Crude Aqueous-methanolic Extract Research Articles

Safety assessment of crude aqueous methanol extract of Annona senegalensis stem bark: acute and sub-chronic toxicity studies.

Annona senegalensis Pers., (wild custard apple), is a shrub used traditionally to treat respiratory and skin diseases. Previous studies have demonstrated its anti-malaria, anti-snake envenomation and anti-cancer activities. However, its toxicological profile remains limited, particularly in male and female animals. This study aims to evaluate the safety of crude aqueous methanol extract of Annona senegalensis stem bark (AMEAS) through acute and sub-chronic toxicity studies. The stem bark of A. senegalensis was collected, air-dried, pulverized, and extracted using 70% methanol. Phytochemical screening, elemental analysis, and acute toxicity evaluation were carried out on AMEAS. Sub-chronic toxicity study was conducted on Wistar rats of both sexes at different doses administered orally for 28 days. Elemental analysis revealed the presence of heavy metals and essential mineral elements with the highest contents being calcium (59.88%) and potassium (25.39%). Acute toxicity testing showed no mortality up to 5000 mg/kg, suggesting an LD50 greater than 5000 mg/kg. In the sub-chronic toxicity study, no mortality or significant harmful effects were observed. The blood glucose decreased from 13.68 mMol/L at 250 mg/kg to 10.71 mMol/L at 1000 mg/kg, much lower than the distilled water group (17.06 mMol/L). In conclusion, the extract appeared to be well-tolerated, with no obvious adverse effects. AMEAS is rich in Calcium (Ca) and potassium (K). It has been shown to have LD50 greater than 5000 mg/kg and is assumed to be safe. On repeated use, AMEAS may cause hypoglycemia and weight loss which may be useful in managing diabetes and obesity respectively.

In vitro evaluation of anticancer, antiviral, and antioxidant properties of an aqueous methanolic extract of Rhus typhina L. leaves

In recent decades, natural products of plant origin have predominated among the sources of new therapeutic agents. Plants, which are intensively used in folk medicine, are the focus of particular interest, and being subjected to in-depth analyses for the assessment of their pharmacological properties. The objective of the present work was to evaluate the in vitro anticancer, antiviral, and antioxidant properties of a crude aqueous methanolic extract from the leaves of Rhus typhina L., which is a plant species with broad applications in traditional phytotherapy. The anticancer capacity of the extract was assessed on two human breast cancer cell lines (MCF7 and MDA-MB-231) using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) cell proliferation assay, and its cell growth inhibitory effect on cancer cells was compared with its action on a non-cancerous breast epithelial cell line (MCF10A). Moreover, the in vitro antiviral activity of the plant extract was studied against viral strains of three taxonomic groups: herpes simplex virus type 1 (HSV-1), coxsackievirus B1 (CVB1), and influenza A virus (IAV/H3N2) using a cytopathic effect inhibition test, extract cytotoxicity evaluation through the neutral red uptake assay, a virucidal assay, a viral attachment assay, and cell pre-treatment with the extract. In turn, a radical scavenging assay with 2,2-diphenyl-1-picrylhydrazyl (DPPH) was applied to explore the antioxidant potential of R. typhina leaf extract. Results indicated that the plant extract possessed antiproliferative activity against the tested cell lines which was stronger toward cancer cells. A modest inhibitory effect on IAV/H3N2 replication was detected; as well as on an effect on herpes virions and on the adsorption of the HSV-1 and IAV/H3N2 strains; and a protective effect on uninfected cells before HSV-1 contact. The leaf extract of R. typhina also exhibited a strong free radical scavenging activity. The results of the present work demonstrated the pharmacological potentials of the plant extract, which warrants further and more detailed study in the future.

EVALUATION OF ANTICOCCIDIAL ACTIVITY OF METHANOLIC EXTRACT OF RICINUS COMMUNIS IN BROILER

Aims and Objectives: Coccidiosis is recognized as the parasitic disease which has the greatest economic impact on poultry production. The emergence of resistant strains to available drugs has become a major problem in order to treat/control coccidiosis. Botanicals can act as alternative to anticoccidial drugs. This study has therefore, been planned to evaluate both in vitro and in vivo anticoccidial activity of the plant R. communis.
 Methods: In in vitro trial 20, 10, 5, 2.5, 1.25 and 0.625 percent, DMSO dissolved crude aqueous methanolic extract of R. communis was used to investigate its inhibitory effect upon sporulation of oocysts. The research was done in University of Agriculture, Faisalabad, Pakistan, with the collaboration of Parasitology, Pathology and Poultry departments. In in vivo trial, the plant was used at three graded concentrations for evaluation of its anticoccidial activity in broiler birds. A total of 144 (one-day-old) broiler chicks were divided into six groups each having 24 chicks.  At age of 15 days, groups I, II and III were given 4%, 5% and 6% of dried powder of R. communis respectively. Group IV was served as positive control (infected, toltrazuril treated), group V as negative control (infected, non-medicated) and group VI was serve as non-infected and non-medicated control. All groups except group VI were infected orally with 50,000 sporulated oocysts of mixed Eimeria species at 18th day of age.
 Result: After 7 days of inoculation, six birds from each group were slaughtered to get results on oocysts score, lesion score, relative organ weight, hematology and immunomodulatory effect. Data was analyzed by using analysis of variance (ANOVA) and group means were compared by Duncan’s multiple range tests.
 Conclusion: It was concluded that R. communis can be used as a prophylactic and therapeutic agent at local and regional level for the control of coccidiosis in broilers.
                    
 Peer Review History: 
 Received: 18 July 2021; Revised: 7 August; Accepted: 6 September, Available online: 15 September 2021
 Academic Editor:  Dr. Nuray Arı, Ankara University, Turkiye, ari@ankara.edu.tr
 UJPR follows the most transparent and toughest ‘Advanced OPEN peer review’ system. The identity of the authors and, reviewers will be known to each other. This transparent process will help to eradicate any possible malicious/purposeful interference by any person (publishing staff, reviewer, editor, author, etc) during peer review. As a result of this unique system, all reviewers will get their due recognition and respect, once their names are published in the papers. We expect that, by publishing peer review reports with published papers, will be helpful to many authors for drafting their article according to the specifications. Auhors will remove any error of their article and they will improve their article(s) according to the previous reports displayed with published article(s). The main purpose of it is ‘to improve the quality of a candidate manuscript’. Our reviewers check the ‘strength and weakness of a manuscript honestly’. There will increase in the perfection, and transparency. 
 Received file:                Reviewer's Comments:
 Average Peer review marks at initial stage: 5.5/10
 Average Peer review marks at publication stage: 7.0/10
 Reviewers:
 Ahmad Najib, Universitas Muslim Indonesia, Makassar, Indonesia, ahmad.najib@umi.ac.id
 Dr. Dennis Amaechi, MrsFoluBabade Mini Estate , Flat 5 by Old Soldiers Quarter, Sabongari/Bwari, Abuja- Federal Capital Territory, Nigeria. amaechitoexcel@yahoo.com 
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 ANTI-INFLAMMATORY, ANALGESIC AND COGNITIVE ENHANCER PLANTS PRESENT IN BANGLADESH: A STUDY REVIEW

Antioxidant and enzyme-inhibiting activity of lyophilized extract from Clinopodium vulgare L. (Lamiaceae)

Clinopodium vulgare L. (Lamiaceae) was used in the traditional Bulgarian medicine for treatment of wounds, diabetes and gastric ulcers. The aim of the present study was to evaluate the antioxidant capacity of the extract (CVE) and fractions from C. vulgare (CV) using DPPH, ABTS and FRAP methods. Enzyme inhibitory activity against acetylcholinesterase, α-glucosidase and α-amylase was also investigated. Rosmarinic acid was used as a positive control. The fraction CV3 demonstrated the highest radical scavenging activity with IC50 values of 0.02 mg/ml (DPPH) and 0.0002 mg/ml (ABTS), as well as the strongest ferric reducing potential (FRAP) of 0.89 mM TE/mg dw. The crude aqueous-methanol extract of C. vulgare also showed high activity with IC50 values of 0.05 mg/ml (DPPH), 0.04 mg/ml (ABTS) and 0.89 mM TE/mg dw (FRAP). Moreover, CV3 demonstrated moderate α-glucosidase and α-amylase inhibitory potential.

Comparative anthelmintic efficacy of Arundo donax, Areca catechu, and Ferula assa-foetida against Haemonchus contortus.

In the present study, anthelmintic activities of Arundo (A.) donax L., Areca (Ar.) catechu L., and Ferula (F.) assa-foetida L. were determined. Leaves of A. donax L., latex of F. assa-foetida L. and seeds of Ar. catechu L. in different solvent fractions were subjected to in vitro (egg hatch assay; EHA, and adult motility assay; AMA) and in vivo (faecal egg count reduction test; FECRT) tests of anthelmintic activity using Haemonchus contortus model. In the AMA, crude aqueous methanol extracts (CAME) and ethyl acetate fractions of F. assa-foetida at 10 hr post-treatment showed maximum mortality of H. contortus at 12.5-50 mg mL-1. In the EHA, CAME of F. assa-foetida was identified as a potent ovicide based on its low LC50 (16.9 µg mL-1), followed in order by Ar. catechu and A. donax. Results from the FECRT also showed the extract of F. assa-foetida L. to be more effective than those of Ar. catechu L. and A. donax L., against the gastrointestinal parasitic nematodes. Chloroform and ethyl acetate fractions showed better anthelmintic activities against the adult worms in vitro, while CAME of these plants were better than their crude powders in vivo. It is recommended to document and investigate indigenous knowledge of possible medicinal plants to plan scientific trials that may justify their endorsement.

Ameliorative effect of Parinari curatellifolia seed extracts on sodium nitroprusside–induced cardiovascular toxicity in rats

The pharmacological potential of Parinari curatellifolia has remained largely unexplored by the scientific community and little information is available to support the folkloric and ethnobotanical uses of the plant. This present study was carried out to investigate the ameliorative effect of P. curatellifolia seed extracts on sodium nitroprusside (SNP) toxicity on the heart and artery in rats. P. curatellifolia seed crude aqueous-methanolic extract (CMPC) as well as n-hexane (HFPC), dichloromethane (DFPC), ethyl acetate (EFPC), and n-butanolic (BFPC) fractions was prepared and screened for phytoconstituents. In the in vitro test, crude extract and fractions were assessed for antiperoxidative potential in SNP-induced lipid peroxidation (LPO) in rat heart and artery. For in vivo assessment, CMPC (400, 500, and 600 mg/kg bwt) and the fractions (5 and 6 mg/kg bwt) were co-administered with 5 mg/kg SNP to Wistar rats for 7 days. Extent of LPO in the heart and artery and serum activities of aspartate transaminase (AST) and alanine transaminase (ALT) as well as concentrations of bilirubin, creatinine, urea, and uric acid were evaluated. The result of phytochemical screening revealed the presence of alkaloids, tannins, saponins, flavonoids, terpenoids, phlobatannins, and cardiac glycosides. SNP toxicity significantly (p < 0.05) increased LPO, bilirubin, creatinine, urea, and uric acid levels as well as the activities of AST and ALT. However, the extracts reversed the effect of SNP toxicity. The study revealed the protective effect of P. curatellifolia against SNP-induced cardiovascular toxicity in rats. The study plant could serve as a good therapeutic agent to combat cardiovascular-related complications.

In Vitro Antimicrobial Activity and GC-MS Analysis of Crude Aqueous Methanolic Extract Produced from Leaves of Eucalyptus species

Objectives:To evaluate the antimicrobial activity of the aqueous methanolic extract was given by the leaves of Eucalyptus sp.growing in Thi-Qar Province, south of Iraq. Additionally, to detect the compounds within the extract by using the technique of gas chromatography-mass spectrometry (GC-MS). Methodology: The healthy leaves of Eucalyptus sp. were collected, washed with tap water then by distilled water, dried, and pulverized through using a small mill to make the dried leaves as a powder. The leaves powder was extracted by absolute petroleum ether then aqueous methanol (80%) for obtaining a sticky crude extract of the aqueous methanol filtrate. The crude extract was tested against some microbial pathogens as well as analysis of the extract using the technique of GC-MS and phytochemical tests. Results:The current study showed the antimicrobial activity of the crude aqueous methanolic extract obtained from Leaves ofEucalyptus sp. by which 40000 µg∕ml of the extract exhibited inhibition zones around Staphylococcus aureus, Streptococcus mutans, Escherichia coli, and Klebsiella sp. besides Candida albicans that the inhibition zones were measured to be 30, 27, 26, 30, and 38 mm respectively. The chemical tests appeared the crude extract containsalkaloids, glycosides, flavonoids, tannins, andsaponin glycosides. GC-MS analysis detected constituents of the extract are dimethylsulfoxoniumformylmethylide, diethyl phthalate, benzene,1,1'-(1,2-cyclobutanediyl)bis-trans-,diisoctyladipate, and 2-methyl-7-phenylindole. Conclusion: The present study concluded that the leaves ofEucalyptussp. possess the bioactive products which are needed to be separatedas the pure compoundsby using techniques of chromatography in order to separately tested in vitro and in vivo, then application of spectrometry techniques for characterization of their chemical structure. Finally, these compounds can be used to treat diseasesas drugs in hospitals.

In Vitro Antimicrobial Activity and GC-MS Analysis of Crude Aqueous Methanolic Extract Produced from Leaves of Eucalyptus species..

In Vitro Antimicrobial Activity and GC-MS Analysis of Crude Aqueous Methanolic Extract Produced from Leaves of Eucalyptus species..

Oestrous cycle of Wistar rats altered by sterol and triterpenes rich fraction of Adansonia digitata (Linn) root bark - A scientific rationale for contraceptive use

Objective: To investigate the effect of different fractions from the aqueous methanolic root bark of Adansonia digitata (Linn) on the estrous cycle of female Wistar rats and to elucidate their phytochemical content.Methods: Crude aqueous methanol extract of Adansonia digitata (Linn) root bark was fractionated by column chromatography using various ratios of hexane, acetyl acetate, methanol and distilled water. Fractions with similar trails on a thin layer chromatography were pooled into four fractions. Each fraction was then screened for its phytochemical content. Forty mature Wistar female rats weighing (135±28) g with regular estrous cycles were given each fraction at two dose levels while a control group received distilled water of equal volume. Treatments were given per os for 15 days starting from the day of estrus. Vaginal smears were observed daily for the duration of each cycle, frequency of occurrence of each stage of the estrous cycle and number of cycles for 15 days before and during treatment. At the end of treatment, rats were humanely sacrificed. Ovaries and uteri were weighed and processed for histopathology.Results: Hexane acetyl acetate and acetyl acetate methanol fractions at 150 mg/kg body weight and 300 mg/kg body weight prolonged the occurrence of proestrus and diestrus and produced atretic cyst-like ovaries at 300 mg/kg body weight. Acetyl acetate methanol treated rats also had low relative ovarian weights when compared to the control.Conclusions: Sterols and triterpenes in hexane acetyl acetate and acetyl acetate methanol fractions of the root bark may be responsible for the disruption of estrous cycle in female Wistar rats and may have a contraceptive effect.

Chemical, biological and molecular modelling analyses to probe into the pharmacological potential of Antidesma madagascariense Lam.: A multifunctional agent for developing novel therapeutic formulations

Antidesma madagascariense Lam. (AM), an indigenous medicinal plant to the Mascarene Islands, is used for the treatment of several diseases. We endeavoured to validate its use via evaluating the kinetics of inhibition of crude aqueous extract (CAE) and crude methanol extract (CME) of AM against key metabolic enzymes (pancreatic lipase, cholesterol esterase [CEase], acetylcholinesterase [AChE], and urease). In vitro antiglycation, antioxidant, cytotoxicity using iCELLigence real time cell analysis system and WST-1 methods, were used. LC–ESI–MS/MS was employed to determine the phenolic composition of the extracts and interaction of selected compounds to the studied enzymes was determined using in silico docking. AChE was inhibited by the CME of AM and CEase by the CAE. Both extracts were active inhibitors of urease and pancreatic lipase. Hyperoside (271.97 μg/g extract), present in large amount in the CME, docked to the enzymatic pocket of urease and CEase. The extracts showed competitive and mixed inhibition of urease and pancreatic lipase, respectively. The antioxidant capacity of the CME (6.61 μg GAE/mg crude extract) was higher compared to CAE (2.20 μg GAE/mg crude extract). AM extracts were significantly (p < 0.05) less potent than aminoguanidine in preventing advanced glycation end products formation. Toxicological screening revealed that both extracts were non-toxic on HEK-293 cells. AM crude extracts at concentrations ranging from 78 to 312 μg/ml did not cause a visible change in cell morphology compared to control. This study supports the safe use of AM as a biomedicine for the management and/or treatment of common non-communicable diseases.

IN VITRO FREE RADICAL SCAVENGING ACTIVITY OF ARISTOLOCHIA TAGALA

Objective: Study of antioxidant activity of a crude aqueous-methanol extract of Aristolochia tagala and its fractions and identification of the compounds with antioxidant activity.Methods: The antioxidant activity was assayed by the ability to scavenge free radicals such as superoxide, nitric oxide and ABTS radical cation and the identification of compounds was carried out by LC/MS analysis.Results: Fraction I of Aristolochia tagala showed the highest free radical scavenging activity and compounds responsible for its activity were identified as magnoflorine, apigenin dimethyl ether, aristolone, and N-trans-feruloyldopamine.Conclusion: The free radical scavenging property of the compounds present in Aristolochia tagala may be one mechanism that contributed to medicinal property exhibited by this plant.

Identification of compounds of Aristolochia tagala and apoptotic activity in HeLa cells

Background: Plants contain secondary metabolite used as drugs/medicines for the treatment of various diseases. Aristolochia tagala is used for the treatment of several diseases. Our study reported the chemopreventive potential of crude aqueous-methanol extract against diethylnitrosamine-induced hepatocellular carcinoma in BALB/c mice. A few articles have reported the presence of pharmacologically active compounds. Objective: Identification of biologically active compounds can give an insight into the mechanism of action of A. tagala and its potential development into modern drugs for the treatment of various diseases including cancer. Materials and Methods: Aqueous methanol extract (ATC) was prepared from roots of A. tagala and fractionated by column chromatography. The compounds present in ATC were identified and characterized by liquid chromatography (LC)–high-resolution mass spectrometry. ATC as well as the fractions (FI–FIV) were tested for their cytotoxic effect in HeLa cells by 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide assay, and apoptotic events were analyzed by flow cytometry. The fraction that showed the most efficient cytotoxic effect against HeLa cells was purified by high-performance thin-layer chromatography. Purified compounds were again assayed for their apoptotic and cytotoxic effect. The most active compound was identified and characterized by electrospray ionization high-resolution mass spectrometry and LC–tandem-mass spectrometry. Statistical analysis was carried out using one-way anova followed by Tukey's multiple comparisons test. Results: A total of 21 compounds were identified; aristolochic acid I, aristolactam IIIa, β-sitosterol, kaempferol, and stigmasterol were previously reported in A. tagala and other compounds in other species of Aristolochia, and some compounds were reported to have anticancer, anti-inflammatory activities. From our study, compound S7 showed the highest cytotoxic and apoptotic activity and was identified as aristolochic acid I. Aristolochic acid earlier has been reported to have antitumor and anticancer effects, but lately, it has also been reported to have nephrotoxic effect. Conclusions: A. tagala was found to contain a number of compounds with reported biological activity. This plant and its related species can, therefore, be exploited for the extraction and isolation of these compounds with no toxicity. Abbreviations used: ATC: Aqueous-methanol extract, AA-I: Aristolochic acid I, CQA: Caffeoylquinic acid, CHCl3: Chloroform, MTT: 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide, ESI: Electrospray ionization, FITC: Fluorescein isothiocyanate, HCC: Hepatocellular carcinoma, HPLC: High-performance liquid chromatography, HRMS: High-resolution mass spectrometry, LC-MS/MS: LC-tandem-mass spectrometry, MeOH: Methanol, PBS: Phosphate buffered saline, PVDF: Polyvinylidene difluoride, RF: Radio frequency, R F: Retardation factor, XEVO-TQD: Xevo triple quadrupole

Anti-ulcer and anti-Helicobacter pylori potentials of the ethyl acetate fraction of Physalis alkekengi L. var. franchetii (Solanaceae) in rodent

Ethnopharmacological relevancePhysalis alkekengi L. var. franchetii (Solanaceae) has been widely used in Chinese folk medicine due to its wide distribution throughout the country, for the treatment of a wide range of diseases including heat and cold, sore throat, fever, fungal infection, inflammation, toothache, rheumatism, burn, analgesic, ulcer and urinary diseases. However, the effect of P. alkekengi var. franchetii on ulcer and Helicobacter pylori infection has not been reported to date. Aim of the studyThis study was designed to investigate the anti-inflammatory, anti-ulcer, anti-Helicobacter pylori and analgesic properties of ethyl acetate fraction of the crude aqueous methanolic extract from the aerial parts of the plant P. alkekengi L. var. franchetii in rodents. Materials and methodsAcute toxicity of the crude extract of P. alkekengi L. var. franchetii (PAF) was evaluated in rats. The petroleum ether fraction (PEF), butanol fraction (BF), ethyl acetate fraction (EAF) and aqueous fraction (AF) of crude aqueous methanolic extract from PAF were screened for anti-inflammatory and anti-ulcer potential at doses of 100, 250 and 500mg/kg (p.o.), using carrageenin-induced hind paw edema and ethanol-induced gastric lesions test in rats. In vitro anti-Helicobacter pylori activity of EAF was assayed subsequently. In addition, three doses of EAF were evaluated for analgesic activity using hot plate and writhing tests, respectively. Finally, we performed a phytochemical analysis of EAF. ResultsFour fractions of crude extract from PAF significantly reduced the paw volume in carrageenin-induced hind paw edema model at different doses (100, 250 and 500mg/kg, p.o.). The fraction EAF at a dose of 500mg/kg exhibited the highest (75.92%) (0.150 ± 0.045***, ***p < 0.001) anti-inflammatory potential, which is similar to indomethacin (***P < 0.001)（0.120 ± 0.014***, 80.74% inhibition of inflammation） at 5mg/kg. Pretreatment with EAF (500mg/kg, p.o.) significantly reduced the intensity of gastric mucosal damage and showed higher gastroprotective activity (90.6%) when compared to the standard drug famotidine (84.6%). In addition, EAF fraction also showed a moderate (P < 0.05) anti-Helicobacter pylori activity with a minimal inhibition concentration (MIC) of 500μg/ml. Furthermore, pain sensation was effectively inhibited at 500mg/kg, p.o. of EAF as manifested by an increase (p < 0.001) of latency time in hot plate from 30 to 90min and a decrease (p < 0.001) in count of writhing induced by acetic acid. By HPLC, we determined some steroid, terpenoid and flavonoids (four compounds): kaempferol, quercetin, Blumenol A and physalindicanols A, which were isolated from the ethyl acetate fraction and identified using 1H NMR and 13C NMR spectra analysis. ConclusionThis study demonstrated the anti-inflammatory, anti-ulcer, anti-Helicobacter pylori and analgesic properties of EAF of the crude extract from PAF thus justifying its traditional usage.

English

Current treatment of trypanosomosis in sub-Saharan Africa is associated with widespread inefficiency. There is therefore the need to find more effective drugs against the disease from promising traditional medicinal herbs. This work is aimed at evaluating ex vivo anti-trypanosomal effect of stem-bark extracts of S. longipedunculata against T. brucei brucei. One hundred microlitre of crude methanol, ethyl acetate and aqueous methanol extracts of S. longipedunculata at concentrations of 3 and 6 mg/ml each were mixed with 50 µl of blood containing 8.1 × 106 trypanosomes and incubated at 37°C for 90 min. Similarly, diminazene aceturate (10 µg/ml), physiological saline solution (50 µl) and blood (100 µl) containing trypanosomes only served as treated, negative and untreated controls, respectively. Motility of the parasite was monitored under light microscope (×400) at 5 min interval throughout the 90 min observation period. All experiments were done in duplicate. The mixtures were subsequently inoculated into rats that were not previously infected with trypanosomes. Phytochemical screening of the extracts revealed the presence of carbohydrates, cardiac glycosides, saponins, steroid, triterpenes, flavonoids and tannins. However, aqueous and ethyl acetate fractions were devoid of flavonoids. The crude methanol immobilized the parasites within 75 min, while ethyl acetate and aqueous extracts induced slight reduction in motility of the parasite at 90 min of incubation. However, inoculated rats developed infection and succumbed to the infection. It is concluded that the stem-bark of the plant possesses trypanostatic, but not trypanocidal, activity against the parasite.   Key words: Antitrypanosomal, drug incubation infectivity test, in vitro, phytochemical screening, stem-bark.

Ramalin, a Novel Phenyl Hydrazide from the Lichen Ramalina Terebrata; Isolation, Total Synthesis and Biological Activities

Ramalin, a new L-glutamic acid derivative of phenylhydrazide (Y-glutamyl-N'-(2-hydroxyphenyl) hydrazide, 1) was isolated from the Antarctic lichen, Ramalina terebrata after a series of bioactivity guided fractionation of crude aqueous methanolic extract. Ramalin showed stronger antioxidant activities than commercially available standards, ascorbic acid, trolox, BHA, kojic acid in both, in vitro and in vivo test systems. In addition, ramalin showed no/less toxicity effects against two human cell lines; fibroblast (CCD-986SK) cells and keratinocyte (HaCaT). Thus, realign merits for cosmetic application and industrial scale production were needed. We developed a cost effective total synthesis of ramalin with 71.5% yield and described here.

Antidiarrhoeal, antisecretory and antispasmodic activities of Matricaria chamomilla are mediated predominantly through K+-channels activation

BackgroundMatricaria chamomilla commonly known as “Chamomile” (Asteraceae) is a popular medicinal herb widely used in indigenous system of medicine for a variety of ailments. However, there is no detailed study available showing its effectiveness in hyperactive gut disorders like, abdominal colic and diarrhoea. This study was designed to determine the pharmacological basis for the folkloric use of Matricaria chamomilla in diarrhoea.MethodsThe crude aqueous-methanolic extract of Matricaria chamomilla (Mc.Cr) was studied for its protective effect in mice against castor oil-induced diarrhoea and intestinal fluid accumulation. The isolated rabbit jejunum was selected for the in-vitro experiments using tissue bath assembly coupled with PowerLab data acquisition system.ResultsOral administration of Mc.Cr to mice at 150 and 300 mg/kg showed marked antidiarrhoeal and antisecretory effects against castor oil-induced diarrhoea and intestinal fluid accumulation, simultaneously, similar to the effects of cromakalim and loperamide. These effects of plant extract were attenuated in animals pretreated with K+ channel antagonist, glibenclamide (GB) or 4-aminopyridine (4-AP). When tested in isolated rabbit jejunum, Mc.Cr caused a dose-dependent (0.3-3 mg/ml) relaxation of spontaneous and low K+ (25 mM)-induced contractions, while it exhibited weak inhibitory effect on high K+ (80 mM). The inhibitory effect of Mc.Cr on low K+-induced contractions was partially inhibited in the presence of GB, while completely blocked by 4-AP. Cromakalim, an ATP-sensitive K+ channel opener, caused complete relaxation of low K+-induced contractions with little effect on high K+. Pretreatment of tissues with GB blocked the inhibitory effects of cromakalim on low K+, while the presence of 4-AP did not alter the original effect. Verapamil, a Ca++ channel antagonist, caused complete relaxation of both low and high K+-induced contractions with similar potency. The inhibitory effect of verapamil was insensitive to GB or 4-AP. When assessed for Ca++ antagonist like activity, Mc.Cr at high concentrations caused rightward shift in the Ca++ concentration-response curves with suppression of the maximum response, similar to the effect of verapamil, while cromakalim did not show similar effect.ConclusionsThis study indicates that Matricaria chamomilla possesses antidiarrhoeal, antisecretory and antispasmodic activities mediated predominantly through K+-channels activation along with weak Ca++ antagonist effect.

Aqueous-Methanol Extract of Gracilaria verrucosa Induces Cytochrome c Release From Mitochondria

Seaweeds is one of many sources of natural bioactive compounds that can be utilized in the fields of pharmaceuticals and health. However, research on utilization of the seaweed's bioactive compounds from Indonesia is still very limited. Therefore, the objective of this research is to study the ability to induce apoptosis of the extracts of Gracilaria verrucosa harvested from Sayang Heulang, a beach in Pameungpeuk, Garut, West Java. The study begins with bioactive compounds extraction using chloroform p.a, methanol p.a and 50mM phosphate buffer pH 7.6 with a ratio 2:1:0.8, resulting crude aqueous-methanol and chloroform extracts. G. verrucosa produced 62.73% (w/w) crude aqueous-methanol extract and 1.07% (w/w) chloroform extract. Mitochondria for apoptosis test were isolated from rat liver. The treatment of 10% v/v crude aqueous-methanol extract from G. verrucosa for five hours at 37 oC toward mitochondrial suspension leads to formation of mitochondrial aggregates and study of the released of cytochrome c showed that those treatment released 8.7μM cytochrome c which has equal to 11.4% of mitoxantrone 500μM activity. In conclusion, our results indicate that aqueous-methanol extract of G. verrucosa able to induce apoptosis through mitochondrial pathway involving cytochrome c release from mitochondria.

Pharmacological basis for the medicinal use of Linum usitatissimum (Flaxseed) in infectious and non-infectious diarrhea

Ethnopharmacological relevanceLinum usitatissimum, commonly known as Flaxseed has traditionally been used for the management of diarrhea and gastrointestinal infections. This study was planned to assess pharmacological basis for the medicinal use of Flaxseed in infectious and non-infectious diarrhea. Materials and methodsThe crude aqueous-methanolic extract of Flaxseed was studied using the in vivo castor oil-induced diarrhea, gut motility and enteropooling assays. Mechanistic basis was further elucidated by testing the inhibitory effect on spontaneously contracting isolated rabbit jejunum preparations, suspended in a 10ml tissue bath containing Tyrode׳ solution, maintained at 37°C and aerated with carbogen. Antibacterial efficacy of the Flaxseed extract was tested against different enteric and non-enteric pathogenic bacteria using in vitro antibacterial assays. ResultsFlaxseed extract reduced the diarrheal score in mice, by 39%, 63.90% and 68.34% at the respective doses of 100, 300 and 500mg/kg. Intestinal secretions were reduced by 24.12%, 28.09% and 38.80%, whereas the intestinal motility was reduced by 31.66%, 46.98% and 56.20% at respective doses of 100, 300 and 500mg/kg. When tested on isolated rabbit jejunum preparations, Flaxseed extract produced a dose-dependent inhibition of both spontaneous and high K+ (80mM)-induced contractions, and shifted the concentration–response curves of Ca++ to the right with suppression of the maximum response, similar to that caused by verapamil. Flaxseed extract was found to possess bactericidal activity at the tested concentrations of 12.5mg/ml, against vancomycin-resistant Enterococcus faecalis (100%), Escherichia coli K1 (88.88%), methicillin-resistant Staphylococcus aureus (98.76%), Bacillus cereus (92.64%), Pseudomonas aeruginosa (76.83%) and Salmonella typhi (26.91±3.35%). The concentration of 10mg/ml showed bactericidal effects against all the aforementioned pathogens except Escherichia coli K1, whereas for Pseudomonas aeruginosa and Salmonella typhi, it was bacteriostatic at this concentration. ConclusionsOur results indicate that Linum usitatissimum (Flaxseed) extract exhibits antidiarrheal and antispasmodic activities by virtue of its antimotility and antisecretory effects which are mediated possibly through inhibition of Ca++ channels, though additional mechanism(s) cannot be ruled out. Flaxseed extract proved effective against both enteric and non-enteric pathogens causing diarrhea, thus ensuring wide coverage and rationalizing its medicinal use in both the infectious and non-infectious diarrhea.

Potential antiradical and alpha-glucosidase inhibitors from Ecklonia maxima (Osbeck) Papenfuss

Alpha-glucosidase inhibitors play a potential role in the treatment of type 2 diabetes by delaying glucose absorption in the small intestine. Ecklonia maxima, a brown alga which grows abundantly on the west coast of South Africa, is used to produce alginate, animal feed, nutritional supplements and fertilizer. The crude aqueous methanol extract, four solvent fractions and three phlorotannins: 1,3,5-trihydroxybenezene (phloroglucinol) (1), dibenzo [1,4] dioxine-2,4,7,9-tetraol (2) and hexahydroxyphenoxydibenzo [1,4] dioxine (eckol) (3) isolated from E. maxima were evaluated for antiradical and alpha-glucosidase inhibitory activities. All the phlorotannins tested had strong antioxidant activities on DPPH free radicals with EC50 values ranging from 0.008 to 0.128μM. Compounds 2 and 3 demonstrated stronger antioxidant activity and an alpha-glucosidase inhibitory property than positive controls. These results suggest that E. maxima could be a natural source of potent antioxidants and alpha-glucosidase inhibitors. This study could facilitate effective utilization of E. maxima as an oral antidiabetic drug or functional food ingredient with a promising role in the formulation of medicines and nutrition supplements.

Anti-leukemic activity of Wattakaka volubilis leaf extract against human myeloid leukemia cell lines

Ethnopharmacological relevanceWattakaka volubilis has been traditionally used in Ayurvedic medicine in India for treatment of several ailments such as bronchial asthma, inflammations, tumors, piles, leucoderma, application to boils, rat bite etc. Aim of the studyThe present study was designed to investigate anti-leukemic activity of the crude aqueous methanolic extract and to identify active compounds from the leaves of Wattakaka volubilis. Materials and methodsThe leaves of Wattakaka volubilis were extracted with aqueous methanol. Liquid–liquid fractionation of the crude methanolic extract with different organic solvents was done and the fractions were screened for in vitro anti-leukemic activity using different leukemic cell lines. The active fractions were then subjected to chromatographic separation for isolation of bioactive compounds. Structure of isolated compound was elucidated by spectroscopic methods. The in vitro anti-leukemic activities of different extracts of the leaves and isolated compound WVP were studied in U-937, HL-60 and K-562 cell-lines by using cell count, MTT [(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-tetrazolium bromide] and DNA laddering assays, flow-cytometric and confocal microscopic techniques. ResultsKaempferol-3-O-[α-l-rhamnopyranosyl-(1→4)-O-α-l-rhamnopyranosyl-(1→6)-O]-β-d-glucopyranoside (WVP) was isolated from crude leaves extract of Wattakaka volubilis. Both the n-butanolic extract (WVB) of Wattakaka volubilis and its isolate WVP were found to be responsible for in vitro anti-leukemic activity. The IC50 values of WVB were found be 120, 100 and 50(μg/ml) in U937, K562, and HL-60 cell lines, respectively. Whereas, the pure isolate WVP exhibited anti-leukemic activity with IC50 values of 13.5, 10.8, and 13.2(μg/ml) in U937, K562, and HL-60 cell lines, respectively. The flow-cytometric analysis confirms that the cell cycle arrest occurs at G1 phase in case of U937 and K562 cell lines and G2/M phase in case of HL60 cell lines. Similarly both confocal microsocopic analysis and DNA laddering assay confirm the apoptosis and cell cycle arrests of leukemic cells. ConclusionThe overall results provide evidence for the ethnopharmacological relevance for use of the plant Wattakaka volubilis in developing novel agents for the treatment of leukemia.