help of HPLC. Diabetes was induced in mice using streptozotocin and the wound healing potential of EAC was evaluated using excision and incision wound models on topical and oral treatment. Various biochemical parameters, in vivo antioxidants, cytokine profiling, VEGF, and histopathological examination were also performed. Further, molecular docking studies were performed using ligand (-) epicatechin on human inducible nitric oxide synthase. Results and conclusion: Phytochemically, EAC showed the presence of tannins, flavonoids, phenolic compounds, and saponins, while the content of (-) epicatechin was reported to be 7.81% w/w. The maximum healing of wounds (91.84 ± 1.10%) was observed in mice treated with a combination of both topical (10% gel) and oral (extract at 200 mg/kg) followed by topically and orally treated groups respectively after 14 days of treatment. These groups also showed significant restoration of altered biochemical parameters, antioxidant enzymes and cytokines. The molecular docking studies confirmed the role of (-) epicatechin in stabilizing the human inducible nitric oxide synthase with inhibitor showing binding energy of -8.31 kcal/mol. The present study confirmed the role of (-) epicatechin as a major marker in diabetic wound healing potential of A. catechu.

Evaluation of pancreatic regeneration activity of Tephrosia purpurea leaves in rats with streptozotocin-induced diabetes.

BACKGROUND AND AIM: Flavonoid rich plant Tephrosia purpurea (T. purpurea), commonly known as Sarpunkha has been used in traditional systems of medicine to treat diabetes mellitus. However, its effectiveness in promoting regeneration of pancreas in diabetes has not been investigated. Therefore, the present study was undertaken to evaluate pancreatic ß-cells regeneration, antioxidant and antihyperlipidemic potentials of T. purpurea leaves extract, its fractions and main constituent Rutin in diabetic rats. EXPERIMENTAL PROCEDURE: The leaves extract and its fractions were first screened for acute and sub-chronic antidiabetic activity in a dose range of 250-500 mg/kg orally. Further, fractions with potent antidiabetic activity were screened for pancreatic ß-cells regeneration activity using histopathological studies and morphometric analysis, which was followed by estimation of biochemical parameters. RESULTS AND CONCLUSION: The most significant antidiabetic, pancreatic regeneration and antihyperlipidemic activity was exhibited by n-butanol soluble fraction of ethanol extract at the dose level of 500 mg/kg. Histopathology revealed that treatment with this fraction improved the ß-cell granulation of islets and prevented the ß-cells damage which was further confirmed by morphometric analysis. Thus, the present study validated the traditional use of T. purpurea plant in the treatment of diabetes, which might be attributed to pancreatic ß-cells regeneration potential of its active constituent Rutin. TAXONOMY CLASSIFICATION BY EVISE: Traditional Medicine; Metabolic Disorder; Experimental Design; Cell Regeneration and Histopathology.

The antidiarrhoeal evaluation of Psidium guajava L. against enteropathogenic Escherichia coli induced infectious diarrhoea.

ETHNOPHARMACOLOGICAL RELEVANCE: The plant Psidium guajava L. (Myrtaceae), commonly used as an edible fruit is traditionally used worldwide in treatment of various gastrointestinal problems including diarrhoea. The leaves of the plant have also been evaluated for antidiarrhoeal activity in various chemical induced diarrhoea models. OBJECTIVE: The main objective of the present study was to evaluate the potency of P. guajava leaves and its major biomarker quercetin against enteropathogenic Escherichia coli (EPEC) induced infectious diarrhoea using preclinical and computational model. MATERIAL AND METHODS: P. guajava alcoholic leaf extract (PGE) was initially standardized using HPLC taking quercetin as a biomarker and was then subjected to antidiarrhoeal evaluation on rats in an EPEC induced diarrhoea rat model. The study included assessment of various behavioral parameters, initially for 6 h and then for up to 24 h of induction which was followed by estimation of stool water content, density of EPEC in stools and blood parameters evaluation. The colonic and small intestinal tissues of the treated animals were subjected to various biochemical estimations, in vivo antioxidant evaluation, estimation of ion concentration, Na+/K+-ATPase activity, assessment of pro-inflammatory cytokines and histopathological studies. Further, the major biomarker off PGE, quercetin was subjected to molecular docking studies with Na+/K+-ATPase and EPEC. RESULTS: The results demonstrated a significant antidiarrhoeal activity of quercetin (50 mg/kg), PGE at 200 and 400 mg/kg, p.o., where quercetin and PFGE at 200 mg/kg, p.o. were found to be more prominent, as confirmed through higher % protection, water content of stools and density of EPEC in stools. PGE and its biomarker quercetin also significantly recovered the WBC, Hb, platelets loss and also revealed a significant restoration of altered antioxidants level, pro-inflammatory cytokines (IL-1ß and TNF-α) expression and had positive influence on Na+/K+-ATPase activity. The docking studies of quercetin with Na+/K+-ATPase showed favourable interactions and residues Glu 327, Ser 775, Asn 776, Glu 779 and Asp 804 of Na+/K+-ATPase were adequately similar to quercetin for donating ligands for binding, while quercetin was also found to terminate the linkage between mammalian cells and EPEC thus, preventing further infection from EPEC. CONCLUSION: Inhibition in intestinal secretion, reduced nitric oxide production and inflammatory expression along with reactivation of Na+/K-ATPase activity could be attributed to the observed antidiarrhoeal potential of PGE against infectious diarrhoea, where quercetin was confirmed to be the main contributing factor.

Toxicological and pharmacological profiling of organically and non-organically cultivated Cymbopogon citratus.

BACKGROUND: With the rapid depletion of forests, impairing the availability of raw drugs, Ayurveda, has reached a very critical phase. Consequently, cultivation of medicinal plants is essential to ensure their availability to the industry. In view of the above concept, organic farming of medicinal plants needs scientific validity. OBJECTIVES: The present study includes the organic and non-organic cultivation of Cymbopogon citratus, followed by toxicological and pharmacological profiling of extracts. MATERIALS AND METHODS: C. citratus was simultaneously cultivated organically (OCC) and non-organically (NCC). Toxicity profile of aqueous extracts was recorded on prokaryotes using bioluminescent bacteria, Vibrio harveyi and evaluated its type 2 anti-diabetic activity. RESULTS: OCC have shown the higher mean values of height, number of germplasms and root lengths compared to NCC. The higher level of toxicity was shown by NCC with decrease in bioluminescence with increasing concentration of extract. In acute type 2 anti-diabetic study, OCC showed prominent decrease in blood glucose at postprandial condition (6th h) (48.86% OCC-200). The order of sub-chronic anti-diabetic activity was observed as positive control > OCC-200 > NCC-200, while OCC at 200 mg/kg corrected the altered lipid profile and antioxidant status with significant increase in body weights of animals. Histopathological examination of pancreas showed the enlargement of pancreatic islets and formation of neo islets with degenerative changes in OCC treated animals. CONCLUSION: The study confirms that organically grown C. citratus is better in terms of nourishment, biological activity and safety measures.

Anti-psoriatic potential of Solanum xanthocarpum stem in Imiquimod-induced psoriatic mice model.

ETHNOPHARMACOLOGICAL RELEVANCE: The plant Solanum xanthocarpum Schrad. & Wendl. (Solanaceae) is one of the members of the dashamula (ten roots) in Ayurvedic system of medicine. The stem and fruits are used as an antipyretic, antiasthmatic and is prescribed in skin infections and for relief in burning sensation in the feet accompanied by vesicular eruptions. OBJECTIVE: To scientifically validate the anti-psoriatic potential of Solanum xanthocarpum stem in Imiquimod-induced psoriatic mice model. MATERIALS AND METHODS: Ethanolic stems extract of Solanum xanthocarpum (ESX) was first subjected to phytochemical screening and quantification of identified phytoconstituents, which was further standardized with the help of HPTLC using chlorogenic acid as a marker. The extract was then subjected to acute oral toxicity and skin irritability study for determining the safety profile of the extract. Imiquimod-induced psoriatic mouse model was then performed to check the efficacy of extract against psoriasis, where treatment was carried out for 15 days both topically (Gel at 2.5%, 5% and 10%) as well as orally (at 100, 200 and 400mg/kg p.o.) and their Psoriasis Area Severity Index (PASI) was calculated. The study also included determination of levels of TNF-α, IL-1ß, IL-6 and IL-17 in the animal tissues, which further included biochemical evaluations such as total collagen, hexosamine, hyaluronic acid DNA, protein antioxidant profiles such as lipid peroxidation, nitric oxide, superoxide dismutase and catalase along with histopathological studies of the tissues. RESULT: ESX showed the presence of mainly phenols, tannins, flavonoids, alkaloids and carbohydrates, while chlorogenic acid was reported to be 3.49% w/w. The Imiquimod-induced psoriatic mouse model, depicted a potent anti-psoriatic activity of the extract both topical (10%) and oral (200 and 400mg/kg p.o., as evident through PASI grading The effect was found to be more prominent in case of topically treated as compared to orally treated mice. The results also showed a significant inhibition in the expression of TNF-α, IL-1ß, IL-6 and IL-17 in treated animal tissues and also showed significant restoration of the altered biochemical parameters along with reduced hyperkeratinisation as observed through histopathology. CONCLUSION: The study scientifically justified the anti-psoriatic activity of the ESX, which may be attributed to inhibition in the expression of cytokines such as TNF-α, IL-1ß, IL-6 and IL-17. Further, the observed antioxidant, antimicrobial and cellular proliferative activities may act as a contributing factor in treatment of psoriasis, which may be attributed to the presence of chlorogenic acid along with other phytochemicals in combination.

High performance thin layer chromatography fingerprinting, phytochemical and physico-chemical studies of anti-diabetic herbal extracts.

INTRODUCTION: Herbal medicines have gained increasing popularity in the last few decades, and this global resurgence of herbal medicines increases their commercial value. However, this increasing demand has resulted in a decline in their quality, primarily due to a lack of adequate regulations pertaining to herbal medicines. AIM: To develop an optimized methodology for the standardization of herbal raw materials. MATERIALS AND METHODS: The present study has been designed to examine each of the five herbal anti-diabetic drugs, Gymnema sylvester R. Br., Pterocarpus marsupium Roxburgh., Enicostema littorale Blume., Syzygium cumini (L.) Skeels. and Emblica officinalis Gaertn. The in-house extracts and marketed extracts were evaluated using physicochemical parameters, preliminary phytochemical screening, quantification of polyphenols (Folin-Ciocalteu colorimetric method) and high performance thin layer chromatography (HPTLC) fingerprint profiling with reference to marker compounds in plant extracts. RESULTS: All the plants mainly contain polyphenolic compounds and are quantified in the range of 3.6-21.72% w/w. E. officinalis contain the highest and E. littorale contain the lowest content of polyphenol among plant extracts analyzed. HPTLC fingerprinting showed that the in-house extracts were of better quality than marketed extracts. CONCLUSION: The results obtained from the study could be utilized for setting limits for the reference phytoconstituents (biomarker) for the quality control and quality assurance of these anti-diabetic drugs.

Antioxidant potential and its relationship with polyphenol content and degree of polymerization in Opuntia elatior Mill. fruits.

BACKGROUND: Opuntia elatior Mill. (Nagaphani) fruits are traditionally recommended as an expectorant, remedy for whooping cough, asthma, gonorrhea, ulcers, tumors, in the treatment of diarrhea and syphilis. Many of these diseases are allied with oxidative stress caused by free radicals. Thus, current research is directed towards finding naturally-occurring antioxidants of plant origin. AIM: To evaluate antioxidant potential of hydro-alcoholic extract of the O. elatior fruits (HAOE) and its fractions. MATERIALS AND METHODS: Using 1,1-diphenyl-2-picrylhydrazyl (DPPH) and nitric oxide radical scavenging assay, total polyphenolic, flavonoid (FA), flavanone (FO) contents and degree of polymerization in relation with its antioxidant activity were examined. RESULTS: The experimental data indicated that the HAOE, ethyl acetate (EAOE) and butanol (BFOE) soluble fractions have shown significant antioxidant activity. The highest polyphenolic, FA, FO contents and degree of polymerization were found in EAOE. The scavenging potential was in the order of Ascorbic Acid > EAOE > BFOE > HAOE > BIOE, where ascorbic acid was used as a positive control. The increased antioxidant potential of EAOE and BFOE fractions over HAOE extract may be attributed to the purification achieved by fractionation of the extract which in turn resulted in an increase in the degree of polymerization and segregation of secondary metabolites. CONCLUSION: The fruit of O. elatior can be used as the best alternative for synthetic antioxidants.

Involvement of NFκB in the antirheumatic potential of Chenopodium album L., aerial parts extracts.

ETHNOPHARMACOLOGICAL RELEVANCE: Chenopodium album L. (C. album) is commonly known as Bathua in Hindi (Family: Chenopodiaceae). Traditionally, the plant is used as a laxative, diuretic, sedative and the infusion of the plant is used for the treatment of rheumatism. However, no scientific validation is available on the antirheumatic potential of the plant. In the present investigation, role of NF kappa B (NFκB) in the antiarthritic potential of extracts of aerial parts of Chenopodium album was explored and evaluated. METHODS: The defatted aerial parts of Chenopodium album were successively extracted with ethylacetate, acetone, methanol and 50% methanol to study their antioxidant capacity followed by antiarthritic potential using Complete Freund׳s adjuvant (CFA) induced arthritis model in rats. The polyphenol, flavonoid and flavanone contents of different extracts were quantified and correlated with their antioxidant capacity, antiarthritic activity and NFκB inhibition potential. RESULTS: The experimental data indicated that the acetone extract of Chenopodium album (ACCA) has shown significant reduction in rat paw edema (80.13%) at dose level of 200 mg/kg per oral in 21 days of this study. On 22nd day, hematological and biochemical parameters were estimated and it was observed that the altered hematological parameters (Hb, RBC, WBC and ESR), biochemical parameters (Serum creatinine, total proteins and acute phase proteins) and loss in body weight in the arthritic rats were significantly brought back to near normal level by the ACCA extract. ACCA extract significantly decreased the NFκB expression in paraventricular nucleus of hypothalamus and this effect is comparable with standard indomethacine in CFA treated rats. The polyphenolic and flavonoid content of different extracts were in the range of 14.56±0.21-42.00±0.2 mg (gallic acid equivalent/g extract) and 2.20±0.003-7.33±0.5 mg (rutin equivalent/g extract) respectively. CONCLUSION: The antiarthritic activity possessed by ACCA extract can be correlated directly to its antioxidant potential, high flavonoidal content achieved by successive extraction and its capacity to inhibit the NFκB protein, as proven by immunohistochemistry study.

Antidiabetic potential of unripe Carissa carandas Linn. fruit extract.

ETHNOPHARMACOLOGICAL RELEVANCE: Carissa carandas commonly known as Karanda have a long history of use in traditional system of medicine. It is used by tribal healers of Western Ghat region of Karnataka as hepatoprotective and antihyperglycemic. However, no scientific data is available to validate the folklore claim. The present study has been designed to evaluate its unripe fruit for the antidiabetic activity. AIM: In the present study, methanol extract of unripe fruits and its fractions were studied for its antidiabetic potential. MATERIALS AND METHODS: The methanol extract and its fractions were screened for antidiabetic activity in alloxan induced diabetic rats. The polyphenolic, flavonoid and flavanone contents of methanolic extract and its fractions were also determined and correlated with its antidiabetic activity. RESULTS: The experimental data indicated that the methanol extract and its ethyl acetate soluble fraction has significantly lowered the elevated blood glucose levels by 48% (p<0.001) and 64.5% (p<0.001) respectively at dose level of 400mg/kg per oral after 24h as compared to diabetic control. In order to assess the role of polyphenolic components in the relevant activity, polyphenolic and flavonoid contents were determined. The polyphenolic and flavonoid content of methanol extract and its ethyl acetate soluble fraction were found to be 15.8 ± 1.2mg and 18.55 ± 0.34 mg (gallic acid equivalent/g extract) and flavonoid content 2.92 ± 0.03 mg and 1.534 ± 0.30 mg (rutin equivalent/g extract) respectively. CONCLUSION: The increased antidiabetic potential of ethyl acetate fraction over methanol extract is due to its partial purification achieved by fractionation which resulted in increase in degree of polymerization and segregation of secondary metabolites.

Evaluation of antidiabetic antihyperlipidemic and pancreatic regeneration, potential of aerial parts of Clitoria ternatea

The aim of the present study was to investigate the pancreatic regeneration potential of of diferent fractions of the ethanol extract Clitoria ternatea L., Fabaceae. The antidiabetic and antihyperlipidemic potential was evaluated in streptozotocin-induced diabetic rats and correlated with its in vivo and in vitro antioxidant activity. The extract and its fractions were initially screened for acute and sub-chronic antidiabetic activity in the dose range of 100200 mg/kg. The most potent extract and fractions were further evaluated for pancreatic β-cells regeneration activity along with antioxidant and antihyperlipidemic activity. The polyphenolic, flavonoid and flavanone contents were assessed and correlated with its antidiabetic activity. The most significant pancreatic regeneration activity, antidiabetic and antihyperlipidemic activity and was shown by ethanol extract and butanol soluble fraction at a dose level of 200 mg/kg, while rutin was found to be least potent. In conclusion, pancreatic regeneration studies of ethanol extract treated rats show nesidioblastosis. It is also suggested that the factors causing regeneration are present within the pancreas. The newly generated islets may have formed from the ductal precursor cells and reduced oxidative stress helps in restoration of β-cell function.