Curcuma longa L. constituents inhibit sortase A and Staphylococcus aureus cell adhesion to fibronectin

The inhibitory activity of Curcuma longa L. (turmeric) rhizome constituents against sortase A, a bacterial surface protein anchoring transpeptidase, from Staphylococcus aureus ATCC 6538p was evaluated. The activity of the isolated compounds (1-4) was compared to that of the positive control,p-hydroxymecuribenzoic acid (pHMB). The biologically active components of C. longa rhizome were characterized by spectroscopic analysis as the curcuminoids curcumin (1), demethoxycurcumin (2), and bisdemethoxycurcumin (3). Curcumin was a potent inhibitor of sortase A, with an IC50 value of 13.8 +/- 0.7 microg/mL. Bisdemethoxycurcumin (IC50 = 31.9 +/- 1.2 microg/mL) and demethoxycurcumin (IC50 = 23.8 +/- 0.6 microg/mL) were more effective than pHMB (IC50 = 40.6 +/- 1.2 microg/mL). The three isolated compounds (1-3) showed no growth inhibitory activity against S. aureus strain Newman, with minimum inhibitory concentrations (MICs) greater than 200 microg/mL. Curcumin also exhibited potent inhibitory activity against S. aureus cell adhesion to fibronectin. The suppression of fibronectin-binding activity by curcumin highlights its potential for the treatment of S. aureus infections via inhibition of sortase activity. These results indicate that curcumin is a possible candidate in the development of a bacterial sortase A inhibitor.     

In vitro activity of the essential oil of Cinnamomum zeylanicum and eugenol in peroxynitrite-induced oxidative processes

The essential oil obtained from the bark of Cinnamomum zeylanicum Blume (Lauraceae) and three of its main components, eugenol, (E)-cinnamaldehyde, and linalool (representing 82.5% of the total composition), were tested in two in vitro models of peroxynitrite-induced nitration and lipid peroxidation. The essential oil and eugenol showed very powerful activities, decreasing 3-nitrotyrosine formation with IC50 values of 18.4 microg/mL and 46.7 microM, respectively (reference compound, ascorbic acid, 71.3 microg/mL and 405.0 microM) and also inhibiting the peroxynitrite-induced lipid peroxidation showing an IC50 of 2.0 microg/mL and 13.1 microM, respectively, against 59.0 microg/mL (235.5 microM) of the reference compound Trolox. On the contrary, (E)-cinnamaldehyde and linalool were completely inactive.     

Antiviral activity of esterified alpha-lactalbumin and beta-lactoglobulin against herpes simplex virus type 1. Comparison with the effect of acyclovir and L-polylysines

The antiviral activity of methylated alpha-lactalbumin (Met-ALA), methylated and ethylated beta-lactoglobulins (Met- and Et-BLG) was evaluated against acyclovir (ACV)-sensitive and -resistant strains of herpes simplex virus type 1 (HSV-1) and compared to that of ACV and L-polylysines (4-15 kDa) using fixed or suspended Vero cell lines. Esterified whey proteins and their peptic hydrolyzates displayed protective action against HSV-1, which was relatively lower than that induced by ACV or L-polylysines. The higher activity of L-polylysines was maintained against an ACV-resistant strain of HSV-1, whereas ACV lost much of its activity. The mean 50% inhibitory concentration (IC50) was about 0.8-0.9 microg/mL for L-polylysines against ACV-sensitive and -resistant strains of HSV-1 when using two concentrations of virus (50% and 100% cytopathic effect, CPE). The IC50 values of ACV against the sensitive strain of HSV-1 were 3 and 15 microg/mL when using the low and high concentrations of virus, respectively. When using 50% CPE, IC50 values for esterified whey proteins ranged from 20 to 95 microg/mL, depending on the nature of the ester group, the degree of esterification, and the nature of the protein. Using the real-time PCR technique, it was shown that Met-ALA inhibited HSV-1 replication.     

Evaluation of the dietetic and therapeutic potential of a high molecular weight hydroxycinnamate-derived polymer from Symphytum asperum Lepech. Regarding its antioxidant, antilipoperoxidant, antiinflammatory, and cytotoxic properties

A water-soluble hydroxycinnamate-derived polymer (>1000 kDa) from Symphytum asperum Lepech. (Boraginaceae) strongly reduced the diphenylpicrylhydrazyl radical (IC(50) approximately 0.7 microg/mL) and inhibited the nonenzymatic lipid peroxidation of bovine brain extracts (IC(50) approximately 10 ng). This polymer exhibited only a low hydroxyl radical scavenging effect in the Fe(3+)-EDTA-H(2)O(2) deoxyribose system (IC(50) > 100 microg/mL) but strongly decreased superoxide anion generation in either the reaction of phenazine methosulfate with NADH and molecular oxygen (IC(50) approximately 13.4 microg/mL) or in rat PMA-activated leukocytes (IC(50) approximately 5 microg/mL). The ability to inhibit both degranulation of azurophil granules and superoxide generation in primed leukocytes indicates that the NADPH oxidase responsible for this later effect is inhibited, pointing to the Symphytum asperum polymer as a potent antiinflammatory and vasoprotective agent. At all concentrations tested (0-200 microg/mL), we observed no cytotoxicity on normal human fibroblasts and neither antiproliferative effects nor proliferation activation on neoplastic cells.     

Effects of grape cell culture extracts on human topoisomerase II catalytic activity and characterization of active fractions

Grape and its cell culture extracts are rich in flavonoids and stilbenes that are biologically active. The objective of this study was to evaluate possible inhibitory effects of grape (a Vitis hybrid Bailey Alicant A) cell culture extract and subfractions on human DNA topoisomerase II catalytic activity and to characterize constituents in the most potent fractions. At 5 microg/mL, grape cell crude extract and Toyopearl (TP) fractions 2-6 provided significantly greater inhibition of topoisomerase II catalytic activity than quercetin, a chemopreventive agent previously known as a topoisomerase catalytic inhibitor. The most potent topoisomerase II catalytic inhibitors from grape cell culture extracts in descending order of potency were TP fractions 4 and 6 (IC(50) = 0.28-0.29 microg/mL), TP-3 (IC(50) = 0.74 microg/mL), and crude extract (IC(50) = 1.02 microg/mL); each was significantly more potent than resveratrol (IC(50) = 18.0 microg/mL), another well-known chemopreventive topoisomerase II catalytic inhibitor. Using both high-performance liquid chromatography and liquid chromatography electrospray ionization mass spectrometry, constituents in TP-4 and TP-6 were characterized. These constituents included cyanidin-3,5-diglucoside, malvidin-3-acetylglucoside, peonidin-3-coumaryl-5-diglucoside, procyanidin B(1), procyanidin B(2), procyanidin B(5), procyanidin dimer digallate, procyanidin C(1), myricetin, and rutin, none of which have been previously characterized from grape cell cultures. The significant potency especially of TP-4 and TP-6 from grape cell cultures suggests that these fractions may have potential as chemopreventive agents.     

Inhibition of gastric H(+),K(+)-ATPase and Helicobacter pylori growth by phenolic antioxidants of Curcuma amada

Gastric ulcer is the most prevalent gastrointestinal disorder, resulting from oxidative stress, Helicobacter pylori infection, up-regulation of proton potassium ATPase (PPA) activity, down-regulation of gastric mucosal defense, etc. In this paper it is reported that phenolic fractions of Curcuma amada, commonly known as mango ginger, acted as potent inhibitors of PPA and H. pylori growth. Mango ginger free phenolics (MGFP) and mango ginger bound phenolics (MGBP) inhibited PPA at IC50 values of 2.2 +/- 0.21 and 0.7 +/- 0.08 microg/mL, respectively, exhibiting 9-27-fold better potency over lansoprazole (IC(50) of 19.3 +/- 2.2 microg/mL). MGFP is constituted by caffeic (26%), gentisic (24%), ferulic (20%), gallic (10%), cinnamic (7%), and protocatechuic acids (7%) and MGBP by ferulic (47%), cinnamic (29%), p-coumaric acid (11%), and syringic (5%) acids as major phenolic acids. MGFP and MGBP further exhibited free radical scavenging (IC(50) of 2.2 +/- 0.17 and 4.2 +/- 0.36 microg/mL), reducing power abilities (193-104 units/g), inhibition of lipid peroxidation (IC(50) of 10.3 +/- 0.91 and 15.6 +/- 1.6 microg/mL), and DNA protection (80% at 4 microg), indicating strong antioxidative properties. MGFP and MGBP thus may be potential and inexpensive multistep blockers against ulcers.     

Investigations into inhibitor type and mode, simulated gastrointestinal digestion, and cell transport of the angiotensin I-converting enzyme-inhibitory peptides in Pacific hake (Merluccius productus) fillet hydrolysate

Fish protein hydrolysate (FPH) produced by incubation of Pacific hake fillet with 3.00% Protamex at pH 6.5 and 40 degrees C for 125 min demonstrated in vitro ACE-inhibitory activity (IC50 = 165 microg/mL), which was enhanced by ultrafiltration through a 10 kDa molecular weight cutoff membrane (IC50 = 44 microg/mL). However, after simulated gastrointestinal digestion, FPH and ultrafiltrate had similar ACE-inhibitory activity (IC 50 = 90 microg/mL), indicating that FPH peptides act as "pro-drug type" inhibitors and that enrichment by ultrafiltration may be unnecessary. Matrix-assisted laser desorption/ionization-time of flight mass spectrometry confirmed that the molecular weights of major peaks were <1 kDa regardless of ultrafiltration. ACE-inhibitory activities of digested hydrolysates were not significantly affected by preincubation with ACE ( P > 0.05) and exhibited a competitive inhibitory mode. A permeability assay using fully differentiated colorectal adenocarcinoma (Caco-2) cells showed an apical to basolateral transport of peptides that ranged from approximately 2 to 20% after 2 h at 37 degrees C. Pacific hake fillet hydrolysates are a potentially bioavailable source of ACE-inhibitory peptides awaiting further in vivo study.     

Effect of yerba mate (Ilex paraguariensis) tea on topoisomerase inhibition and oral carcinoma cell proliferation

Tea flavonoids have antitopoisomerase activity and can inhibit cell proliferation. The objectives of this study were to determine the phenolic content of yerba mate tea products (MT) (Ilex paraguariensis) and evaluate their capacity to inhibit topoisomerase I (Topo I) and II (Topo II) activities and oral carcinoma cell proliferation. Total polyphenols of aqueous extracts of dried MT leaves were measured by the Folin-Ciocalteau assay, using chlorogenic (CH) and gallic (GA) acids as standards. Topoisomerase inhibition was determined by a clone-forming assay, which uses yeast (Saccharomyces cerevisiae) strains as a model. Controls included dimethyl sulfoxide (1.66%); camptothecin (50 microg/mL), a Topo I inhibitor; and amsacrine (100 microg/mL), a Topo II inhibitor. Cytotoxicity studies were conducted using a nontumorigenic human keratinocyte cell line HaCaT and two human squamous cancer cell lines (SCC-61 and OSCC-3). MT was found to be a rich source of phenolic compounds. Total polyphenol content of various commercially available traditional MT products ranged from 236 to 490 mg equiv of CH/g of dry leaves. Such levels were significantly different among products depending on their origin (P < 0.001). Higher anti-topoisomerase II activity was observed against JN394t(2-4) strain for Nobleza Gaucha MT (IC50 = 0.43 microg equiv of CH) in comparison to GA (IC50 = 112 mM) and CH (IC50 > 1500 mM). MT showed catalytic anti-topoisomerase activity against Topo II but not against Topo I. In addititon, MT exhibited dose-dependent cytotoxicity against all squamous cell lines tested. In comparison to premalignant cell line HaCaT [28 microg equiv of (+)-catechin mL(-1)], the cell line SCC-61 [21 microg equiv of (+)-catechin mL(-1)] was the most sensitive to MT, resulting in 50% inhibition of net cell growth. It is concluded that MT is rich in phenolic constituents and can also inhibit oral cancer proliferation. The effect on cancer cell proliferation may be mediated via inhibition of topoisomerase II. The lack of correlation between polyphenol content and the inhibition of topoisomerases suggests that the effect of MT on topoisomerase inhibition may be due to other still unidentified biologically active phytochemicals constituents.     

Evaluation of antioxidant activity and preventing DNA damage effect of pomegranate extracts by chemiluminescence method

The antioxidant activities of three parts (peel, juice, and seed) and extracts of three pomegranate varieties in China were investigated by using a chemiluminescence (CL) method in vitro. The scavenging ability of pomegranate extracts (PEs) on superoxide anion, hydroxide radical, and hydrogen peroxide was determined by the pyrogallol-luminol system, the CuSO4-Phen-Vc-H2O2 system, and the luminol-H2O2 system, respectively. DNA damage preventing the effect of PE was determined by the CuSO4-Phen-Vc-H2O2-DNA CL system. The results showed that the peel extract of red pomegranate had the best effect on the scavenging ability of superoxide anion because its IC50 value (4.01 +/- 0.09 microg/mL) was the lowest in all PEs. The seed extract of white pomegranate could scavenge hydroxide radical most effectively of the nine extracts (the IC50 value was 1.69 +/- 0.03 microg/mL). The peel extract of white pomegranate had the best scavenging ability on hydrogen peroxide, which had the lowest IC50 value (0.032 +/- 0.003 microg/mL) in the nine extracts. The seed extract of white pomegranate (the IC50 value was 3.67 +/- 0.03 microg/mL) was the most powerful on the DNA damage-preventing effect in all of the PEs. Also, the statistical analysis indicated that there were significant differences (at P < 0.05) among the extracts of the different varieties and parts in each system.     

In vitro anti-inflammatory and anti-proliferative activity of sulfolipids from the red alga Porphyridium cruentum

A sulfoglycolipidic fraction (SF) isolated from the red microalga Porphyridium cruentum was analyzed for fatty acid composition and assayed for ability to inhibit, in vitro, the generation of superoxide anion in primed leucocytes and the proliferation of a panel of human cancer cell-lines. Results demonstrated that SF contained large amounts of palmitic acid (26.1%), arachidonic acid (C20: 4 omega-6, 36.8%), and eicopentaenoic (C20:5 omega-3, 16.6%) acids, and noticeable amounts of 16:1n-9 fatty acid (10.5%). It strongly inhibited both the production of superoxide anion generated by peritoneal leukocytes primed with phorbol myristate acetate (IC(50): 29.5 microg/mL), and the growth of human colon adenocarcinoma DLD-1 and to a lesser extent of human breast adenocarcinoma MCF-7, human prostate adenocarcinoma PC-3, and human malignant melanoma M4 Beu cell-lines, and therefore might have a chemopreventive or chemotherapeutic potential, or both. It was found markedly more cytotoxic than sulfoquinovosyldiacylglycerols from plant used as a standard (STD), due to a stronger ability to inhibit DNA alpha-polymerase (IC(50): 378 microg/mL, vs 1784 microg/mL for STD). After a 48-h continuous treatment, IC(50) values for growth inhibition were in the range of 20-46 microg/mL instead of 94 to >250 microg/mL for STD, and those for inhibition of metabolic activity were in the range of 34-87 microg/mL instead of >250 microg/mL for STD. The higher anti-proliferative effect was observed on colon adenocarcinoma DLD-1 cells, and the weaker effect was observed on breast adenocarcinoma MCF-7.     

Chemical composition and antifungal activity of essential oils from different tissues of Japanese Cedar (Cryptomeria japonica)

In this study antifungal activities of essential oils from different tissues of Japanese cedar (Cryptomeria japonica D. Don) against four wood decay fungi and six tree pathogenic fungi were investigated. In addition, the yields of essential oils obtained by water distillation were compared and their constituents determined by GC-MS analyses. The yield of essential oils from four tissues of Japanese cedar is in the decreasing order of leaf (27.38 mL/kg) > bark (6.31 mL/kg) > heartwood (3.80 mL/kg) > sapwood (1.27 mL/kg). Results obtained from the antifungal tests demonstrate that the essential oil of Japanese cedar heartwood used against Laetiporus sulphureus and Trametes versicolor and sapwood essential oil used against L. sulphureus had strong antifungal activities at 500 mug/mL, with IC(50) values of 39, 91, and 94 microg/mL, respectively. Besides, the essential oils of Japanese cedar heartwood used against Rhizoctonia solani, Collectotrichum gloeosporioides, Fusarium solani, and Ganoderma australe had strong antifungal activities at 500 microg/mL, with IC(50) values of 65, 80, 80, and 110 microg/mL, respectively. GC-MS analyses showed that the sesquiterpene hydrocarbon compounds dominate in the essential oil from Japanese cedar heartwood, amounting to a total percentage of 82.56%, with the major compounds of delta-cadinene (18.60%), isoledene (12.41%), and gamma-muurolene (11.82%). It is proposed that the excellent antifungal activities of Japanese cedar heartwood essential oils might correlate with the presence of these compounds.     

Venturia inaequalis-inhibiting Diels-Alder adducts from Morus root bark

In organic apple orcharding there is a continuous need for natural fungicides effective against Venturia inaequalis (Cooke) Winter, the causal agent of apple scab. In this study an in vitro assay is presented for determining the germination inhibitory potential of extracts and pure compounds. From a screening of plant extracts, the methanol extract of Morus root bark revealed distinct V. inaequalis inhibiting qualities, which were subjected to a bioguided fractionation. Among the isolated metabolites [moracins M (1), O/P (2), kuwanon L (3), and sanggenons D (4), B (5), G (6), O (7), E (8), and C (9)] all the Diels-Alder adducts (3-9) showed an antifungal activity with IC50 values between 10 and 123 microM. The in vitro activity of the most active fraction (A5, IC50 39.0 +/- 4.2 microg/mL) was evaluated in vivo, confirming a distinct antifungal activity against V. inaequalis for the tested natural material.     

Antioxidative and antiplatelet effects of aqueous inflorescence Piper betle extract

Piper betle, belonging to the Piperaceae family, is a tropical plant, and its leaf and inflorescence are popularly consumed by betel quid (BQ) chewers in Taiwan and many other South and Southeast Asian countries. However, little is known about the biochemical properties of inflorescence Piper betle (IPB) toward reactive oxygen species (ROS) and platelet functions. In the present work, aqueous IPB extract was shown to be a scavenger of H(2)O(2), superoxide radical, and hydroxyl radical with a 50% inhibitory concentration (IC(50)) of about 80, 28, and 73 microg/mL, respectively. IPB extract also prevented the hydroxyl radical induced PUC18 plasmid DNA breaks at concentrations higher than 40 microg/mL. Since ROS are crucial for platelet aggregation, we further found that IPB extract also inhibited the arachidonic acid (AA) induced and collagen-induced platelet aggregation, with an IC(50) of 207 and 335 microg/mL, respectively. IPB extract also inhibited the AA-, collagen- (>100 microg/mL of IPB), and thrombin (>250 microg/mL of IPB)-induced thromboxane B(2) (TXB(2)) production by more than 90%. However, IPB extract showed little effect on thrombin-induced aggregation. These results indicated that aqueous components of IPB are potential ROS scavengers and may prevent the platelet aggregation possibly via scavenging ROS or inhibition of TXB(2) production.     

Antimicrobial and antioxidant activities of Melissa officinalis L. (Lamiaceae) essential oil

The present study describes antimicrobial and free radical scavenging capacity (RSC) together with the effects on lipid peroxidation (LP) of Melissa officinalis essential oil. The chemical profile of essential oil was evaluated by the means of gas chromatography-mass spectrometry (GC-MS) and thin-layer chromatography (TLC). RSC was assessed measuring the scavenging activity of essential oil on the 2,2-diphenyl-1-picrylhydrazyl (DPPH(*)) and OH(*) radicals. The effect on LP was evaluated following the activities on Fe(2+)/ascorbate and Fe(2+)/H(2)O(2) systems of induction. The antimicrobial activity was tested against 13 bacterial strains and six fungi. The examined essential oil exhibited very strong RSC, reducing the DPPH radical formation (IC(50) = 7.58 microg/mL) and OH radical generation (IC(50) = 1.74 microg/mL) in a dose-dependent manner. According to the GC-MS and TLC (dot-blot techniques), the most powerful scavenging compounds were monoterpene aldehydes and ketones (neral/geranial, citronellal, isomenthone, and menthone) and mono- and sesquiterpene hydrocarbons (E-caryophyllene). Very strong inhibition of LP, particularly in the Fe(2+)/H(2)O(2) system of induction (94.59% for 2.13 microg/mL), was observed in both cases, also in a dose-dependent manner. The most effective antibacterial activity was expressed on a multiresistant strain of Shigella sonei. A significant rate of antifungal activity was exhibited on Trichophyton species.     

Black soybean promotes the formation of active components with antihepatoma activity in the fermentation product of Agaricus blazei

The antihepatoma activity and related active components in the fermentation products of Agaricus blazei (AB) cultured in the medium containing soybean (S) or black soybean (BS) were investigated. AB(BS)-pE and AB(S)-pE were the ethanolic extracts from the fermentation products of AB(BS) and AB(S), respectively. According to the IC 50 values, AB(BS)-pE (161.1 and 24.0 microg/mL for Hep 3B and Hep G2 cells, respectively) exhibited stronger cytotoxicities against hepatoma cells than AB(S)-pE (>200 and 99.9 microg/mL for Hep 3B and Hep G2 cells, respectively). AB(BS)-pE was separated by silica gel column chromatography and eluted with n-hexane/ethyl acetate/methanol gradient solvent system into 21 fractions. Fraction 3 [AB(BS)-pE-F3], eluted with n-hexane/ethyl acetate (97:3 and 19:1, v/v), was the most active fraction having inhibitory activity on the proliferation of Hep 3B and Hep G2 cells (IC 50 of 3.6 and 1.9 microg/mL, respectively). Three major compounds, compounds 1- 3, were further isolated from the AB(BS)-pE-F3 fraction by reversed-phase semipreparative high-performance liquid chromatography. Compounds 2 and 3 gave better antihepatoma activity than that of compound 1. The IC 50 values of compounds 2 and 3 were 2.8 and 4.5 microg/mL for Hep 3B cells and 1.4 and 2.0 microg/mL for Hep G2 cells, respectively. The structures of compounds 2 and 3 were identified by UV, IR, electron impact mass spectrometry, and (1)H and (13)C NMR to be blazeispirols A and C, respectively. Blazeispirols A and C existed in the mycelia but not in the broth and were more in AB(BS)-pE (49.9 +/- 8.9 and 14.2 +/- 2.4 mg/g, respectively) than AB(S)-pE (15.9 +/- 1.7 and 3.9 +/- 0.6 mg/g, respectively). Additionally, the result shows that the production of blazeispirols A and C was increased after cultivation in the medium containing black soybean on day 6 and reached the maximum on day 12, and the contents of blazeispirols A and C were negatively correlated with Hep 3B and Hep G2 cell viabilities ( r = -0.84 to -0.93, P < 0.01). It suggests that blazeispirols A and C could be used as biomarkers to produce the fermentation product of A. blazei with antihepatoma activity.     

Bioactive metabolites from the fungus Nectria galligena, the main apple canker agent in Chile

The phytopathogenic fungus Nectria galligena Bres. is the most common canker disease agent of hardwood trees. The terpenoids colletochlorin B, colletorin B, ilicicolin C, E, and F, as well as the phytotoxin alpha,beta-dehydrocurvularin have been isolated from liquid cultures of N. galligena obtained from the xylem of infected apple trees in central Chile. Ilicicolin C and F and alpha,beta-dehydrocurvularin were active against Pseudomonas syringae with IC50 values of 28.5, 28.5, and 14.2 microg/mL, respectively, in the same range as streptomycin and penicillin G (11 and 15 microg/mL, respectively). All of the compounds showed moderate inhibitory activity toward the enzymes acetylcholinesterase (AChE) and beta-glucuronidase. The most active enzyme inhibitors were colletochlorin B and ilicicolin C and E, with IC50 values of 30-36 microg/mL in the AChE assay and 32-43 microg/mL in the beta-glucuronidase test. All of the chlorinated compounds showed some toxicity toward human lung fibroblasts, with IC50 values in the range of 64-120 microg/mL. alpha,beta-Dehydrocurvularin proved to be the most toxic compound, showing IC50 values less than 12 microg/mL. The effect of the isolated compounds on seed germination and radicle and epicotyl growth was assessed in lettuce and millet seeds. At 100 and 200 microg/disk, alpha,beta-dehydrocurvularin significantly reduced radicle length and epicotyl growth in Lactuca sativa. This is the first report on the occurrence of colletochlorin B, colletorin B, ilicicolin C, E, and F, as well as alpha,beta-dehydrocurvularin associated to N. galligena.     

Development of an enzyme-linked immunosorbent assay for the detection of glyphosate

A competitive indirect enzyme-linked immunosorbent assay (CI-ELISA) was developed to quantitate the herbicide glyphosate [N-(phosphonomethyl)glycine] in water. The ELISA has a detection limit of 7.6 microg mL(-1) and a linear working range of 10-1000 microg mL(-1) with an IC(50) value of 154 microg mL(-1). The glyphosate polyclonal antisera did not cross-react with a number of other herbicides tested but did cross-react with the glyphosate metabolite aminomethylphosphonic acid and a structurally related herbicide, glyphosine [(N,N-bis(phosphonomethyl)glycine]. The assay was used to estimate, quantitatively with accuracy and precision, glyphosate concentrations in water samples. Water samples were analyzed directly, and no sample preparation was required. To improve detection limits, water samples were concentrated prior to analysis, resulting in the increase of the detection limits by 100-fold. After the sample preconcentration step, the detection limit improved to 0.076 microg mL(-1) with an IC(50) value of 1.54 microg mL(-1), and a linear working range was 0.1-10 microg mL(-1). Glyphosate concentrations determined by ELISA correlated well with those determined by high-pressure liquid chromatography (r(2) = 0.99). This assay contributes to reducing the costs associated with conventional residue analysis techniques for the quantitation of glyphosate in water.     

Inhibitory effects of oolong tea polyphenols on pancreatic lipase in vitro

Fifty-four polyphenols isolated from tea leaves were evaluated for their inhibitory activities against pancreatic lipase, the key enzyme of lipid absorption in the gut. (-)-Epigallocatechin 3-O-gallate (EGCG), which is one of major polyphenols in green tea, showed lipase inhibition with an IC50 of 0.349 microM. Moreover, flavan-3-ol digallate esters, such as (-)-epigallocatechin-3,5-digallate, showed higher activities of inhibition on lipase with an IC50 of 0.098 microM. On the other hand, nonesterified flavan-3-ols, such as (+)-catechin, (-)-epicatechin, (+)-gallocatechin, and (-)-epigallocatechin, showed zero and/or the lowest activities against pancreatic lipase (IC50 > 20 microM). These data suggested that the presence of galloyl moieties within the structure was required for enhancement of pancreatic lipase inhibition. It is well-known that flavan-3-ols are polymerized by polyphenol oxidase and/or heating in a manufacturing process of oolong tea. Oolonghomobisflavans A and B and oolongtheanin 3'-O-gallate, which are typical in oolong tea leaves, showed strong inhibitory activities with IC50 values of 0.048, 0.108, and 0.068 microM, respectively, even higher than that of EGCG. The oolong tea polymerized polyphenols (OTPP) were prepared for the assay from oolong tea extract, from which the preparation effectively subtracted the zero and/or less-active monomeric flavan-3-ols by preparative high-performance liquid chromatography. The weight-average molecular weight (Mw) and number-average molecular-weight (Mn) values of OTPP were 2017 and 903, respectively, by using gel permeation choromatography. OTPP showed a 5-fold stronger inhibition against pancreatic lipase (IC50 = 0.28 microg/mL) by comparison with that of the tannase-treated OTPP (IC50 = 1.38 microg/mL). These data suggested that the presence of galloyl moieties within their chemical structures and/or the polymerization of flavan-3-ols were required for enhancement of pancreatic lipase inhibition.     

Tumor cell proliferation and cyclooxygenase enzyme inhibitory compounds in Amaranthus tricolor

Amaranthus tricolor is consumed as a vegetable in Asia. Bioassay-directed isolation of leaves and stems of A. tricolor yielded three galactosyl diacylglycerols (1-3) with potent cyclooxygenase and human tumor cell growth inhibitory activities. The purified compounds were characterized by spectroscopic methods. In addition, the fatty acid moieties in diacyl galactosyl glyerols were characterized by GC-MS analyses. The galactosyl diacylglycerols 1-3 inhibited the cyclooxygenase-1 (COX-1) enzyme by 78, 63, and 93% and the cyclooxygenase-2 (COX-2) enzyme by 87, 74, and 95%, respectively. These compounds were tested for antiproliferative activity using human AGS (gastric), CNS (central nervous system; SF-268), HCT-116 (colon), NCI-H460 (lung), and MCF-7 (breast) cancer cell lines. Compound 1 inhibited the growth of AGS, SF-268, HCT-116, NCI-H460, and MCF-7 tumor cell lines with IC50 values of 49.1, 71.8, 42.8, 62.5, and 39.2 mug/mL, respectively. For AGS, HCT-116, and MCF-7 tumor cell lines, the IC50 values of compounds 2 and 3 were 74.3, 71.3, and 58.7 microg/mL and 83.4, 73.1, and 85.4, respectively. This is the first report of the COX enzyme inhibitory activity for galactosyl glycerols and antiproliferative activities against human colon, breast, lung, stomach, and CNS tumor cell lines.