Mutagenicity and preclinical safety assessment of the aqueous extract of Clinacanthus nutans leaves.

CONTEXT: Clinacanthus nutans (CN) is used traditionally for treating various illnesses. Robust safety data to support its use is lacking. OBJECTIVE: To evaluate the adverse effects of aqueous extract of CN leaves (AECNL). MATERIALS AND METHODS: The oral toxicity of the AECNL was tested following Organisation for Economic Co-operation and Development (OECD) guidelines. Mutagenicity (Ames test) of AECNL was evaluated using TA98 and TA100 Salmonella typhimurium strains. RESULTS: No mortality or morbidity was found in the animals upon single and repeated dose administration. However, significant body weight loss was observed at 2000 mg/kg during sub-chronic (90 d) exposure. In addition, increased eosinophil at 500 mg/kg and decreased serum alkaline phosphatase levels at 2000 mg/kg were observed in male rats. Variations in glucose and lipid profiles in treated groups were also observed compared to control. Ames test revealed no evidence of mutagenic or carcinogenic effects at 500 µg/well of AECNL. CONCLUSION: The median lethal dose (LD50) of the AECNL is >5000 mg/kg and the no-observed-adverse-effect level is identified to be greater than 2000 mg/kg/day in 90-d study.

Flavonoids-Rich Orthosiphon stamineus Extract as New Candidate for Angiotensin I-Converting Enzyme Inhibition: A Molecular Docking Study.

This study aims to evaluate the in vitro angiotensin-converting enzyme (ACE) inhibition activity of different extracts of Orthosiphon stamineus (OS) leaves and their main flavonoids, namely rosmarinic acid (RA), sinensetin (SIN), eupatorin (EUP) and 3'-hydroxy-5,6,7,4'-tetramethoxyflavone (TMF). Furthermore, to identify possible mechanisms of action based on structure-activity relationships and molecular docking. The in vitro ACE inhibition activity relied on determining hippuric acid (HA) formation from ACE-specific substrate (hippuryl-histidyl-leucine (HHL)) by the action of ACE enzyme. A High Performance Liquid Chromatography method combined with UV detection was developed and validated for measurement the concentration of produced HA. The chelation ability of OS extract and its reference compounds was evaluated by tetramethylmurexide reagent. Furthermore, molecular docking study was performed by LeadIT-FlexX: BioSolveIT's LeadIT program. OS ethanolic extract (OS-E) exhibited highest inhibition and lowest IC50 value (45.77 ± 1.17 µg/mL) against ACE compared to the other extracts. Among the tested reference compounds, EUP with IC50 15.35 ± 4.49 µg/mL had highest inhibition against ACE and binding ability with Zn (II) (56.03% ± 1.26%) compared to RA, TMF and SIN. Molecular docking studies also confirmed that flavonoids inhibit ACE via interaction with the zinc ion and this interaction is stabilized by other interactions with amino acids in the active site. In this study, we have demonstrated that changes in flavonoids active core affect their capacity to inhibit ACE. Moreover, we showed that ACE inhibition activity of flavonoids compounds is directly related to their ability to bind with zinc ion in the active site of ACE enzyme. It was also revealed that OS extract contained high amount of flavonoids other than RA, TMF, SIN and EUP. As such, application of OS extract is useful as inhibitors of ACE.

Genotoxicity, acute and subchronic toxicity studies of nano liposomes of Orthosiphon stamineus ethanolic extract in Sprague Dawley rats.

BACKGROUND: Orthosiphon stamineus (OS) Benth is a medicinal plant and native in Southeast Asia. Pharmacological effects of OS are attributed to the presence of lipophilic flavones. However; lipophilic compounds suffer from poor aqueous solubility which limits the OS oral bioavailability and therapeutic applications. Therefore, OS was prepared in nano formulation form using liposomes from soybean phospholipids. The aim of the present study is to evaluate the in vitro genotoxicity and in vivo oral toxicity of nano liposomes of OS ethanolic extract (OS-EL). METHODS: In the acute toxicity study Sprague Dawley female rats were given a single dose of the OS-EL at 5000 mg/kg/day orally and screened for two weeks after administration. In the subchronic study, three different doses of OS-EL were administered for 28 days. Mortality, clinical signs, body weight changes, hematological and biochemical parameters, gross findings, organ weights, and histological parameters were monitored during the study. Genotoxicity was assessed using the Ames test with the TA98 and TA100 Salmonella typhimurium strains. High-performance liquid chromatography was performed for identification and quantification of the major marker compounds in OS-EL. Heavy metal detection was performed using an atomic absorption spectrometer. RESULTS: The acute toxicity study showed that the LD50 of the extract was greater than 5000 mg/kg. In the repeated dose 28-day oral toxicity study, the administration of 250 mg/kg, 500 mg/kg, and 1000 mg/kg/day of OS-EL per body weight revealed no significant difference in food and water consumptions, bodyweight change, haematological and biochemical parameters, relative organ weights, gross findings or histopathology compared to the control group. The Ames test revealed that the OS-EL did not have any potential to induce gene mutations in S. Typhimurium. CONCLUSIONS: Analyses of these results with the information of signs, behaviour, and health monitoring could lead to the conclusion that the long-term oral administration of OS-EL for 28 days does not cause sub-chronic toxicity.

Selected metabolites profiling of Orthosiphon stamineus Benth leaves extracts combined with chemometrics analysis and correlation with biological activities.

BACKGROUND: Studies on selected metabolites profiling of Orthosiphon stamineus extracts using chromatographic and spectroscopic techniques combined with chemometric tools have not been fully elucidated. Thus present study was performed to profile selected metabolites in O. stamineus leaves extracts using HPLC and FTIR combined with chemometric tools and correlated with biological activities. METHODS: Five different extracts were prepared using three methods; maceration, soxhlet and reflux. The extracts were analyzed using UV-Vis, HPLC and FTIR techniques. Analysis of selected primary and secondary metabolites was also evaluated. The antioxidant and cytotoxic activities of the extracts were evaluated. Chemometric tools were employed to classify the extracts based on HPLC analysis and FTIR fingerprints. RESULTS: The ethanolic extract using maceration characterized high content of phenolics and flavonoids, (rosmarinic acid and eupatorin) with high antioxidant activity. Ethanolic (50%) and methanolic extracts using soxhlet showed high proteins and glycosaponins. Water extracts using reflux and maceration showed high polysaccharides. Methanolic extract (50%) using soxhlet and methanolic extract using maceration showed strong cytotoxic effect against MCF7 and HCT116 cell lines, respectively. Antioxidant and cytotoxic activities showed significant correlation with selected primary and secondary metabolites. HPLC fingerprints combined with chemometrics showed the extracts have been clustered based on selected major peaks profile. FTIR fingerprints combined with chemometrics showed that the extracts have been clustered based on protein and polysaccharide contents. CONCLUSION: Ten different extracts of O. stamineus have showed significant differences in the content of selected primary and secondary metabolites as well as the biological activities. Chemometric tools were able to classify and discriminate the distinctive features of extracts thus can be correlated with the biological activities.

Isolation, Characterization, Crystal Structure Elucidation of Two Flavanones and Simultaneous RP-HPLC Determination of Five Major Compounds from Syzygium campanulatum Korth.

Two flavanones named (2S)-7-Hydroxy-5-methoxy-6,8-dimethyl flavanone (1), (S)-5,7-dihydroxy-6,8-dimethyl-flavanone (2), along with known chalcone, namely, (E)-2',4'- dihydroxy-6'-methoxy-3',5'-dimethylchalcone (3) and two triterpenoids, namely, betulinic and ursolic acids (4 and 5), were isolated from the leaves of Syzygium campanulatum Korth (Myrtaceae). The structures of compounds (1 and 2) were determined on the basis of UV-visible, FTIR, NMR spectroscopies and LC-EIMS analytical techniques. Furthermore, new, simple, precise, selective, accurate, highly sensitive, efficient and reproducible RP-HPLC method was developed and validated for the quantitative analysis of the compounds (1-5) from S. campanulatum plants of five different age. RP-HPLC method was validated in terms of specificity, linearity (r2 ≤ 0.999), precision (2.0% RSD), and recoveries (94.4%-105%). The LOD and LOQ of these compounds ranged from 0.13-0.38 and 0.10-2.23 µg·mL-1, OPEN ACCESS respectively. Anti-proliferative activity of isolated flavanones (1 and 2) and standardized extract of S. campanulatum was evaluated on human colon cancer (HCT 116) cell line. Compounds (1 and 2) and extract revealed potent and dose-dependent activity with IC50 67.6, 132.9 and 93.4 µg·mL-1, respectively. To the best of our knowledge, this is the first study on isolation, characterization, X-ray crystallographic analysis of compounds (1 and 2) and simultaneous RP-HPLC determination of five major compounds (1-5) from different age of S. campanulatum plants.

Anti-Obesity Effects of Melastoma malabathricum var Alba Linn in Rats Fed with a High-Fat Diet.

Obesity is one of the major public health problems worldwide and it is generally associated with many diseases. Although synthetic drugs are available for the treatment of obesity, herbal remedies may provide safe, natural, and cost-effective alternative to synthetic drugs. One example of such drugs is Melastoma malabathricum var Alba Linn (MM). Although several studies have been reported for the pharmacological activities of MM, there is no report on the anti-obesity effect of MM. The aim of the present study is to evaluate the anti-obesity potential of methanolic extract of MM. The anti-obesity effect of MM on rats fed with a high-fat diet was investigated through determination of the changes in body weight, fat weight, organ weights, and blood biochemicals. The animals in this study were divided into three groups: a normal group with a standard diet (N), a control group fed with high-fat diet (C), and a MM treatment group fed with high-fat (HFD + MM) diet for 8 weeks. There was no significant difference in the amount of food intake between control and HFD + MM treatments. These results also suggest that MM does not induce a dislike for the diet due to its smell or taste. The study shows that MM significantly prevented increases in body weight, cholesterol, LDL, HDL, and total lipids that resulted from the high-fat diet. MM also decreased the epididymal fat (E-fat) and retroperitoneal fat (R-fat) weights and phospholipid concentrations induced by the high-fat diet. On the basis of these findings, it was concluded that MM had anti-obesity effects by suppressing body weight gain and abdominal fat formation.

Preparation and characterization of nano liposomes of Orthosiphon stamineus ethanolic extract in soybean phospholipids.

BACKGROUND: O. stamineus is a medicinal herb with remarkable pharmacological properties. However, poor solubility of the active principles limits its medicinal value. This study sought to prepare nano liposomes of OS ethanolic extract in unpurified soybean phospholipids in order to improve its solubility and permeability. OS liposomes were prepared by the conventional film method, and were characterized for solubility, entrapment efficiency, Fourier transform infrared spectroscopy (FTIR), transmission electron microscopy (TEM), particle size and zeta potential, release, absorption in everted rat intestinal sacs, and DPPH scavenging effect. RESULTS: OS liposomes showed substantial enhancement of extract's solubility from 956 ± 34 to 3979 ± 139 µg/ml, with entrapment efficiency of 66.2 ± 0.9%. FTIR study indicates interaction between soybean phospholipids and OS extract. TEM and dynamic light scattering showed presence of round anionic nano liposomes with particle size and zeta potential of 152.5 ± 1.1 nm and -49.8 ± 1.0 mV, respectively. A study using the fluorescent probe pyrene showed the critical micellar concentration is 9.2 ± 2.9 µg/ml. Release studies showed 94 ± 0.1% release in non-formulated extract and 62.4 ± 0.1% in OS liposomes. Released extract from OS liposomes showed improvement in DPPH scavenging effect, IC50 = 23.5 ± 1.1 µg/ml compared to 32.4 ± 0.5 µg/ml in non-formulated extract. OS liposomes were stable at pH 5.5 and 7.4, but showed reversible agglomeration at pH 1.6. Absorption in everted rat intestinal sacs showed substantial improvement in permeability of 3'-hydroxy-5, 6, 7, 4″-tetramethoxyflavone, sinensetin, eupatorin, and 3 other unknown compounds. CONCLUSIONS: Enhanced solubility, absorption and antioxidant effect may improve the overall pharmacological effects and medicinal value of OS ethanolic extract.

Syzygium campanulatum korth methanolic extract inhibits angiogenesis and tumor growth in nude mice.

BACKGROUND: Syzygium campanulatum Korth (Myrtaceae) is an evergreen shrub rich in phenolics, flavonoid antioxidants, and betulinic acid. This study sought to investigate antiangiogenic and anti-colon cancer effects of S.C. standardized methanolic extract. METHODS: Betulinic acid was isolated from methanolic extract by crystallization and chromatography techniques. S.C. methanolic extract was analyzed by UV-Vis spectrophotometry, FTIR, LC-MS, and HPLC. Antiangiogenic effect was studied on rat aortic rings, matrigel tube formation, cell proliferation and migration, and expression of vascular endothelial growth factor (VEGF). Antitumor effect was studied using a subcutaneous tumor model of HCT 116 colorectal carcinoma cells established in nude mice. RESULTS: Analysis by HPLC, LC-MS and FTIR confirm presence of betulinic acid in S.C. methanolic extract. Quantitative analysis by HPLC indicates presence of betulinic acid in S.C. extract at 5.42 ± 0.09% (w/w). Antiangiogenesis study showed potent inhibition of microvessels outgrowth in rat aortic rings, and studies on normal and cancer cells did not show any significant cytotoxic effect. Antiangiogenic effect was further confirmed by inhibition of tube formation on matrigel matrix that involves human endothelial cells (IC50 = 17.6 ± 2.9 µg/ml). S.C. extract also inhibited migration of endothelial cells and suppressed expression of VEGF. In vivo antiangiogenic study showed inhibition of new blood vessels in chicken embryo chorioallantoic membrane (CAM), and in vivo antitumor study showed significant inhibition of tumor growth due to reduction of intratumor blood vessels and induction of cell death. CONCLUSION: Collectively, our results indicate S. campanulatum as antiangiogenic and antitumor candidate, and a new source of betulinic acid.

Simultaneous quantification of flavonoids in blood plasma by a high-performance liquid chromatography method after oral administration of Blumea balsamifera leaf extracts in rats.

The leaves of Blumea balsamifera are used as a folk medicine in kidney stone diseases in South-East Asia. Phytochemical investigation revealed leaves contained a number of flavonoids. In view of these, the present work was aimed to quantify and preliminary pharmacokinetic investigation of five flavonoids viz. dihydroquercetin-7,4¢-dimethyl ether (I), dihydroquercetin-4¢-methyl ether (II), 5,7,3¢,5¢-tetrahydroxyflavanone (III), blumeatin (IV) and quercetin (V) in rat plasma following oral administration (0.5g/Kg) of B. balsamifera leaf extract in rats. Quantification was achieved by using a validated, reproducible high-performance liquid chromatographic method. The mean recoveries of I, II, III, IV and V were 90.6, 93.4, 93.5, 91.2 and 90.3% respectively. The limit of quantification was 25 ng/mL for I and IV, 10 ng/mL for II and III and 100 ng/mL for V respectively. The within day and day-to-day precision for all the compounds were < 10%. The validated HPLC method herein was applied for pharmacokinetic studies and the main pharmacokinetic parameters were: t1/2 (hr) 5.8, 4.3, 2.9, 5.7 and 7.3, Cmax (ng/mL) 594.9, 1542.9 1659.9, 208.9 and 3040.4; Tmax (hr) 4.7, 1.0, 1.0, 3.5 and 2.3; AUC0-oo (ng hr/mL) 5040, 5893, 9260, 1064 and 27233 for I, II, III, IV and V respectively. The developed method was suitable for pharmacokinetic studies and this preliminary study also revealed significant absorption after oral dosing in rats.

In vitro and in vivo anti-colon cancer effects of Garcinia mangostana xanthones extract.

BACKGROUND: Xanthones are a group of oxygen-containing heterocyclic compounds with remarkable pharmacological effects such as anti-cancer, antioxidant, anti-inflammatory, and antimicrobial activities. METHODS: A xanthones extract (81% α-mangostin and 16% γ-mangostin), was prepared by crystallization of a toluene extract of G. mangostana fruit rinds and was analyzed by LC-MS. Anti-colon cancer effect was investigated on HCT 116 human colorectal carcinoma cells including cytotoxicity, apoptosis, anti-tumorigenicity, and effect on cell signalling pathways. The in vivo anti-colon cancer activity was also investigated on subcutaneous tumors established in nude mice. RESULTS: The extract showed potent cytotoxicity (median inhibitory concentration 6.5 ± 1.0 µg/ml), due to induction of the mitochondrial pathway of apoptosis. Three key steps in tumor metastasis including the cell migration, cell invasion and clonogenicity, were also inhibited. The extract and α-mangostin up-regulate the MAPK/ERK, c-Myc/Max, and p53 cell signalling pathways. The xanthones extract, when fed to nude mice, caused significant growth inhibition of the subcutaneous tumor of HCT 116 colorectal carcinoma cells. CONCLUSIONS: Our data suggest new mechanisms of action of α-mangostin and the G. mangostana xanthones, and suggest the xanthones extract of as a potential anti-colon cancer candidate.

Antiangiogenesis and antioxidant activity of ethanol extracts of Pithecellobium jiringa.

BACKGROUND: Angiogenesis plays a critical role in embryonic development and various physiological processes. However, excessive angiogenesis is associated with several pathological conditions including cancer. Pithecellobium jiringa (Jack) Prain is a traditional medicinal plant from the family Leguminosae. It is native to the Southeast Asia, where it has been used traditionally for treatment of various ailments such as hypertension and diabetes. The present work is aimed to study antioxidant and antiangiogenesis activities of P. jiringa ethanol extracts. METHODS: P. jiringa fruit rinds were extracted with ethanol and 50% ethanol. The antioxidant property was analysed using, 1,1-diphenyl-2-picryl-hydrazyl free radical scavenging assay. Phytochemical analysis was performed using thin layer chromatography and colorimetric methods. Then, cell growth inhibition was studied against a panel of human cell lines by MTT test. In vitro inhibition of angiogenesis was studied by the following assays: isolated rat aortic rings cell viability, colony formation, endothelial cell migration, endothelial tube formation on matrigel, and expression of vascular endothelial growth factor by endothelial cells. In vivo antiangiogenesis effect was studied by utilising fertilised chick embryos assay. The results were statistically analysed by analysis of variance. RESULTS: Ethanolic and 50% hydro-ethanolic extracts showed relatively high concentration of total phenolics associated with potent antioxidant activity. The rat aortic rings study conducted showed potent inhibition of the microvessels outgrowth with IC50s 5.27 ± 0.81 µg/ml (ethanolic) and 4.45 ± 0.63 µg/ml (50% hydro-ethanolic). Both extracts arrested the growth of human endothelial cells via down-regulation of VEGF expression, leading to inhibition of other angiogenesis cascades including migration of endothelial cells, and formation of capillary network on matrigel matrix. The extracts also inhibited the neovascularisation of chick embryo chorioallantoic membrane. CONCLUSIONS: P. jiringa extracts inhibit angiogenesis by blocking the VEGF expression thus inhibiting endothelial cells proliferation, migration and differentiation most likely due to presence of the antioxidant phenolics.

alpha-Mangostin enhances betulinic acid cytotoxicity and inhibits cisplatin cytotoxicity on HCT 116 colorectal carcinoma cells.

Despite the progress in colon cancer treatment, relapse is still a major obstacle. Hence, new drugs or drug combinations are required in the battle against colon cancer. α-Mangostin and betulinic acid (BA) are cytotoxic compounds that work by inducing the mitochondrial apoptosis pathway, and cisplatin is one of the most potent broad spectrum anti-tumor agents. This study aims to investigate the enhancement of BA cytotoxicity by α-mangostin, and the cytoprotection effect of α-mangostin and BA on cisplatin-induced cytotoxicity on HCT 116 human colorectal carcinoma cells. Cytotoxicity was investigated by the XTT cell proliferation test, and the apoptotic effects were investigated on early and late markers including caspases-3/7, mitochondrial membrane potential, cytoplasmic shrinkage, and chromatin condensation. The effect of α-mangostin on four signalling pathways was also investigated by the luciferase assay. α-Mangostin and BA were more cytotoxic to the colon cancer cells than to the normal colonic cells, and both compounds showed a cytoprotective effect against cisplatin-induced cytotoxicity. On the other hand, α-mangostin enhanced the cytotoxic and apoptotic effects of BA. Combination therapy hits multiple targets, which may improve the overall response to the treatment, and may reduce the likelihood of developing drug resistance by the tumor cells. Therefore, α-mangostin and BA may provide a novel combination for the treatment of colorectal carcinoma. The cytoprotective effect of the compounds against cisplatin-induced cytotoxicity may find applications as chemopreventive agents against carcinogens, irradiation and oxidative stress, or to neutralize cisplatin side effects.

A review of the literature and latest advances in research of Piper sarmentosum.

CONTEXT: Piper sarmentosum Roxb. (Piperaceae) is a traditional medicinal as well as a culinary plant in South East Asian countries, whereby aerial parts of the plant are consumed as a vegetable in various forms and the whole plant or parts are used as folk remedies, alone or in combination with other herbs, to treat various ailments. The plant has extensively been investigated in a broad range of studies to provide scientific evidence for folklore claims or to find new therapeutic uses; however, heretofore, a summary of the data are not available. OBJECTIVE: In order to describe nutritional and therapeutic potential of P. sarmentosum and summarize scientific evidence that supports traditional claims, a literature review and latest advances in research of the plant are given herein. MATERIALS AND METHODS: The literature has been retrieved from a number of databases such as Google Scholar, PubMed, Medline, Science Direct and SciFinder. The articles related to synthetic work, ecology and agriculture have been excluded. RESULTS AND DISCUSSION: The review has not only revealed a number of pharmacological activities supporting the traditional claims but indicates new prospects for the plant. Antiangiogenic activity and toxicity studies suggest the usage of the plant in treating diseases involving neo-vascularization. The available efficacy, safety, pharmacokinetic and stability data urge clinical studies on extracts of the plant. CONCLUSION: The present review may be helpful to future researchers intending to investigate the plant and natural pharmaceutical industry for preparing evidence-based formulations.

Rapid separation and determination of betulinic acid from a complex matrix using combination of TLC and RP-HPLC.

Hitherto, only a few studies are reported about using the combination of TLC and RP-HPLC for the separation and determination of analyte(s) from a complex matrix. The present study is aimed to develop a simple and rapid method for the separation and determination of betulinic acid from a complex matrix, extracts of Orthosiphon stamineus, using a combination of the two techniques. The samples having higher contents of the analyte and fewer interfering species were prepared using TLC. The samples were then eluted through C(18) column using isocratic solvent system comprising acetonitrile, methanol and acetic acid acidified water of pH 2.8 in a ratio of 70 : 20 : 10 (v/v/v), respectively, and detection was carried out at 210 nm. The method was validated and applied successfully to quantify betulinic acid in various types of extracts of the plant. The limit of detection (LOD) and limit of quantification (LOQ) were found to be 0.0005 and 0.0050 µg/ml, respectively. The method exhibited linearity in a concentration range of 0.005-100.00 µg/ml (R(2)= 0.9999). The recovery was found to be 97.10 - 97.60% (RSD < 5%), whereas, intra-day and inter-days accuracy values were 97.13 - 98.67% (RSD < 5%) and 96.45 - 98.00% (RSD < 5%), respectively. The results of the present study indicate that the developed method is simple, rapid, sensitive and accurate, and may be of a value to natural product industry and researchers for the standardization of extracts containing betulinic acid in a lesser time and consuming fewer solvents.

Evaluation of antiangiogenic and antoxidant properties of Parkia speciosa Hassk extracts.

Parkia speciosa Hassk is a traditional medicinal plant with strong antioxidant and hypoglycemic properties. This study aims to investigate the total phenolic content, antioxidant, cytotoxic and antiangiogenic effect of eight extracts from P. speciosa empty pods. The extracts were found to contain high levels of total phenols and demonstrated strong antioxidant effect in DPPH scavenging test. In rat aortic rings, P. speciosa extracts significantly inhibited the microvessel outgrowth from aortic tissue explants by more than 50%. The antiangiogenic activity was further confirmed by tube formation on matrigel matrix involving human endothelial cells. Cytotoxic effect was evaluated by XTT test on endothelial cells as a model of angiogenesis versus a panel of human cancer and normal cell lines. Basically the extracts did not show acute cytotoxicity. Morphology examination of endothelial cells indicated induction of autophagy characterized by formation of plenty of cytoplasmic vacuoles. The extracts were found to work by decreasing expression of vascular endothelial growth factor in endothelial cells.

Antiangiogenic properties of Koetjapic acid, a natural triterpene isolated from Sandoricum koetjaoe Merr.

BACKGROUND: Angiogenesis, the formation of new blood vessels, has become an important target in cancer therapy. Angiogenesis plays an important role in tumor growth and metastasis. Koetjapic acid (KA) is a seco-A-ring oleanene triterpene isolated from S. koetjape. The solvent extract of this plant species was shown previously to have strong antiangiogenic activity; however the active ingredient(s) that conferred the biological activity and the mode of action was not established. Given the high concentration of KA in S. koetjape, an attempt has been made in this study to investigate the antiangiogenic properties of KA. RESULTS: Treatment with 10-50 µg/ml KA resulted in dose dependent inhibition of new blood vessels growth in ex vivo rat aortic ring assay. KA was found to be non-cytotoxic against HUVECs with IC50 40.97 ± 0.37 µg/ml. KA inhibited major angiogenesis process steps, endothelial cell migration and differentiation as well as VEGF expression. CONCLUSIONS: The non-cytotoxic compound, KA, may be a potent antiangiogenic agent; its activity may be attributed to inhibition of endothelial cells migration and differentiation as well VEGF suppression.

Bioactive Markers Based Pharmacokinetic Evaluation of Extracts of a Traditional Medicinal Plant, Piper sarmentosum.

In vitro assays are economical and easy to perform but to establish relevance of their results to real clinical outcome in animals or human, pharmacokinetics is prerequisite. Despite various in vitro pharmacological activities of extracts of Piper sarmentosum, there is no report of pharmacokinetics. Therefore, the present study aimed to evaluate ethanol extract of fruit of the plant in dose of 500 mg kg(-1) orally for pharmacokinetics. Sprague-Dawley rats were randomly divided into groups 1, 2, and 3 (each n = 6) to study absorption, distribution and excretion, respectively. High performance liquid chromatography (HPLC) with ultraviolet detection was applied to quantify pellitorine, sarmentine and sarmentosine in plasma, tissues, feces and urine to calculate pharmacokinetic parameters. Pellitorine exhibited maximum plasma concentration (C(max)) 34.77 ng mL(-1) ± 1.040, time to achieve C(max) (T(max)) 8 h, mean resident time (MRT) 26.00 ± 0.149 h and half life (t(1/2)) 18.64 ± 1.65 h. Sarmentine showed C(max) 191.50 ± 12.69 ng mL(-1), T(max) 6 h, MRT 11.12 ± 0.44 h and t(1/2) 10.30 ± 1.98 h. Sarmentosine exhibited zero oral bioavailability because it was neither detected in plasma nor in tissues, and in urine. Pellitorine was found to be distributed in intestinal wall, liver, lungs, kidney, and heart, whereas sarmentine was found only in intestinal wall and heart. The cumulative excretion of pellitorine, sarmentine and sarmentosine in feces in 72 h was 0.0773, 0.976, and 0.438 µg, respectively. This study shows that pellitorine and sarmentine have good oral bioavailability while sarmentosine is not absorbed from the gastrointestinal tract.

Standardization and in vivo antioxidant activity of ethanol extracts of fruit and leaf of Piper sarmentosum.

The present study aimed to investigate standardized ethanol extracts of fruit and leaves of Piper sarmentosum for their in vivo antioxidant activity in rats using a CCl (4)-induced oxidative stress model. The standardization was based on the quantification of the markers pellitorine, sarmentine and sarmentosine by high performance liquid chromatography (HPLC), and determination of total primary and secondary metabolites. The rats, divided into 7 groups each (n = 6), were used as follows: group 1 (CCl (4), negative control), group 2 (untreated, control), groups 3 and 4 (fruit extract 250 and 500 mg/kg, respectively), groups 5 and 6 (leaf extract 250 and 500 mg/kg, respectively) and group 7 (vitamin-E 100 mg/kg, positive control). The doses were administered orally for 14 days; 4 h following the last dose, a single dose of CCl (4) (1.5 mg/kg) was given orally to all the groups except group 2, and after 24 h, blood and liver of each animal were obtained. Analysis of plasma and liver homogenate exhibited significant preservation of markers of antioxidant activity, total plasma antioxidant activity (TPAA), total protein (TP), superoxide dismutase (SOD), catalase (CAT), and thiobarbituric acid reactive species (TBARS), in the pretreated groups as compared to the CCl (4) group (p < 0.05). Histology of the liver also evidenced the protection of hepatocytes against CCl (4) metabolites in the pretreated groups. The results of this study indicate the IN VIVO antioxidant activity of both extracts of the plant, which may be valuable to combat diseases involving free radicals.

7-Hy-droxy-6-meth-oxy-2H-chromen-2-one.

The title compound, C(10)H(8)O(4), is one of the coumarins existing in Morinda citrifolia L (Noni). The chromenone ring system is approximately planar with a maximum deviation of 0.0208 (14) Å. The meth-oxy group does not deviate from this plane [C-O-C-C torsion angle = -1.5 (3)°], indicating that the whole mol-ecule is almost planar. In the crystal packing, inter-molecular O-H⋯O hydrogen bonds link the mol-ecules into chains. These are further connected by C-H⋯O hydrogen bonds.

Xanthine oxidase inhibitory activities of extracts and flavonoids of the leaves of Blumea balsamifera.

CONTEXT: Blumea balsamifera DC (Compositae) leaves have been recommended for use as a folk medicine in the treatment of various diseases related to urolithiasis in southeast Asia. Phytochemical studies of this plant revealed it contains four classes of flavonoids (e.g., flavonols, flavones, flavanones, and dihydroflavonol derivatives). OBJECTIVE: In view of the broad pharmacological activity of flavonoids, this study was carried out to determine the xanthine oxidase (XO) inhibitory and enzymatically produced superoxide radical scavenging activity of different organic extracts and that of the isolated flavonoids from B. balsamifera leaves. MATERIALS AND METHODS: The inhibitory activity of XO was assayed spectrophotometrically at 295 nm. The superoxide radicals scavenging activity was assessed by NBT reduction method, spectrophotometrically at 560 nm. A dose response curve was plotted for determining IC50 values. RESULTS: The methanol extract (IC50 = 0.111 mg/mL) showed higher XO inhibitory activity than the chloroform (0.138 mg/mL) and pet-ether extracts (0.516 mg/mL). IC50 values of scavenging of superoxide radicals for extracts decreased in the order of: methanol (0.063 mg/mL) > chloroform (0.092 mg/mL) > pet-ether (0.321 mg/mL). The XO inhibitory activity of the isolated flavonoids and reference compounds tested decreased in the order of: allopurinol > luteolin > quercetin > tamarixetin > 5,7,3',5'-tetrahydroxyflavanone > rhamnetin > luteolin-7-methyl ether > blumeatin > dihydroquercetin-4'-methyl ether > dihydroquercetin-7,4'-dimethyl ether > L-ascorbic acid. DISCUSSION AND CONCLUSION: The results indicated that the flavone derivatives were more active than the flavonol derivatives. The flavanone derivatives were moderately active and the dihydroflavonol derivatives were the least. The higher flavonoid content of extracts contributed to their higher XO inhibitory activity.