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Dioscoreanone, a 1,4-phenanthraquinone isolated from an ethanolic extract of the rhizome of Dioscorea membranacea, Pierre ex Prain & Burkill, a plant which has been used to treat inflammation and cancer in Thai Traditional Medicine. In this study, the mechanisms of dioscoreanone on LPS-induced NO production and cytokine expression through the activation of NF-κB and ERK1/2 are demonstrated in RAW 264.7 cells. Through measurement with Griess reagent, dioscoreanone was found to reduce NO levels with an IC(50) value of 2.50 ± 0.64 µM, due to the significant suppression of LPS-induced iNOS mRNA expression, as well as IL-1β and IL-6 levels at a concentration of 6 µM. At the signal transduction level, using the pNFκB-Luciferase reporter system, dioscoreanone significantly inhibited NF-κB transcriptional activity, which resulted from the prevention of IκBα degradation. In addition, dioscoreanone in the range of 1.2-5 µM significantly enhanced LPS-induced ERK1/2 activation and dioscoreanone alone induced the activation of ERK1/2 proteins in a concentration- and time-dependent response. The activation of ERK1/2 proteins by dioscoreanone was due to both an arylating reaction, which was suppressed by N-acetyl cysteine, and a redox cycling reaction of NQOR, which was inhibited by dicoumarol. In conclusion, the mechanisms of dioscoreanone on the inhibition of NO production and mRNA expression of iNOS, IL-1β, and IL-6 were due to both the inhibition of NF-κB activation and the activation of ERK1/2 proteins. The activation of dioscoreanone may in turn inhibit the binding of NF-κB to pro-inflammatory gene promoters in LPS-induced RAW264.7 macrophage cells.

Mulberry (Morus alba L.) leaf tea is promoted for its health benefits and the control of diabetes in Asian nations. The blood glucose lowering activity of mulberry leaf extract (MA) has been proven; however, the molecular basis underlying this effect remains unclear. The aim of the present work is to elucidate its mechanism of the antihyperglycemic action, by examining the effect of MA on glucose uptake and the translocation of glucose transporter 4 protein (GLUT4) to the plasma membrane of adipocytes isolated from diabetic rats. The incubation of adipocytes with 5–45 μg/ml MA resulted in 31–54% increase of glucose uptake in a dose-dependent manner. This glucose uptake enhancing effect was inhibited by the phosphoinositol 3-kinase (PI3-K) inhibitor, wortmannin (100 nM). The GLUT4 protein on the plasma membrane fraction of adipocytes was markedly increased after treatment with 15 μg/ml MA extract. Interestingly, gallic acid, one of the phenolic compounds found in MA extract, increased glucose uptake and enhanced the translocation of GLUT4 at concentrations comparable to the amount of gallic acid in the effective concentration ranges of MA. Thus, it is likely that gallic acid contributes, at least in part, to its antihyperglycemic activity. The present results suggest that the antihyperglycemic action of MA is mediated by increasing glucose uptake via the activation of PI3-K signaling pathway and translocation of GLUT4 to the plasma membrane. These findings are the first molecular evidence supporting the mulberry tea as herbal medicine for diabetic patients.

Thirteen plants and their parts acquired from southern Thailand were investigated for their polysaccharide contentsand prebiotic properties. The fresh, ground samples were extracted with 50% and 95% ethanol and water at ambient and boilingtemperatures. The extracts were freeze-dried, digested with HCl buffer and -amylase, and indigestible polysaccharidecontents were determined. Base on extract yields and indigestible polysaccharide contents, ten samples were chosen aspotential sources of prebiotics. These included embryo, flesh and pericarp of palm fruit (Borrassus flabellifer L.), skin, fleshand seed of jackfruit (Artocarpus heterophyllus Lam.), flesh of rambutan (Nephelium lappaceum L.), jampadah (Artocarpusinteger Merr.), and young coconut (Cocos nucifera Linn.), and okra pod (Hibiscus esculentus Linn.). Their extract yields (%dry wt) were 26.54, 44.94, 51.69, 71.54, 59.43, 16.00, 55.73, 34.11, 22.66, and 12.39, respectively, and indigestible polysaccharidecontents (mg/g dry extract) were 409.85, 334.87, 705.80, 689.08, 605.76, 403.44, 566.83, 542.56, 513.87, and 460.73, respectively.The amounts of oligosaccharide were 33.69, 47.20, 14.13, 0.00, 98.05, 29.35, 9.43, 2.40, 0.00, and 49.15 mg/g dry extract, respectively. Subsequently, five samples were chosen for further studies and possible commercial development based on theirextract yield, the amount and type of oligosaccharides, i.e. palm flesh, palm embryo, jackfruit flesh, jackfruit seed, and okrapod. Molecular weights of the polysaccharides from the five samples were 190-1,600 Daltons with a degree of polymerizationof 5-6.

To compare Pueraria mirifica 25 and 50 mg dosages to relieve menopausal symptoms. A double-blind study was conducted on 52 hysterectomized women with menopausal symptoms who had an indication for hormone therapy. The women were randomly allocated into two groups receiving either Pueraria mirifica 25 mg tablet (Arm A; n = 26) or Pueraria mirifica 50 mg (Arm B; n = 26) for 6 months. Menopausal symptoms, physical examination findings, and laboratory data were recorded at baseline. Symptoms were reevaluated at 3 and 6 months, while physical and laboratory examinations were evaluated at 1 month and at the end of treatment. A modified Green climacteric scale was used to evaluate menopausal symptoms; a score of ≥15 indicated estrogen deficiency. All women completed the study. Mean baseline climacteric scores of women who received 25 and 50 mg were: 24.19 ± 9.11 versus 23.19 ± 7.89, respectively (p = 0.674). After 3 and 6 months of treatment, scores were 17.92 ± 10.40 versus 15.35 ± 8.44 (p = 0.332) and 14.08 ± 10.30 versus 12.46 ± 6.38 (p = 0.500), respectively. No significant side effects were observed with Pueraria mirifica 25 or 50 mg. Both dosages of Pueraria mirifica were similarly effective and safe in the treatment of menopausal symptoms.

Dioscorealide B (DB), a naphthofuranoxepin has been purified from an ethanolic extract of the rhizome of Dioscorea membranacea Pierre ex Prain & Burkill which has been used to treat inflammation and cancer in Thai Traditional Medicine. Previously, DB has been reported to have anti-inflammatory activities through reducing nitric oxide (NO) and tumor necrosis factor-a (TNF-a) production in lipopolysaccharides (LPS)-induced RAW 264.7 macrophage cells. In this study, the mechanisms of DB on LPS-induced NO production and cytokine expression through the activation of nuclear factor-kB (NF-kB) and ERK1/2 are demonstrated in RAW 264.7 cells. Through measurement with Griess’s reagent, DB reduced NO level with an IC50 value of 2.85 � 0.62 mM that was due to the significant suppression of LPS-induced iNOS mRNA expression as well as IL-1b, IL-6, and IL-10 mRNA at a concentration of 6 mM. At the signal transduction level, DB significantly inhibited NF-kB binding activity, as determined using pNFkB-Luciferase reporter system, which action resulted from the prevention of IkBa degradation. In addition, DB in the range of 1.5‐6 mM significantly suppressed the activation of the ERK1/2 protein. In conclusion, the molecular mechanisms of DB on the inhibition of NO production and mRNA expression of iNOS, IL-1b, IL-6, and IL-10 were due to the inhibition of the upstream kinases activation, which further alleviated the NF-kB and MAPK/ERK signaling pathway in LPS-induced RAW264.7 macrophage cells. J. Cell. Biochem. 109: 1057‐1063, 2010. 2010 Wiley-Liss, Inc.

Background Skew deviation is an acquired vertical ocular misalignment caused by damage to the prenuclear vestibular inputs to the ocular motor nuclei. A-pattern strabismus often has bilaterally symmetric vertical incomitance and overdepression in adduction (superior oblique overaction) and can be associated with developmental delay, cerebral palsy, hydrocephalus, spina bifida, or posterior fossa or other brainstem disease. The purpose of this study is to describe the ocular motility and torsion findings in patients with A-pattern strabismus and bilateral overdepression in adduction (superior oblique muscle overaction) and to propose a possible brainstem mechanism underlying these observations. Results Most of the 13 patients identified had other neurologic abnormalities, including spina bifida, hydrocephalus, perinatal stroke, or global delay. Only 2 patients had vertical ocular misalignment in primary gaze. Of the 13, 7 had incomitant vertical tropias during lateral gaze, and 12 had bilateral incyclotorsion documented on fundus examination. Despite having bilateral overdepression in adduction (superior oblique overaction), 11 of the 13 had no difference in vertical ocular misalignment with alternating head tilt rather than reversing hypotropias as would be expected from primary oblique dysfunction. The findings are consistent with damage to the utricular pathways corresponding to the anterior semicircular canal and a resulting posterior canal predominance to the extraocular muscle subnuclei that creates increased tonus to the depressors, bilaterally. Conclusions A-pattern strabismus may, in some cases, represent a special form of skew deviation. The ocular motility and clinical findings are consistent with bilateral damage to the utricular pathways corresponding to the anterior semicircular canals rather than bilateral primary superior oblique muscle overaction.

Nutraceuticals and functional foods have become an important tool for consumers to manage their health and wellness.Pre-, pro-, and synbiotics are a part of this group of products shown to have properties that can modulate gastrointestinalproblems and improve general health and well being. Prebiotics are polysaccharides that can withstand acidic and enzymaticdigestion in the small intestine and can be utilized by probiotics, and gut microflora, in the large intestine for their growth andactivities that benefit the host’s health, e.g., by enhancing the immunity and mineral absorption, preventing colon cancer andother gastrointestinal diseases, and lowering cholesterol. Prebiotics, e.g., inulin, oligofructose, oligolactose, and lactulose, occur naturally in many plants, or may be synthesized from starch or other carbohydrates using appropriate enzymes.A number of plants from southern Thailand, e.g., palm fruit (Borrassus flabellifer L.), jackfruit (Artocarpus heterophyllusLam.), young coconut (Cocos nucifera Linn.), rambutan (Nephelium lappaceum L.), jampadah (Artocarpus integer Merr.), and okra (Abelmoschus esculentus Moench.) contain a considerable amount of polysaccharides that have been shown to have prebiotic properties. Prebiotics may be used as ingredients in functional foods, or may be presented as nutraceuticals in the form of capsules, tablets or powder, sometimes together with probiotic cultures.

Free radical-induced vascular dysfunction plays a key role in the pathogenesis of vascular disease found in chronic diabetic patients. Morus alba (MA) leaf extract is promoted for good health especially in diabetic patients. Interestingly, antidiabetic and antioxidant activities of MA have been reported in experimental animals. Thus, the hypothesis of this study was that the long-term treatment with MA could improve vascular reactivity of chronic diabetic rats. To test this hypothesis, we examined the effect of long-term treatment with MA on the vascular responses to vasoactive agents in streptozotocin-induced chronic diabetic rats. The diabetic rats were either orally administered with distilled water, MA (0.25, 0.5 and 1 g/kg per day) or subcutaneously injected with insulin (4 U/kg per day) for 8 weeks. After each treatment, the fasting blood glucose, blood pressure, vascular responses to vasoactive agents and tissue malondialdehyde were examined. Morus alba at the doses of 0.5 and 1 g/kg, which significantly reduced blood glucose level, also significantly decreased the high blood pressure in diabetic rats. Vascular responses of the chronic diabetic rats to vasodilators, acetylcholine (3-30 nmol/kg) and sodium nitroprusside (1-10 nmol/kg) were significantly suppressed by 26% to 44% and 45% to 77% respectively, whereas those to vasoconstrictor, phenylephrine (0.01-0.1 micromol/kg) were significantly increased by 23% to 38% as compared to normal rats. Interestingly, the administration of 0.5 and 1 g/kg MA or 4 U/kg insulin significantly restored the vascular reactivities of diabetic rats. Moreover, 8 weeks of diabetes resulted in the elevation of malondialdehyde content in tissues (liver, kidney, heart, and aorta), and MA treatment significantly lessened this increase. These results provide the first evidence for the efficacy of MA in restoring the vascular reactivity of diabetic rats, the mechanism of which may associate with the alleviation of oxidative stress.

Thirteen Thai crops including banana, okra, jackfruit, germinated rice, rambutan, durian, jampadah, huasa potato,tamarind, coconut, mango, fan palm fruit and dioscorea tuber were tested for anti-allergic effect using RBL-2H3 cells andanti-microbial activity. These 13 crops, some of which included different parts, e.g. skin, flesh, and seed, were extracted withfour solvents separately [(95% ethanol (EtOH), 50% EtOH, water (W) and hot water (HW)], respectively, to obtain 112extracts. Among these extracts, mango seed in 50% EtOH possessed the highest anti-allergic activity against antigen-inducedb-hexosaminidase release as a marker of degranulation in RBL-2H3 cells with an IC50 value of 7.5±0.8 mg/ml, followed bybanana (W, IC50 = 13.5±2.4 mg/ml), okra (W, IC50 = 13.6±3.1 mg/ml), jampadah skin (HW, IC50 = 13.8±3.9 mg/ml), tamarindseed coat (HW, IC50 = 14.2±3.1 mg/ml), jampadah flesh (W, IC50 = 14.6±3.1 mg/ml); whereas other crops possessed IC50values from 21.5->100 mg/ml. Moreover, the plants showing high anti-allergic effects were also possessed marked antibacterialactivity. Rambutan peel, mango peel, mango seed and tamarind seed coat exhibited appropriate anti-bacterialactivity against Staphylococcus aureus, Bacillus subtilis and Pseudomonas aeruginosa with MIC values ranging from 250-2,000 mg/ml, but did not show any effect towards Escherichia coli and Candida albicans. This study indicates that these Thaicrops may have potential as functional foods and neutraceuticals for treatment of allergy, allergy-related diseases and somebacterial infections.

A reversed-phase high-performance liquid chromatographic method has been developed and validated for the determination of pyridostemin, the major pesticidal alkaloid found in Stemona curtisii. This methodology was applied to the investigation of plant extracts and water dispersible granule formulations. Stability indicating procedures have also been carried out. The chromatographic separation was on a C18 column with a mixture of acetonitrile–water–triethylamine (30:70:0.12, v/v/v), using UV detection at 300 nm. Validation procedures showed that the method was specific, accurate and precise. The response was linear over a range of 5–25 μg mL−1 with recoveries in the range of 98.28–102.85%. The RSD for intra- and inter-day precision were

High-performance liquid chromatography-photodiode array detection (HPLC-PDA) was developed for chromatographic fingerprint analysis of the ethanolic extract from Dioscorea membranacea Pierre (known as Hua-Khow-Yen in Thai medicinal plant). Chromatographic fingerprint, together with the contents of the dioscorealide A, dioscorealide B and dioscoreanone, which are three markers of the main constituents were first established and applied for quality control. The samples were separated with reversed phase HPLC by linear gradient elution using water and acetonitrile as mobile phase at a flow rate of 1.0 ml/min and detector wavelength at 245 nm for dioscoreanone and at 270 nm for dioscorealide A and dioscorealide B. This HPLC fingerprint analysis has proven its usefulness for identification and assessment for quality evaluation of D. membranacea extract.

The effects of the extracts from Hibiscus sabdariffa calyces L. (H. sabdariffa) on nociceptive response using writhing, hot plate and formalin test in mice and the antipyretic activity in yeast-induced fever in rats, were examined. Anti-inflammatory activity was also investigated on carrageenin-induced paw edema in rats. No acute toxicity was observed in mice after oral administration of the ethanol and aqueous extract of H. sabdariffa calyces at the dose of 15 g/kg. Oral administration of the ethanol extract at the dose of 800 mg/kg significantly decreased the number of contortions and stretchings induced by acetic acid in mice. The aqueous extracts had no effect on this test. Neither the ethanol nor aqueous extract had an effect in the formalin and hot plate tests in mice. The ethanol and the vacuum dried extract of H. sabdariffa calyces (200-800 mg/kg, p.o.) decreased the yeast-induced fever in rats. The H. sabdariffa extract had no effect on carrageenininduced paw edema in rats. These results suggest that the ethanol and aqueous extract (vacuum dry) of H. sabdariffa calyces possess antipyretic action through mechanisms that are different from that of aspirin.

The effects of either ethanol or aqueous extract from rhizomes of Smilax corbularia Kunth (S. corbularia) on anti-inflammatory activity in carrageenin-induced edema in rats, antinociceptive activity using writhing, hot plate and formalin tests in mice and the antipyretic activity in yeast-induced fever were examined. Oral administration of the ethanol extract of S. corbularia rhizomes significantly suppressed the paw edema induced by carrageenin in rats while the aqueous extract had no effect. Neither the ethanol extract nor aqueous extract significantly affected the antinociceptive tests in mice and yeast-induced fever in rats. These results suggest that the ethanol extract of S. corbularia rhizomes possess anti-inflammatory activity, and that its actions on the inflammation may be different from those of aspirin.

Thai medicinal plants locally known as Hua-Khao-Yen were examined for their inhibitory activities against lipopolysaccharide (LPS) induced nitric oxide (NO) production in RAW 264.7 cell lines. Among the plant species studied, an ethanolic extract of Dioscorea membranacea exhibited the most potent inhibitory activity, with an IC(50) value of 23.6 microg/ml. From this extract, eight compounds [two naphthofuranoxepins (1, 2), one phenanthraquinone (3), three steroids (4-6) and two steroidal saponins (7, 8)] were isolated and further investigated for their inhibitory properties of NO production. It was found that diosgenin-3-O-alpha-L-rhamnosyl (1-->2)-beta-D-glucopyranoside (7) possessed the highest activity (IC(50)=3.5 microM), followed by dioscoreanone (3, IC(50)=9.8 microM) and dioscorealide B (2, IC(50)=24.9 microM). Regarding structural requirements of diosgenin derivatives for NO production inhibitory activity, compound 7 which has a rhamnoglucosyl moiety at C-3 exhibited much higher activity than compounds that have either a diglucosyl substitution (8) or its aglycone (9); whereas, hydroxyl substitution at position 8 of naphthofuranoxepin derivatives conferred higher activity than the methoxyl group. It is concluded that diosgenin-3-O-alpha-L-rhamnosyl (1-->2)-beta-D-glucopyranoside (7), dioscoreanone (3) and dioscorealide B (2) are active principles for NO inhibitory activity of Dioscorea membranacea. Compounds 1-3 were also tested for the inhibitory effect on LPS-induced TNF-alpha release in RAW 264.7 cells. The result revealed that 3 possessed potent activity against TNF-alpha release with an IC(50) value of 17.6 microM, whereas, 1 and 2 exhibited mild activity. The present study may support the use of Dioscorea membranacea by Thai traditional doctors for treatment of the inflammatory diseases.

The rhizomes of Smilax glabra (SG) has long been used in Traditional Chinese and Thai herbal medicine to treat a variety of infectious diseases and immunological disorders.To investigate the in vitro anti-allergic activities of crude extracts andpure isolated flavonoid compounds from SG by determination of inhibitory effect on antigen-induced release of β-hexosaminidasefrom RBL-2H3 cells.The in vitro inhibitory effects ofcrude aqueous and organic extracts on β-hexosaminidase release in RBL-2H3 cells were evaluated as an in vitro indication ofpossible anti-allergic activity. Bioassay-guided fractionation of extracts was used to isolate flavonoid compounds from the ethanolic extracts.The 95% and 50% ethanolic extracts of SG showed remarkably high anti-allergic activity, with IC50 values of 5.74 ± 2.44 and 23.54 ± 4.75 µg/ml, much higher activity than that for Ketotifen (IC50 58.90 µM). The water extract had negligible activity (IC50100 µg/ml). The two isolated flavonols, Engeletin and Astilbin, showed weak anti-allergic activity, IC50 values 97.46 ± 2.04 and100 µg/ml, respectively.The 95% and 50% ethanolic extracts of SG showed strong anti-allergic activity, but two flavonol constituents did not show any significant anti-allergic activity. These findings suggest that a combination of effects of various phytochemicals in crude extracts used in traditional medicine, are responsible for the purported anti-allergic activity of SG herbal preparations. The plethora of constituents in crude extracts, as yet unidentified, are likely to be acting synergistically to account for the strong observed anti-allergic in vitro activity.

Several natural phytochemicals are increasingly used, as an adjunct to chemotherapy, to reduce drug adverse effects. Zingiber officinale rhizome (ginger) product has been reported to be effective against nausea and vomiting in patients receiving emetogenic chemotherapy such as cisplatin/doxorubicin (DXR). In addition, its ethanolic extract of Zingiber officinale rhizome (EEZOR) has been reported to possess anticarcinogenic properties. However, the mechanism for anticancer activity of ginger especially when used in combination with chemotherapy has not well elucidated, one of its possible mechanisms might involve its genotoxicity.To investigate genotoxic and cytotoxic potentials of EEZOR alone and EEZOR pretreatments followed by 0.1 mcg/ ml DXR, a genotoxic chemotherapeutic agent in human lymphocytes by sister chromatid exchange (SCE) assay in vitro. The effect on cell cycle kinetics was also explored.Human lymphocytes were treated with EEZOR alone at 25-500 mcg/ml and EEZOR pretreated at 12.5-200 mcg/ml followed by 0.1 mcg/ml DXR. SCE levels and cell cycle kinetics were evaluated.EEZOR significantly induced biphasic SCE at 50 and 400 mcg/ml (p0.05). However, cytotoxicity manifested at 500 mcg/ml. All EEZOR pretreatments at 12.5, 25, 50, and 100 mcg/ml, except at 200 mcg/ml, prior to DXR, moderately enhanced DXR-induced genotoxicity by 1.3 times (p0.05). Both EEZOR alone and EEZOR prior to DXR at certain concentrations delayed cell cycle.At specific doses, EEZOR could induce genotoxicity and in pretreatments could moderately enhance DXR-induced genotoxicity and delay cell cycle. This finding suggests that dosage use of EEZOR needs to be adjusted for long-term safety. In addition, EEZOR in adjunct to DXM might have potential benefits not only as an emetic agent but also in chemotherapy. Further in vivo animal and human studies to support this evidence is essentially needed.

Combinations of modern medicines with herbal medicines are being developedfor more effectiveness. Data on the safety and drug-herb interactions are needed to be clarified. Ethanolic extract of Plumbago indica root (EEPIR) is medicinally usedfor cancer treatment in Asian traditional medicine. However its mechanism of action is still inconclusive. Our previous study demonstrated that EEPIR was genotoxic and induced cell cycle delay in human lymphocytes in vitro.To investigate genotoxic potency and interference with cell cycle of EEPIR in combination with doxorubicin (DXR), a standard chemotherapeutic agent, in human lymphocytes by in vitro sister chromatid exchange (SCE) assay.Human lymphocytes were pretreated with EEPIR at 6.25-100 mcg/mlfollowed by DXR (0.1 mcg/ml). SCE levels and cell cycle kinetics were evaluated.EEPIR pretreatments (6.5-50 mcg/ml) significantly enhanced genetic damage induced by DXR (p0. 05). Delaying of the cell cycle was detected and related to EEPIR concentration. EEPIR at 100 mcg/ml, on the contrary, did not enhance DXR-induced genotoxicity but tended to lower genotoxicity compared to DXR treatment alone. It significantly delayed cell cycle the most (p0.05).EEPIR pretreatments at proper doses enhanced genotoxic damage induced by DXR in human lymphocytes. Delaying cell cycle by EEPIR could lower that potency. Usage of EEPIR, therefore, should be adjusted for safety. Combination of EEPIR with DXR might be usefulfor more efficient cancer treatment with less DXR toxicity. Further in vivo study is needed to support this in vitro evidence.

Purpose: To investigate the cytotoxic and antioxidant activities of the fruit and fruit waste (residue and marc) extracts of Antidesma thwaitesianum Müll. Arg., known as mamao in Thai, using chemical and cell-based assays.Methods: The cytotoxicity of mamao fruit and fruit waste extracts obtained by expression, maceration and decoction against a panel of six human cancer cell lines (COR-L23, A549, LS174T, PC-3, MCF7 and HeLa) was determined by sulforhodamine B (SRB) assay. In addition, their antioxidant activities were measured by chemical methods: 2,2′-azino-bis-3-ethylbenzthiazoline-6-sulphonic acid (ABTS) radical scavenging assay and ferric ion reducing antioxidant power (FRAP) assay and cell-based methods: nitroblue tetrazolium (NBT) dye reduction assay and nitric oxide (NO) scavenging assay. The phenolic and flavonoid contents were assessed colorimetrically at 765 nm and 415 nm respectively.Results: Among the test extracts, the ethanol extracts of fresh fruits (FME) and marc left after squeezing fresh fruits (MME) exhibited moderate cytotoxicity against human breast MCF7 cells while the extract obtained by decocting the residue left after maceration of dried fruits (RDW) was moderately cytotoxic to lung large cell carcinoma COR-L23 cells. In the chemical assays, the extract obtained by decocting the residue left after maceration of dried marc (RMW) displayed the strongest ABTS radicalscavenging and ferric-reducing activities among the extracts. In the cell-based assays, however, FME and DME exerted potent nitric oxide scavenging activity whereas the extract obtained by decocting the residue left after maceration of fresh fruits (RFW) showed moderate superoxide radical-scavenging activity relative to the test extracts. The ABTS radical-scavenging and ferric-reducing activities of these extracts strongly correlate with their phenolic and flavonoid contents, indicating their specific contributions to such activities.Conclusion: The ethanol extracts of fresh and dried mamao fruits exhibit both cytotoxic and cellular antioxidant activities, and thus possess great potentials for application in the development of effective dietary supplements to prevent oxidative stress-induced diseases.Keywords: Antidesma thwaitesianum Müll. Arg., Cytotoxicity, Lung cell carcinoma, Antioxidant activity, Cellular assays

The extraction of active compounds from plants is one of the most critical steps in the commercial development of natural products for medicinal, herbicidal or pesticidal use. The focus of this study was to compare conventional maceration and pressurized liquid extraction (PLE) techniques for the efficient extraction of rotenone from the stem and root of Derris elliptica Benth and Derris malaccensis Prain. The effects of experimental variables, such as solvent, temperature and pressure, on PLE efficiency have been studied. Chloroform was determined to be a good extraction solvent (rotenone content 40.6%, w/w) compared to commonly used solvent, 95% ethanol (rotenone content 15.0%, w/w). The optimal conditions for PLE were 50 degrees C and 2000 psi. PLE showed higher extraction efficiency (rotenone content 46.1%, w/w) as compared with conventional maceration method (rotenone content 40.6%, w/w). The order of rotenone content found in crude extract obtained by optimized method from the highest to the lowest was root (46.1%, w/w) and stem (9.4%, w/w) of D. elliptica and stem of D. malaccensis (5.2%, w/w), respectively. Moreover, the results from this study indicated that PLE was considerably less time and solvent consuming (30 min, 3 ml/g of dried sample) than the conventional maceration techniques (72 h, 10 ml/g of dried sample).

Ethanolic- and water extracts from five species of Thai medicinal plants known as Hua-Khao-Yen were tested for their inhibitory effects against HIV-1 protease (HIV-PR) and HIV-1 integrase (HIV-1 IN). The result revealed that the ethanolic (EtOH) extract of Smilax corbularia exhibited anti-HIV-1 IN activity with an IC 50 value of 1.9 μg/ml, followed by the water extract of Dioscorea birmanica (IC 50 = 4.5 μg/ml), the EtOH extract of Dioscorea birmanica (IC 50 = 4.7 μg/ml), the water extract of Smilax corbularia (IC 50 = 5.4 μg/ml), the EtOH extract of Smilax glabra (IC 50 = 6.7 μg/ml) and the water extract of Smilax glabra (IC 50 = 8.5 μg/ml). The extracts of Pygmaeopremna herbacea and Dioscorea membranacea were apparently inactive (IC 50 > 100 μg/ml). Interestingly, only the EtOH extract of Dioscorea membranacea showed appreciable activity (IC 50 = 48 μg/ml) against HIV-1 PR, while the other extracts possessed mild activity. This result strongly supported the basis for the use of Smilax corbularia and Dioscorea membranacea for AIDS treatment by Thai traditional doctors.

Thai medicine plants used for Osteoarthritis of knee (OA) treatment consist of twelve plants such as Crinumn asiaticum, Cleome viscosa, Drypetes roxburghii, Piper longum, Piper nigrum, Plumbago indica, Alpinia galanga, Curcuma aromatica, Globba malaccensis, Zingiber montanum, Zingiber officinale andZingiberzerumbet. They showedhighfrequency in OA formula.To investigate inhibitory effect on LPS-induced nitric oxide (NO) release from RAW264. 7 cell and free radical scavenging activity usingDPPH assay of these ethanolic plant extracts.Plant materials were extracted by maceration in 95% ethanol. Anti-inflammatory activity were tested on LPS-induced NO production. Free radical scavenging activity was performed by DPPH assay.All of ethanolic extracts exhibited potent inhibitory effect on NO release. The ethanolic extract of Z. zerumbet exhibited the highest inhibitory effect followed by Z. montanum and G. malaccensis, respectively. Except A. galanga and C. viscosa, all extracts possessed more influential than indomethacin (IC50 = 20.32±3.23 μLg/ml), a positive control. The investigation on antioxidant activity suggested that the ethanolic extracts of D. roxburghii, Z. officinale, Z. montanum, C. aromatic, A. galanga, P indica, G malaccensis, P nigrum exhibited antioxidant activity. By means ofD. roxburghii had the highest electron donating activity,followed by Z. officinale. Moreover both extracts were more effective than BHT apositive control (EC50 = 14.04±1.95 μg/ml).Thai medicinal plants had anti-inflammatory activity and could inhibit destruction of articular cartilage that corresponded to the traditional medicine and supported using these medicinal plants for OA treatment.

Benjalokawichien (BLW) or Ya-Ha-Rak (HR) is a traditional remedy in the Nationaldrug list of herbal medicinal products AD 2012 of Thailand. For traditional use, BLW is used as antipyretic agent. It also has anti-allergic effect, particularly treating allergic rash. The ethanolic extract of BLW exhibited anti-allergic activity via inhibitory effect against a release ofbeta-hexosaminidase in RBL-2H3 cell line. Pectolinarigenin has been identified as the active compound ofBLW extract. In this study, a reversed-phase high performance liquid chromatography (RP-HPLC) method was developed in order to control quality ofpreparation in three aspects such as chemical fingerprint, quantification and stability of the ethanolic extract. The RP-HPLC was performed with a gradient mobile phase composed of 0.1% ortho phosphoric acid and acetronitrile, and peaks were detected at 331 nm. Based on validation results, this analytical method is precise, accurate and stable for quantitative determination ofpectolinarigenin. The amount ofpectolinarigenin in Benjalokawichien extract determined by this method was 18.50 mg/g ofextract. Therefore, this method could be consideredfor quality control ofBLWextract.

Immune system is the most important system ofhuman body. Thaifolk doctors have used some medicinal plants as an adaptogenic drug or immunomodulatory agent. Piper chaba Hunter, Piper sarmentosum Roxb. and Piper interruptum Opiz. are used by folk doctors to activate immune response in cancer patients.To investigate the effect on natural killer cell activity and on lymphocyte proliferation activity of water extract of P chaba Hunter P. sarmentosum Roxb. and P interruptum Opiz. MATERIAL ANDMETHOD: Plant materials were extracted by decoction method. All extracts were testedfor an immunomodulatory effect using PBMCs from twelve healthy donors by chromium release assay. Lymphocyte proliferation was also determined by 3H-thymidine uptake assay. The degree of activation was expressed as the stimulation index.The water extract of P chaba Hunter significantly increased lymphocyte proliferation at concentrations ofl ng/ml, 10 ng/ml, 1 μg/ml, 5 μg/ml, 10 μg/ml and 100 μg/ml. P sarmentosum Roxb., and P interruptum Opiz. extracts at those concentrations significantly stimulated lymphocyteproliferation. P sarmentosum Roxb. extractsignificantly increased natural killer (NK) cell activity at a concentration of 100 μg/ml but P chaba Hunter and P interruptum Opiz. extracts did not significantly stimulate natural killer cell activity.P chaba Hunter, P interruptum Opiz. andP sarmentosum Roxb. have an immunomodulatory effect especially for P sarmentosum Roxb. extract which can activate both lymphocyte proliferation and NK cell activity.

Thai traditional remedy called Kabpi (KP) has long been usedfor oral and throat infection. It composed with four herbs Syzygium aromaticum (L.) Merr etPerry, Nigella sativa L., Limonia acidissima L. andAchasma sphaerocephalum Holtt.To investigate antimicrobial activity ofKP remedy extracts and its plant components.The extract procedures were maceration method with 95% ethanol, 50% ethanol and dried by evaporator and boiling in water, filtrated dried by lyophilizer The residue from the maceration in 95% ethanol and 50% ethanol were boiled in water and dried by lyophilizer In the preliminary studied, all extracts were evaluated antimicrobial activity by disc diffusion method. All extracts were tested against one type of gram positive bacteria Staphylococcus aureus (ATCC 25923) and one type ofyeast Candida albicans (ATCC 90028). The active plant extracts were diluted to determine the minimum inhibitory concentration (MIC) values by microtiter plate-based assay.The 95% ethanol extract of KP remedy showed antimicrobial activity against S. aureus (ATCC 25923) and C. albicans (ATCC 90028) (MIC = 0.625, 0.625 mg/ml, respectively). The zone of inhibition ofall extracts were in the range of 7.33 to 23.33 mm. The 95% ethanol extract ofAchasma sphaerocephalum Holtt. showed the highest inhibition zone against S. aureus 21.00 mm (MIC = 0.625 mg/ml). The 95% ethanol extract ofSyzygium aromaticum (L.) Merr etPerry exhibited the best antimicrobial activity against C. albicans the inhibition zone with inhibition zone of23.33 mm (MIC = 0.156 mg/ml).The present study demonstrated that the extracts ofKP remedy and its plant components had an antimicrobial effect against oral and throat infection such as S. aureus and C. albicans. These results support using this remedy of Thai traditional medicine called Kabpifor oral and throat infection.

Sung Yod rice is a red-violet pigmented rice and grown in the southern part of Thailand. Its rice bran oil has attracted the attention ofscientists who have described anti-oxidant properties ofsome ingredients in Sung Yod rice bran oil. Normally, extraction methods ofcommercial product from rice bran oil are by expression or soxhlet extraction with hexane. Thus, biological activities of Sung Yod rice bran oil related to health and chemical content ofrice bran oilfrom the two methods should be studied.The objectives of this research were to investigate for biological activities and chemical content ofSung Yod rice bran oil obtainedfrom expression or soxhlet extraction method.Biological activities such as cytotoxic, anti-inflammatory and antioxidant activities were investigated. Sulphorhodamine (SRB) assay was used to test cytotoxic activity against four human cancer cell lines: lung (COR-L23), cervical (HeLa), prostate (PC-3) and breast (MCF-7) and normal human lung cells (MRC-5). The inhibitory effect on lipopolysaccharide (LPS) induced nitric oxide (NO) production in RA W264. 7 cell lines was usedfor the determination of anti-inflammatory effect. DPPH, TEAC and FRAP assay were carried outfor antioxidant activity. Total phenolic compound was determined by Folin-Ciocalteu reagent. y-oryzanol and vitamin E content were determined by HPLC. Sung Yod rice bran oil was produced by expression method (EX) or by soxhlet extraction method using hexane (SXH-I).The percentage ofyield ofSung Yod rice bran oil by EX and SXH were 2.16 and 15.23 %w/w, respectively. Only EX showed the selective cytotoxicity against prostate cancer cells (PC-3), (IC50 = 52.06±1.60 μg/ml). It also exhibited high inhibitory effects on NO production (IC50 = 30.09 μg/ml). In contrast, SXH had no anti-inflammatory effect and cytotoxic activity against any of the cancer cells. EXshowed higher antioxidant activity determined using DPPH compared to SXH. It also showed higher amount of yoryzanol and vitamin E than that ofSXH (3.09±1.04 and 1.35±1.56 mg % w/w ofextracts, respectively). Yet, SXH exhibited higher antioxidant power determined by FRAP assay and higher total phenolic content compared with EX CONCLUSION: Sung Yod rice bran oil, produced by expression method, had better benefit for health regarding cytotoxicity against prostate cancer cells (PC-3), anti-inflammatory effect and antioxidant (using DPPH) than that produced by soxhlet method extracted with hexane.

Hibiscus sabdariffa (HS) leaves are a vegetable, which is used as a healthy sour soup for protection against chronic diseases in Thai traditional medicine.To investigate antioxidant, anti-inflammatory and cytotoxic activities of Hibiscus sabdariffa leave extracts from diferent extraction methods.Fresh and dry Hibiscus sabdariffa leaves were extracted by various methods such as maceration with 95% and 50% ethanol, squeeze, and boiling with water or decoction. All extracts were testedfor antioxidant activity by using DPPH radical scavenging assay, anti-inflammatory activity by determination on inhibitory effect of nitric oxide production on RAW264. 7 cell. Cytotoxic activity also tested against human prostate cancer cell line (PC-3) by using sulforhodamine B (SRB) assay. Total phenolic content determined by the Folin-Ciocalteu colorimetric method.The results found that the 95% ethanolic extract of Hibiscus sabdariffa dried leaves (HSDE95) showed the highest antioxidant activity with an EC50 of 34.51±2.62 μg/ml and had the highest phenolic content (57.00±3.73 mg GAE/g). HSDE95 also showed potent cytotoxicity against prostate cancer cell line with an IC50 of 8.58±0.68 μg/ml whereas HSDE95 and all of extracts ofHibiscus sabdariffa leaves had no anti-inflammatory activity.The obtained results revealed that HSDE95 extract showedpotent cytotoxic activity against prostate cancer cells but low antioxidant and anti-inflammatory activities. This extract should be further isolated as active compounds against prostate cancer.

Benjakul, a Thai traditional herbal preparation, comnprises five plants: Piper chaba, Piper sarmentosum, Piper interruptum, Plumbago indica, and Zingiber officinale. It has widely been used to treat cancer patients in folk medicine in Thailand. Benjakul extract, and its isolated compounds should be investigated for cytotoxic activity and analysis isolated compounds from chemical fingerprinting.To study cytotoxicity ofBenjakul extract and its isolatedpure compounds against human small cell lung cancer cell line (NCI-HI 688) and in normal human lungfibroblast cell line (MRC-5) and analysis the content ofisolated compounds for quality control of Benjakul extract.Bioassay-guided fractionation was used for isolated active compounds from ethanolic extract of Benjakul. Cytotoxic activity was carried using the SRB assay. HPLC method was applied to analyze six isolated compound contentfrom Benjakul extract.The ethanolic extract ofBenjakul showed cytotoxicity against NCI-H1688 with IC50 value = 36.15±4.35 μg/ml. Hexane fraction as semi-separation by VLC showed the best cytotoxic activity (21.1 7±7.42 μg/ml). Six isolated compounds were identified as myristicin, plumbagin, methyl piperate, 6-shogaol, 6-gingerol and piperine. Plumbagin exhibited the highest cytotoxic activity and 6-shogaol was the second most effective cytotoxic constituent (IC50 values = 1.41±0.01 and 6.45±0.19 μg/ml, respectively). Piperine showed the highest content in both ofHPLC analysis and column chromatography separation.Benjakul extract exhibited cytotoxicity against NCI-HI 688. Plumbagin and 6-shogaol are bioactive markers for cytotoxicity against this small cell lung cancer cell line. Chromatographic fingerprinting can be used to analyze six cytotoxic compounds isolatedfrom the ethanolic extract ofBenjakul.

Prabchompoothaweep remedy (PT) has long been used in Thai traditional medicine to treat allergic rhinitis and asthma. It is composed of 23 plants. It is on National herbal drug list of Thailand, but there is no reportfor anti-allergic, anti-inflammatory and antioxidant activities.To investigate anti-allergic, anti-inflammatory and antioxidant activities of the crude extract from PTby different extraction method.The method of extract used was maceration in 95% ethanol and 50% ethanol; the residue of these extracts were continued extracted by boiling water, they obtained PTE95, PTE50, PTR95 andPTR50, respectively. The other method of extraction was boiling and drying by lyophilizer that obtained PTW Five crude extracts were determined anti- allergic activity by the inhibition of β-hexosaminidase release from RBL-2H3 cell lines, anti-inflammatory activity were determined by the inhibition ofnitric oxide (NO) production from RA W264. 7 cell lines induced by lipopolysaccharide (LPS) and antioxidant activity were tested by DPPH radical scavenging assay.PTE95 showed the most potent ofanti-allergic activity, anti-inflammatory activity and antioxidant activity (IC5 = 12.97, 22.51 and EC50 = 14.62 μg/ml, respectively).These results suggest that the method of extraction PT that showed the best anti-allergy, anti-inflamation and antioxidant activity was maceration in 95% ethanol.

Sa-Tri-Lhung-Klod is a Thai traditional medicine remedy for postpartum in the lists of The National Drug List ofHerbal Medicine Products AD. It consists ofseventeen herbs and were obtained by maceration and used in the form of liquor for women's health care such as treatment ofamenorrhea, menopause and blood tonic. In addition, it also usedfor postpartum care for being anti-inflammation in postpartum and prevention of cancer in women.To investigate cytotoxic activity ofSa-Tri-Lhung-Klod remedy extracts and its herbal ingredients against human ovarian carcinoma cell line (SKOV-3) and cervical adenocarcinoma (HeLa) cell line.Sa-Tri-Lhung-Klod remedy and its plant ingredients were extracted by maceration in 95% ethanol and dried using evaporator. All extracts were testedfor cytotoxic activity by sulforhodamine B (SRB) assay.Ethanolic extract ofSa-Tri-Lhung-Klod remedy displayed cytotoxic activity against SKOV-3 and HeLa with IC50 values of 72.84±1.07 and 47.24±2.83 μg/ml, respectively. It was classified as "less-active" according to the NCI guideline. However, Caesalpinia sappan, Mammea siamensis and Curcuma comosa showed high cytotoxic activity against SKOV-3 with IC50 values of 9.55±1.38 13.45±0.82 and 14.21±1.30 μg/ml, respectively. The ethanolic extracts ofCaesalpiniasappan and Mammea siamensis also exhibited cytotoxic activity against HeLa with IC50 values of 6.30±0.06 and 7.72±0.11 μg/ml, respectively.These results support the use of Sa-Tri-Lhung-Klod remedy in Thai traditional medicine for preventing of ovarian cancer and cervical cancer Caesalpinia sappan, Curcuma comosa and Mammea siamensis were strikingly active against ovarian and cervical cancer cells. Their extracts have the potential for developing as new anti-cancer drugs for women health.