Antihypertensive activity and vascular reactivity mechanisms of Vitex pubescens leaf extracts in spontaneously hypertensive rats.

BACKGROUND: Vitex pubescens has been used traditionally in hypertension treatment but not yet scientifically assessed. The objective of the study is to investigate the antihypertensive and vasorelaxant activities of V. pubescens, study its underlying pharmacological mechanisms, and identify the relevant vasoactive compounds. METHODS: Successive extractions of V. pubescens leaf were carried out to produce petroleum ether (VPPE), chloroform (VPCE), methanol (VPME), and water (VPWE) extracts. Spontaneously hypertensive rats (SHRs) received a daily oral administration of the extracts (500 mg/kg/day; n = 6) or verapamil (15 mg/kg/day; n = 6) for 2 weeks, while the systolic and diastolic blood pressures were measured using non-invasive tail-cuff method. Vasorelaxation assays of the extracts were later conducted using phenylephrine (PE, 1 µM) pre-contracted aortic ring preparation. Mechanisms of vasorelaxation by the most potent fraction were studied using vasorelaxation assays with selected blockers/inhibitors. GC-MS was conducted to determine the active compounds. RESULTS: VPPE elicited the most significant diminution in systolic and diastolic blood pressure of treated SHRs and produced the most significant vasorelaxation in the aortic rings. Vasorelaxant effects of F2-VPPE were significantly reduced in endothelium-denuded aortic rings by glibenclamide (1 µM), whereas calcium chloride and PE-induced contractions were significantly suppressed. Endothelium removal of the aortic rings or incubation with indomethacin (10 µM), atropine (1 µM), methylene blue (10 µM), propranolol (1µM) and L-NAME (10 µM) did not significantly alter F2-VPPE-induced vasorelaxation. Seven compounds were identified using GC-MS, including spathulenol. CONCLUSION: F2-VPPE exerted its endothelium-independent vasorelaxation by inhibition of vascular smooth muscle contraction induced by extracellular Ca+2 influx through trans-membrane Ca+2 channels and/or Ca+2 release from intracellular stores, and by activation of KATP channels. The vasorelaxation effects of V. pubescens could be mediated by the compound, spathulenol.

Blood pressure lowering effect and vascular activity of Phyllanthus niruri extract: The role of NO/cGMP signaling pathway and ß-adrenoceptor mediated relaxation of isolated aortic rings.

ETHNOPHARMACOLOGICAL RELEVANCE: Phyllanthus niruri have a long history of use in the traditional treatment of various ailments including hypertension. Literature reports have indicated that it is a potent antihypertensive herbal medication used traditionally. AIM OF THE STUDY: This study was carried out to investigate the antihypertensive and vasodilatory activity of four solvents extracts of P. niruri namely; petroleum ether (PEPN), chloroform (CLPN), methanol (MEPN) and water (WEPN), with the aim of elucidating the mechanism of action and identifying the phytochemical constituents. MATERIALS AND METHODS: Male Spontaneous Hypertensive Rats (SHRs) were given oral gavage of P. niruri extract daily for two weeks and the blood pressure was recorded in vivo. We also determine the vasodilation effect of the extracts on rings of isolated thoracic aorta pre-contracted with phenylephrine (PE, 1 µM). Endothelium-intact or endothelium-denuded aorta rings were pre-incubated with various antagonists like 1H-[1,2,4] oxadiazolo-[4,3-a]quinoxalin-1-one (ODQ, 10 µM) and Methylene blue (MB 10 µM), sGC inhibitors; Nω-Nitro-L-arginine methyl ester hydrochloride (L-NAME, 10 µM) a nitric oxide synthase (NOS) inhibitor; atropine (10 µM), a cholinergic receptor blocker; indomethacin (10 µM), a cyclooxygenase inhibitor and various K+ channel blockers such as glibenclamide (10 µM) and tetraethyl ammonium (TEA 10 µM) for mechanism study. RESULTS: SHRs receiving P. niruri extracts showed a significant decrease in their blood pressure (BP) when compared to the baseline value, with PEPN being more potent. The extracts (0.125-4 mg/mL) also induced vasorelaxation on endothelium-intact aorta rings. PEPN elicited the most potent maximum relaxation effect (Rmax). Mechanism assessment of PEPN showed that its relaxation effect is significantly suppressed in endothelium-denuded aorta rings. Pre-incubation of aorta rings with atropine, L-NAME, ODQ, indomethacin, and propranolol also significantly attenuated its relaxation effect. Conversely, incubation with TEA and glibenclamide did not show a significant effect on PEPN-induced relaxation. CONCLUSION: This study indicates that the antihypertensive activity of P. niruri extract is mediated by vasoactive phytoconstituents that dilate the arterial wall via endothelium-dependent pathways and ß-adrenoceptor activity which, in turn, cause vasorelaxation and reduce blood pressure.

Combination of Epicatechin 3-Gallate from Euphorbia hirta and Cefepime Promotes Potential Synergistic Eradication Action against Resistant Clinical Isolate of Pseudomonas aeruginosa.

Pseudomonas aeruginosa is naturally resistant to many classes of antipseudomonal antibiotics due to the species ability to easily acquire resistance. Plant-based antibacterial agent in combination with the existing antibiotic proposes an alternative treatment regimen for the eradication of resistant bacterial infections. The antibacterial effects of the isolated epicatechin 3-gallate compound from Euphorbia hirta in combination with cefepime were investigated in vitro against resistant P. aeruginosa. The fractional inhibitory concentration index of the combination was determined using checkerboard broth microdilution method. Epicatechin 3-gallate combined with cefepime had produced synergistic effect against P. aeruginosa (with average FIC index of 0.24). The MIC of epicatechin 3-gallate was effectively reduced to MIC/4, MIC/8, MIC/16, and MIC/32 in the presence of cefepime. Time-kill study of epicatechin 3-gallate combined with cefepime exhibited remarkable bactericidal activity where the eradication of P. aeruginosa occurred within 4 h of treatment. Scanning electron micrographs revealed apparent cell membrane damage and leakage of cytoplasmic contents from P. aeruginosa cells which eventually led to the cell lysis after the combination treatment of epicatechin 3-gallate and cefepime. The potential of epicatechin 3-gallate to act synergistically with cefepime against clinically resistant P. aeruginosa strain possibly will maximize the successful outcomes when choosing empirical antibiotic treatment in hospitals or health care institutions.

In vivo anti-arthritic and anti-nociceptive effects of ethanol extract of Moringa oleifera leaves on complete Freund's adjuvant (CFA)-induced arthritis in rats.

BACKGROUND: The medicinal uses of plants are in many cases based exclusively on traditional knowledge without enough scientific evidences. Different parts of Moringa oleifera were traditionally used for the treatment of wide variety of ailments including arthritis and joints pain. The present study had been designed to evaluate the anti-arthritic and anti-nociceptive activities of ethanol extract of Moringa leaves, this being the most abundant plant part suitable for commercial mass production of botanical medicinal products. METHODS: Complete Freund's adjuvant (CFA)-induced arthritis in rats was used as disease model. CFA-induced inflammatory paw edema, body weight, arthritic index, X-ray radiography, hematological parameters, and walk track and locomotion analysis were all evaluated for the assessment of disease progression. In addition to that, anti-nociceptive activity was examined at different dose levels in both normal and arthritic-induced rats using Eddy's hot plate and tail flick thermal analgesia. RESULTS: The analysis of various arthritic assessment parameters used in this study revealed that Moringa extract has a considerable effect in preventing development or ameliorate arthritis disease severity. Moreover, the ethanol extract of Moringa leaves revealed significant anti-nociceptive activity at in both normal and CFA-induced arthritis rats in a dose-dependent manner. CONCLUSION: Ethanol extract of Moringa leaves appears to be a really promising as analgesic and arthritis medication, but a larger and more detailed preclinical and clinical studies especially in human is highly recommended.

Mechanism of Action of Isolated Caffeic Acid and Epicatechin 3-gallate from Euphorbia hirta against Pseudomonas aeruginosa.

BACKGROUND: The escalating dominance of resistant Pseudomonas aeruginosa strains as infectious pathogen had urged the researchers to look for alternative and complementary drugs. OBJECTIVE: The objective of this study is to address the biological targets and probable mechanisms of action underlying the potent antibacterial effect of the isolated compounds from Euphorbia hirta (L.) against P. aeruginosa. MATERIALS AND METHODS: The action mechanisms of caffeic acid (CA) and epicatechin 3-gallate (ECG) on P. aeruginosa cells were investigated by several bacterial physiological manifestations involving outer membrane permeabilization, intracellular potassium ion efflux, and nucleotide leakage. RESULTS: The findings revealed that ECG and CA targeted both cell wall and cytoplasmic membrane of P. aeruginosa. The cellular membrane destruction and ensuing membrane permeability perturbation of P. aeruginosa had led to the ascending access of hydrophobic antibiotics, release of potassium ions, and leakages of nucleotides. CONCLUSION: The overall study concludes that ECG and CA isolated from E. hirta possess remarkable anti-infective potentials which can be exploited as drug template for the development of new antibacterial agent against resistant P. aeruginosa pathogen. SUMMARY: Epicatechin 3-gallate (ECG) and caffeic acid (CA) exhibited remarkable bactericidal abilities by increasing the outer membrane and plasma membrane permeability of Pseudomonas aeruginosa pathogenECG and CA had facilitated the entry of hydrophobic antibiotics into P. aeruginosa by disintegrating the lipopolysaccharides layer of the outer membraneECG-induced potassium efflux with efficiency close to that obtained with cefepime suggesting mode of action through membrane disruptionBoth ECG and CA had caused consistent leakage of intracellular nucleotide content with the increase in time. Abbreviations used: ECG: Epicatechin 3-gallate; CA: Caffeic acid; E. hirta: Euphoria hirta.

Kigelia africana (Lam.) Benth. (Sausage tree): Phytochemistry and pharmacological review of a quintessential African traditional medicinal plant.

ETHNOPHARMACOLOGICAL RELEVANCE: Kigelia africana is a quintessential African herbal medicinal plant with a pan-African distribution and immense indigenous medicinal and non-medicinal applications. The plant is use traditionally as a remedy for numerous disease such as use wounds healing, rheumatism, psoriasis, diarrhea and stomach ailments. It is also use as an aphrodisiac and for skin care. AIM OF THE REVIEW: The present review aims to compile an up-to-date review of the progress made in the continuous pharmacological and phytochemistry investigation of K. africana and the corresponding commercial and pharmaceutical application of these findings with the ultimate objective of providing a guide for future research on this plant. METHOD: The scholarly information needed for this paper were predominantly sourced from the electronic search engines such as Google, Google scholar; publishing sites such as Elsevier, scienceDirect, BMC, PubMed; other scientific database sites for chemicals such as ChemSpider, PubChem, and also from online books. RESULTS: Pharmacological investigations conducted confirm the anti-inflammatory, analgesic, antioxidant and anticancer activity of the extract of different parts of the plant. Bioactive constituents are found to be present in all parts of the plant. So far, approximately 150 compounds have been characterized from different part of the plant. Iridoids, naphthoquinones, flavonoids, terpenes and phenylethanoglycosides are the major class of compounds isolated. Novel compounds with potent antioxidant, antimicrobial and anticancer effect such as verbascoside, verminoside and pinnatal among others, have been identified. Commercial trade of K. africana has boosted in the las few decades. Its effect in the maintenance of skin has been recognized resulting in a handful of skin formulations in the market. CONCLUSIONS: The pharmaceutical potentials of K. africana has been recognized and have witness a surge in research interest. However, till date, many of its traditional medicinal uses has not been investigated scientifically. Further probing of the existential researches on its pharmacological activity is recommended with the end-goal of unravelling the pharmacodynamics, pharmacokinetics, clinical relevance and possible toxicity and side effects of both the extract and the active ingredients isolated.

Antioxidant and Cytotoxic Effect of Barringtonia racemosa and Hibiscus sabdariffa Fruit Extracts in MCF-7 Human Breast Cancer Cell Line.

BACKGROUND: The fruits of Barringtonia racemosa and Hibiscus sabdariffa have been used in the treatment of abscess, ulcer, cough, asthma, and diarrhea as traditional remedy. OBJECTIVE: This study aims to evaluate cytotoxic effect of B. racemosa and H. sabdariffa methanol fruit extracts toward human breast cancer cell lines (MCF-7) and its antioxidant activities. MATERIALS AND METHODS: Total antioxidant activities of extracts were assayed using 2,2'-diphenyl-1-picrylhydrazyl radical (DPPH) and ß-carotene bleaching assay. Content of phytochemicals, total flavonoid content (TFC), and total phenolic content (TPC) were determined using aluminum chloride colorimetric method and Folin-Ciocalteu's reagent, respectively. Cytotoxic activity in vitro was investigated through 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyl tetrazolium bromide (MTT) assay. RESULTS: B. racemosa extract exhibited high antioxidant activities compared to H. sabdariffa methanol fruit extracts in DPPH radical scavenging assay (inhibitory concentration [IC50] 15.26 ± 1.25 µg/mL) and ί-carotene bleaching assay (I% 98.13 ± 1.83%). B. racemosa also showed higher TPC (14.70 ± 1.05 mg gallic acid equivalents [GAE]/g) and TFC (130 ± 1.18 mg quercetin equivalents [QE]/g) compared to H. sabdariffa (3.80 ± 2.13 mg GAE/g and 40.75 ± 1.15 mg QE/g, respectively). In MTT assay, B. racemosa extract also showed a higher cytotoxic activity (IC50 57.61 ± 2.24 µg/mL) compared to H. sabdariffa. CONCLUSION: The present study indicated that phenolic and flavonoid compounds known for oxidizing activities indicated an important role among the contents of these plants extract. B. racemosa methanol extract have shown potent cytotoxic activity toward MCF-7. Following these promising results, further fractionation of the plant extract is underway to identify important phytochemical bioactives for the development of potential nutraceutical and pharmaceutical use. SUMMARY: The phenolic and flavonoid compounds were present in B. racemosa and H. sabdariffa methanol extractsB. racemosa methanol extract was found to be potent antioxidant activityB. racemosa methanol extract have shown potent cytotoxic activity (IC50 57.61 ± 2.24 µg/mL) toward MCF-7The phenolic and flavonoid compounds may contribute to the antioxidant and cytotoxic activity of B. racemosa. Abbreviations Used: MCF-7: Human breast cancer cell lines, DMEM: Modified eagle medium, DPPH: 2,2'-diphenyl-1-picrylhydrazyl radical, TPC: Total phenolic content, Na2CO3: Sodium carbonate, GAE: Gallic acid equivalents, TFC: Total flavonoid content, NaNO2: Sodium nitrite, AlCl3: Aluminum chloride, NaOH: Sodium hydroxide, QE: Quercetin equivalents, MTT: 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyl tetrazolium bromide, IC50: Inhibitory concentration, ANOVA: Analysis of variance, DLA: Dalton's lymphoma ascitic.

Acute and Sub-Acute Toxicity Evaluation of the Methanolic Extract of Alstonia scholaris Stem Bark.

Alstonia scholaris has been used by traditional medicine practitioners since the medieval ages for the treatment of diseases. The aim of this research was to evaluate the acute and sub-acute oral toxicity of its methanolic extract. The acute toxicity test was conducted using Sprague Dawley (SD) rats. The methanolic extract of Alstonia scholaris stem bark (ASME) was administrated in a single dose of 2000 mg/kg via oral gavage; and the animals were observed for any behavioral changes or mortality. In the sub-acute toxicity study, SD rats received three doses of ASME (250, 500 and 1000 mg/kg) for 28 days via oral gavage. During these 28 days of treatment, the rats were observed weekly for toxicity symptoms. Following the 28-day treatment, the rats were sacrificed for hematological, biochemical and histopathology studies. In the acute toxicity study, Alstonia scholaris was found to be non-toxic at a dose of 2000 mg/kg b.w. In the sub-acute toxicity study, significant variations in body weight, hematological and biochemical parameters were observed in the experimental groups at the dose of 500 and 1000 mg/kg with the death of two female rats being recorded at the highest dose (1000 mg/kg b.w.). Histopathological studies revealed slight degeneration (lesion) and centrilobular necrosis in the liver, which was most expressed in the highest-dose group. These results demonstrate that, while a single dose and short term oral intake of Alstonia scholaris bark extract caused no toxicity up to a dose of 2000 mg/kg b.w., toxic effects manifested in the long term treatment at the highest dose (500 and 1000 mg/kg). The long-term toxic effect was found to be associated with alterations in hematological compositions and end-organ damage to the liver. Thus, prolonged use of high doses of ASME orally should be discouraged and lower doses encouraged.

Mechanisms underlying the antihypertensive effect of Alstonia scholaris.

ETHNOPHARMACOLOGICAL RELEVANCE: Alstonia scholaris has a long history of use in the Ayurveda traditional treatment of various ailments including hypertension. We have reported the blood pressure lowering activity of the extract of A. scholaris. The following research aim to delineate the pharmacological mechanism involve in the antihypertensive action. MATERIALS AND METHOD: Vasorelaxant effect of the n-butanol fraction of A. scholaris (NBF-ASME) was evaluated on rat aorta pre-contracted with phenyelphrine (PE, 1 µM). Aortic rings preparation were pre-incubated with various antagonists like 1H-[1,2,4] oxadiazolo-[4,3-a]quinoxalin-1-one (ODQ 10 µM), methylene blue (MB 10 µM), Nω-nitro-L-arginine methyl ester hydrochloride (l-NAME 10 µM), atropine (10 µM), indomethacin (1 µM), ML-9 and various K(+) channel blockers such as glibenclamide (10 µM) and tetraethyl ammonium (TEA 10 µM) for mechanism study. RESULT: The results showed that pre-incubation of aortic rings with the extract (0.5, 1 and 2mg/mL) significantly inhibit the contractile response of the rings to phenylephrine-induced contraction (p<0.05-0.001). Removal of endothelium, incubation with L-NAME, indomethacin, atropine and propranolol did not significantly affect the relaxation effect of NBF-ASME. Furthermore, the K(+) channel blockers, TEA and glibenclamide showed no inhibitory effect. However, aortic rings pretreated with ODQ and ML-9 showed a significant suppression of the relaxation curve of NBF-ASME (p<0.01-0.001). In Ca(2+)-free solution, NBF-ASME inhibits the release of intracellular Ca(2+) from the sarcoplasmic reticulum. NBF-ASME also inhibits calcium chloride (CaCl2)-induced contraction in endothelium-denuded aortic rings. CONCLUSION: The results from this study suggests that A. scholaris exerts vasodilation via calcium channels blockade, direct activation of soluble guanylate cyclase and possibly by also inhibiting the formation of inositol 1, 4, 5-triphosphate.

Antidiabetic and antioxidant activities of Nypa fruticans Wurmb. vinegar sample from Malaysia.

OBJECTIVES: To study the antidiabetic and antioxidant activities of nipa palm vinegar (NPV) used in traditional Malay medicine for treating diabetes. METHODS: NPV was extracted using liquid-liquid extraction method and the obtained samples were subjected to antidiabetic studies using normal and streptozotocin-induced diabetic rat models whereas antidoxidant activities were investigated via in vitro antioxidant tests namely 2,2-diphenyl-1-picrylhydrazyl and 2,2'-azinobis-3-ethylbenzothiozoline-6-sulfonic acid free radicals scavenging activities and the reducing power assay. RESULTS: Single administration of NPV and its extracts were not effective in both normal and diabetic rats. In intraperitoneal glucose tolerance test, NPV and its aqueous extract showed significant blood glucose lowering effect. In the sub-acute study, compared with the diabetic control, aqueous extract of NPV showed the most notable blood glucose lowering effect (56.6%) and a significant improvement in serum insulin levels (79.8%, P < 0.05). To assess NPV's antioxidant activity, three in vitro antioxidant tests were employed: 2,2-diphenyl-1-picrylhydrazyl and 2,2'-azinobis-3-ethylbenzothiozoline-6-sulfonic acid free radical-scavenging assays, and the reducing power assay. Ethyl acetate extract had the greatest antioxidant potential and content of phenolic and flavonoid compounds. A linear positive correlation between the antioxidant parameters was observed. Chemical profiling analysis of aqueous extract of NPV revealed the presence of acetic acid (35.25%), the main active constituent which significantly contributed to the observed antidiabetic activity. CONCLUSIONS: Aqueous extract of NPV possesses antihyperglycaemic activities comparable to the metformin, while the ethyl acetate extract precipitated significant antioxidant effects attributable to its high phenolic content. These findings suggest that antioxidant compounds of NPV do not contribute much towards the overall observed antidiabetic effect.

Aqueous Extract of Nypa fruticans Wurmb. Vinegar Alleviates Postprandial Hyperglycemia in Normoglycemic Rats.

Nypa fruticans Wurmb. vinegar, commonly known as nipa palm vinegar (NPV) has been used as a folklore medicine among the Malay community to treat diabetes. Early work has shown that aqueous extract (AE) of NPV exerts a potent antihyperglycemic effect. Thus, this study is conducted to evaluate the effect of AE on postprandial hyperglycemia in an attempt to understand its mechanism of antidiabetic action. AE were tested via in vitro intestinal glucose absorption, in vivo carbohydrate tolerance tests and spectrophotometric enzyme inhibition assays. One mg/mL of AE showed a comparable outcome to the use of phloridzin (1 mM) in vitro as it delayed glucose absorption through isolated rat jejunum more effectively than acarbose (1 mg/mL). Further in vivo confirmatory tests showed AE (500 mg/kg) to cause a significant suppression in postprandial hyperglycemia 30 min following respective glucose (2 g/kg), sucrose (4 g/kg) and starch (3 g/kg) loadings in normal rats, compared to the control group. Conversely, in spectrophotometric enzymatic assays, AE showed rather a weak inhibitory activity against both α-glucosidase and α-amylase when compared with acarbose. The findings suggested that NPV exerts its anti-diabetic effect by delaying carbohydrate absorption from the small intestine through selective inhibition of intestinal glucose transporters, therefore suppressing postprandial hyperglycemia.

Chemical analysis, inhibition of biofilm formation and biofilm eradication potential of Euphorbia hirta L. against clinical isolates and standard strains.

BACKGROUND: The frequent occurrences of antibiotic-resistant biofilm forming pathogens have become global issue since various measures that had been taken to curb the situation led to failure. Euphorbia hirta, is a well-known ethnomedicinal plant of Malaysia with diverse biological activities. This plant has been used widely in traditional medicine for the treatment of gastrointestinal, bronchial and respiratory ailments caused by infectious agents. METHODS: In the present study, chemical compositions of methanol extract of E. hirta L. aerial part was analyzed by gas chromatography and gas chromatography coupled to mass spectrometry. A relevant in vitro model was developed to assess the potency of the E. hirta extract to inhibit the bacterial biofilm formation as well as to eradicate the established biofilms. Besides biofilm, E. hirta extract was also evaluated for the inhibition efficacy on planktonic cells using tetrazolium microplate assay. For these purposes, a panel of clinically resistant pathogens and American type culture collection (ATCC) strains were used. RESULTS: The methanolic extract of aerial part of E. hirta was predominantly composed of terpenoid (60.5%) which is often regarded as an active entity accountable for the membrane destruction and biofilm cell detachment. The highest antibacterial effect of crude E. hirta extract was observed in the clinical isolates of Pseudomonas aeruginosa with minimum inhibitory concentration (MIC) value of 0.062 mg/ml. The extract also displayed potent biofilm inhibition and eradication activity against P. aeruginosa with minimum biofilm inhibition concentration (MBIC) and minimum biofilm eradication concentration (MBEC) values of 0.25 mg/ml and 0.5 mg/ml, respectively. CONCLUSIONS: The crude methanol extract of E. hirta has proven to have interesting and potential anti-biofilm properties. The findings from this study will also help to establish a very promising anti-infective phytotherapeutical to be exploited in the pharmaceutical industries.

In vitro and in vivo effects of standardized extract and fractions of Phaleria macrocarpa fruits pericarp on lead carbohydrate digesting enzymes.

BACKGROUND: One vital therapeutic approach for the treatment of type 2 diabetes mellitus is the use of agents that can decrease postprandial hyperglycaemia by inhibiting carbohydrate digesting enzymes. The present study investigated the effects of bioassay-guided extract and fractions of the dried fruit pericarp of Phaleria macrocarpa, a traditional anti-diabetic plant, on α-glucosidase and α-amylase, in a bid to understand their anti-diabetic mechanism, as well as their possible attenuation action on postprandial glucose increase. METHODS: Methanol extract (ME), obtained by successive solvent extraction, its most effective liquid-liquid n-butanol fraction (NBF) and the flash column chromatographic sub-fraction (SFI), were evaluated for in vitro α-glucosidase (yeast) and α-amylase (porcine) activity inhibition. Furthermore, confirmatory in vivo tests were carried out in streptozotocin-induced diabetic rats (SDRs) using oral glucose, sucrose and starch tolerance tests. RESULTS: At the highest concentration employed (100 µg/ml), NBF showed highest inhibition against α-glucosidase (75%) and α-amylase (87%) in vitro (IC50 = 2.40 ± 0.23 µg/ml and 58.50 ± 0.13 µg/ml, respectively) in a dose-dependent fashion; an effect found to be about 20% higher than acarbose (55%), a standard α-glucosidase inhibitor (IC50 = 3.45 ± 0.19 µg/ml). The ME and SFI also inhibited α-glucosidase (IC50 = 7.50 ± 0.15 µg/ml and 11.45 ± 0.28 µg/ml) and α-amylase (IC50 = 43.90 ± 0.19 µg/ml and 69.80 ± 0.25 µg/ml), but to a lesser extent. In in vivo studies with diabetic rats, NBF and SFI effectively reduced peak blood glucose (PBG) by 15.08% and 6.46%, and the area under the tolerance curve (AUC) by 14.23% and 12.46%, respectively, after an oral sucrose challenge (P < 0.05); thereby validating the observed in vitro action. These reduction effects on PBG and AUC were also demonstrated in glucose and starch tolerance tests, but to a lesser degree. CONCLUSIONS: These findings reveal that P. macrocarpa can attenuate hyperglycaemia in both in vitro and in vivo conditions by potently inhibiting carbohydrate hydrolysing enzymes, making it a viable plant for sourcing natural compounds for the management of type 2 diabetes mellitus.

A simple isocratic HPLC method for the simultaneous determination of sinensetin, eupatorin, and 3'-hydroxy-5,6,7,4'-tetramethoxyflavone in Orthosiphon stamineus extracts.

Orthosiphon stamineus extracts contain three flavonoids (3'-hydroxy-5,6,7,4'-tetramethoxyflavone, sinensetin, and eupatorin) as bioactive substances. Previous reported high performance liquid chromatography- ultraviolet (HPLC-UV) methods for the determination of these flavonoids have several disadvantages, including unsatisfactory separation times and not being well validated according to International Conference on Harmonisation of Technical Requirements for Registration of Pharmaceuticals for Human Use (ICH) standard guidelines. A rapid, specific reversed-phase HPLC method with isocratic elution of acetonitrile: isopropyl alcohol: 20mM phosphate buffer (NaH(2)PO(4)) (30:15:55, v/v) (pH 3.5) at a flow-rate of 1ml/minute, a column temperature of 25°C, and ultraviolet (UV) detection at 340 nm was developed. The method was validated and applied for quantification of different types of O stamineus extracts and fractions. The method allowed simultaneous determination of 3'-hydroxy-5,6,7,4'-tetramethoxyflavone, sinensetin, and eupatorin in the concentration range of 0.03052-250 µg/ml. The limits of detection and quantification, respectively, were 0.0076 and 0.061 µg/ml for 3'-hydroxy-5,6,7,4'-tetramethoxyflavone, 0.0153 and 0.122 µg/ml for sinensetin and 0.0305 and 0.122 µg/ml for eupatorin. The percent relative standard deviation (% RSD) values for intraday were 0.048-0.368, 0.025-0.135, and 0.05-0.476 for 3'-hydroxy-5,6,7,4'-tetramethoxyflavone, sinensetin, and eupatorin, respectively, and those for intraday precision were 0.333-1.688, 0.722-1.055, and 0.548-1.819, respectively. The accuracy for intraday were 91.25%-103.38%, 94.32%-109.56%, and 92.85%-109.70% for 3'-hydroxy-5,6,7,4'-tetramethoxyflavone, sinensetin, and eupatorin, respectively, and those for interday accuracy were 97.49%-103.92%, 103.58%-104.57%, and 103.9%-105.33%, respectively. The method was found to be simple, accurate and precise and is recommended for routine quality control analysis of O stamineus extract containing the three flavonoids as the principle components in the extract.

Bioassay-guided antidiabetic study of Phaleria macrocarpa fruit extract.

An earlier anti-hyperglycemic study with serial crude extracts of Phaleria macrocarpa (PM) fruit indicated methanol extract (ME) as the most effective. In the present investigation, the methanol extract was further fractionated to obtain chloroform (CF), ethyl acetate (EAF), n-butanol (NBF) and aqueous (AF) fractions, which were tested for antidiabetic activity. The NBF reduced blood glucose (p < 0.05) 15 min after administration, in an intraperitoneal glucose tolerance test (IPGTT) similar to metformin. Moreover, it lowered blood glucose in diabetic rats by 66.67% (p < 0.05), similar to metformin (51.11%), glibenclamide (66.67%) and insulin (71.43%) after a 12-day treatment, hence considered to be the most active fraction. Further fractionation of NBF yielded sub-fractions I (SFI) and II (SFII), and only SFI lowered blood glucose (p < 0.05), in IPGTT similar to glibenclamide. The ME, NBF, and SFI correspondingly lowered plasma insulin (p < 0.05) and dose-dependently inhibited glucose transport across isolated rat jejunum implying an extra-pancreatic mechanism. Phytochemical screening showed the presence of flavonoids, terpenes and tannins, in ME, NBF and SFI, and LC-MS analyses revealed 9.52%, 33.30% and 22.50% mangiferin respectively. PM fruit possesses anti-hyperglycemic effect, exerted probably through extra-pancreatic action. Magniferin, contained therein may be responsible for this reported activity.

Antioxidant and antiangiogenic activities of the essential oils of Myristica fragrans and Morinda citrifolia.

OBJECTIVE: Toinvestigate the anti-angiogenic activity and antioxidant properties of Myristica fragrans (M. fragrans) (nutmeg) and Morinda citrifolia (M. citrifolia)(mengkudu) oils. METHODS: The nutmeg and megkudu essential oils were obtained by steam distillation. The antioxidant activities of both essential oils were determined by beta-carotene/linoleic acid bleaching assay and reducing power while the anti-angiogenic activity was investigated using rat aortic ring assay using various concentrations. RESULTS: The results showed that nutmeg oil has higher antioxidant activity than mengkudu oil. The nutmeg oil effectively inhibited the oxidation of linoleic acid with (88.68±0.1)% while the inhibition percentage of oxidation of linoleic acid of the mengkudu oil is (69.44±0.4)%. The nutmeg oil and mengkudu oil showed reducing power with an EC(50) value of 181.4 µg/mL and 3 043.0 µg/mL, respectively. The antiangiogenic activity of nutmeg oil showed significant antiangiogenic activity with IC(50) of 77.64 µg/mL comparing to mengkudu oil which exhibits IC(50) of 109.30 µg/mL. CONCLUSIONS: Bioactive compound(s) will be isolated from the nutmeg essential oil to be developed as antiangiogenic drugs.

Chemical composition, antioxidant and cytotoxicity activities of the essential oils of Myristica fragrans and Morinda citrifolia.

BACKGROUND: In this study the chemical composition, antioxidant activities and cytotoxic effect of the essential oils of Myristica fragrans (nutmeg) and Morinda citrifolia (mengkudu) were determined. RESULTS: Thirty-eight compounds in nutmeg oil and six compounds in mengkudu oil were identified by gas chromatography­mass spectrometry. The free radical scavenging activity of nutmeg oil was superior of that mengkudu oil. The MTT assay of nutmeg oil on human colorectal carcinoma (HCT-116) and human breast carcinoma (MCF-7) cell lines showed IC50 values of 78.61 and 66.45 µg mL⁻¹, respectively. The mengkudu oil exhibited IC50 values of 91.46 and 78.15 µg mL⁻¹ for HCT-116 and MCF-7, respectively. CONCLUSION: The results showed that nutmeg oil can be developed as potent anti-cancer and antioxidant drugs.