Surface plasmon resonance and cytotoxicity assays of drug efficacies predicted computationally to inhibit p53/MDM2 interaction.

Docking on the p53-binding site of murine double minute 2 (MDM2) by small molecules restores p53's tumor-suppressor function. We previously assessed 3244 FDA-approved drugs via "computational conformer selection" for inhibiting MDM2 and p53 interaction. Here, we developed a surface plasmon resonance method to experimentally confirm the inhibitory effects of the known MDM2 inhibitor, nutlin-3a, and two drug candidates predicted by our computational method. This p53/MDM2 interaction displayed a dosage-dependent weakening when MDM2 is pre-mixed with drug candidates. The inhibition efficiency order is nutlin-3a (IC50 = 97 nM) > bepridil (206 nM) > azelastine (307 nM). Furthermore, we verified their anti-proliferation effects on SJSA-1 (wild-type p53 and overexpressed MDM2), SW480 (mutated p53), and SaOs-2 (deleted p53) cancer cell lines. The inhibitory order towards SJSA-1 cell line is nutlin-3a (IC50 = 0.8 µM) > bepridil (23 µM) > azelastine (25 µM). Our experimental results are in line with the computational prediction, and the higher IC50 values from the cell-based assays are due to the requirement of higher drug concentrations to penetrate cell membranes. The anti-proliferation effects of bepridil and azelastine on the cell lines with mutated and deleted p53 implied some p53-independent anti-proliferation effects.

Role of the activation gate in determining the extracellular potassium dependency of block of HERG by trapped drugs.

Drug induced long QT syndrome (diLQTS) results primarily from block of the cardiac potassium channel HERG (human-ether-a-go-go related gene). In some cases long QT syndrome can result in the lethal arrhythmia torsade de pointes, an arrhythmia characterized by a rapid heart rate and severely compromised cardiac output. Many patients requiring medication present with serum potassium abnormalities due to a variety of conditions including gastrointestinal dysfunction, renal and endocrine disorders, diuretic use, and aging. Extracellular potassium influences HERG channel inactivation and can alter block of HERG by some drugs. However, block of HERG by a number of drugs is not sensitive to extracellular potassium. In this study, we show that block of WT HERG by bepridil and terfenadine, two drugs previously shown to be trapped inside the HERG channel after the channel closes, is insensitive to extracellular potassium over the range of 0 mM to 20 mM. We also show that bepridil block of the HERG mutant D540K, a mutant channel that is unable to trap drugs, is dependent on extracellular potassium, correlates with the permeant ion, and is independent of HERG inactivation. These results suggest that the lack of extracellular potassium dependency of block of HERG by some drugs may in part be related to the ability of these drugs to be trapped inside the channel after the channel closes.

A computational and experimental approach to investigate bepridil binding with cardiac troponin.

Cardiac troponin is a Ca(2+)-dependent switch for the contraction in heart muscle and a potential target for drugs in the therapy of heart failure. Bepridil is a drug that binds to troponin and increases calcium sensitivity of muscle contraction. Because bepridil has been well studied, it is a good model for analysis by computational and experimental methods. Molecular dynamics (MD) simulations were performed on troponin complexes of different sizes in the presence and absence of bepridil bound within the hydrophobic pocket at the N-terminal domain of troponin C. About 100 ns of simulation trajectory data were generated, which were analyzed using cross-correlation analyses and MMPBSA and MMGBSA techniques. The results indicated that bepridil binding within the hydrophobic pocket of cardiac TnC decreases the interaction of TnC with TnI at both the N-domain of TnC and the C-domain of TnC, and decreases the correlations of motions among the segments of the troponin subunits. The estimated calcium-binding affinities using MMPBSA showed that bepridil has a sensitizing effect for the isolated system of TnC, but loses this effect for the complex. Our experimental measurements of calcium dissociation rates were consistent with that prediction. We also observed that while bepridil enhanced the troponin-tropomyosin-actin-activated ATPase activity of myosin S1 at low calcium concentrations it was slightly inhibitory at high calcium concentrations. Bepridil increases the ATPase activity and force generation in muscle fibers, but its effects appear to depend on the concentration of calcium.

Possible role of calcium ions, calcium channels and calmodulin in excystation and metacystic development of Entamoeba invadens.

The effect of calcium ions (Ca(2+)) and calmodulin (CaM) on the excystation and metacystic development of Entamoeba invadens was examined by transfer of cysts to a growth medium containing calcium antagonists and CaM inhibitors. Excystation, which was assessed by counting the number of metacystic amoebae after induction of excystation, was inhibited by the calcium chelators ethyleneglycol bis (beta-aminoethyl ether)- N,N'-tetraacetate (EGTA) and ethylene-diaminetetraacetate (EDTA), with EDTA being more potent than EGTA. The inhibitory effect of higher concentrations of these chelators on excystation was associated with reduced viability of cysts. Metacystic development, when determined by the number of nuclei in an amoeba, was delayed by EGTA, because the percentage of four-nucleate amoebae was higher than in controls at day 3 of incubation. EDTA made metacystic development unusual by producing a large number of metacystic amoebae with more than ten nuclei. The inhibition of excystation by these chelators was partially abrogated by their removal. A putative antagonist of intracellular calcium flux, 8-( N,N-diethylamino) octyl-3,4,5-trimethoxybenzoate (TMB-8) also inhibited the excystation and metacystic development, but had little effect on cyst viability. The slow Na(+)-Ca(2+) channel blocker bepridil but not verapamil inhibited the excystation and metacystic development, associating with reduced cyst viability at higher concentrations. The inhibitory effect of bepridil on excystation was abrogated by removal of the drug. The CaM inhibitor trifIuoperazine (TFP) but not W-7 [ N-(6-aminohexyl)-chloro-1-naphtalene sulphonamide] inhibited the excystation and metacystic development. The inhibitory effect of TFP on excystation was also abrogated by removal of the drug. These results indicate that extracellular calcium ions, amoebic intracellular calcium flux, calcium channels, and a CaM-dependent process contribute to the excystation and metacystic development of E. invadens.

Antioxidant effects of 1,5-anhydro-D-fructose, a new natural sugar, in vitro.

The antioxidant effects of 1,5-anhydro-D-fructose (1,5-AF), a unique anhydrohexulose, were studied in 1,1-diphenyl-2-picrylhydrazyl (DPPH) solution, in human cells along with lipid peroxidation of low-density lipoprotein (LDL). We have confirmed that 1,5-AF scavenges DPPH radicals directly in solution and inhibits the formation of hydrogen peroxide and superoxide anion, typical reactive oxygen species (ROS), induced by phorbol myristate acetate (PMA) in a dose-dependent manner in THP-1 cells. We also observed the dose-dependent antioxidant effects of 1,5-AF on copper-mediated LDL oxidation. These findings suggest that 1,5-AF might play a role in reducing the risk of atherosclerosis and may help prevent coronary heart disease.

Antioxidant capacities of ten edible North American plants.

The EtOAc extract obtained from ten edible North American plants, Acorus calamus, Clintonia borealis, Gaultheria shallon, Juniperus osteosperma, Opuntia polyacantha, Prunus americana, Prunus virginiana, Sambucus cerulea, Sorbus americana and Vaccinium parvifolium, were tested in the 1,1-diphenyl-2-picrylhydrazyl (DPPH) free radical assay. High antioxidant activity was obtained from the extracts of three fruits, Gaultheria shallon, Sambucus cerulea and Prunus americana and one extracted rhizome, Acorus calamus. Catechin and epicatechin, potent polyphenolic antioxidants, were identified in the EtOAc extracts of Gaultheria shallon and Sambucus cerulea by reversed-phase thin-layer chromatography (TLC) and reversed-phase high-performance liquid chromatography (HPLC).

Antioxidant enzymes and DPPH-radical scavenging activity in chilled and heat-shocked rice (Oryza sativa L.) seedlings radicles.

Chilling whole rice seedlings at 5 degrees C significantly increased the time needed to recover linear growth and reduced the subsequent linear rate of radicle growth. Subjecting nonchilled seedlings to a 45 degrees C heat shock for up to 20 min did not alter subsequent growth, whereas a 3 min heat shock was optimal in reducing growth inhibition caused by 2 days of chilling. The activity of five antioxidant enzymes [superoxide dismutase (EC 1.15.1.1), catalase (CAT; EC 1.11.1.6), ascorbate peroxidase (APX; EC 1.11.1.11), glutathione reductase (GR; EC 1.6.4.2), and guaiacol peroxidase (GPX; EC 1.11.1.7)] and DPPH (1,1-diphenyl-2-picrylhydrazyl)-radical scavenging activity were measured in heat-shocked and/or chilled radicles. Heat shock slightly increased the activity of CAT, APX, and GR and suppressed the increase of GR and GPX activity during recovery from chilling. Increased CAT, APX, GR, and DPPH-radical scavenging activity and protection of CAT activity during chilling appear to be correlated with heat shock-induced chilling tolerance.

Cyclooxygenase-inhibitory and antioxidant constituents of the aerial parts of Antirhea acutata.

Two new compounds, (6S)-hydroxy-29-nor-3,4-seco-cycloart-4(30),24-dien-3-oic acid (1) and 8-[1-(3,4-dihydroxyphenyl)-3-methoxy-3-oxopropyl]epicatechin (3), were isolated by bioassay-guided fractionation from the aerial parts of Antirhea acutata (DC.) Urb. (Rubiaceae). Compound 1 showed moderate inhibitory activities in cyclooxygenase-1 and -2 assays (IC(50) 43.7 and 4.7 microM, respectively), while compound 3 was active in 1,1-diphenyl-2-picrylhydrazyl free-radical and cytochrome c reduction antioxidant assays (IC(50) 29.1 and 16.3 microM, respectively). Additionally, one further new compound was isolated, (3S,24S)-25-trihydroxy-9,19-cycloartane-29-oic acid (2), but this was inactive in the bioassay systems used. Compound 1 is based on the unprecedented 29-nor-3,4-seco-cycloartane skeleton.

Broussochalcone A, a potent antioxidant and effective suppressor of inducible nitric oxide synthase in lipopolysaccharide-activated macrophages.

The antioxidant properties of broussochalcone A (BCA) and its effects on nitric oxide (NO) production in lipopolysaccharide (LPS)-activated macrophages were investigated in this study. BCA, isolated from Broussonetia papyrifera Vent., inhibited iron-induced lipid peroxidation in rat brain homogenate in a concentration-dependent manner with an IC(50) of 0.63 +/- 0.03 microM. It was as potent as butylated hydroxytoluene, a common antioxidant used for food preservation. In a diphenyl-2-picrylhydrazyl assay system, the radical-scavenging activity of BCA seemed to be more potent than that of alpha-tocopherol, its IC(0.200) being 7.6 +/- 0.8 microM. BCA could directly scavenge superoxide anion and hydroxyl radicals. These results indicated that BCA was a powerful antioxidant with versatile free radical-scavenging activity. On the other hand, we found that BCA suppressed NO production concentration-dependently, with an IC(50) of 11.3 microM in LPS-activated macrophages. This effect was not the consequence of a direct inhibitory action on the enzyme activity of inducible NO synthase (iNOS). Our results indicated that BCA exerts potent inhibitory effects on NO production, apparently mediated by its suppression of IkappaBalpha phosphorylation, IkappaBalpha degradation, nuclear factor-kappa B activation, and iNOS expression. Therefore, we conclude that the antioxidant activities of BCA and its inhibition of IkappaBalpha degradation and iNOS protein expression may have therapeutic potential, given that excessive free radicals and NO production have been associated with various inflammatory diseases.

New bis-catechols 5-lipoxygenase inhibitors.

Three polyhydroxy-2-phenylnaphthalenes (1-3) and the oxy analogue of tetrahydroxypavinan (4) were prepared and evaluated for their antioxidant properties (inhibition of diphenylpycrylhydrazyl radical (DPPH), reduction of iron (III) ion) and inhibition of 5-lipoxygenase (5-LO) activity. Their three-dimensional structures were established on the basis of spectroscopic data and semiempirical calculations. Compounds 1 and 2 were found as potent 5-LO inhibitors as nordihydroguaiaretic acid (NDGA), whereas 4 is 2.5 times less potent than NDGA. The reliability of the 3-D structures with the 5-LO inhibition properties is discussed. Their antioxidant properties show that tested compounds are expected to act as redox inhibitors.

Antioxidant activity of Nigella sativa essential oil.

The essential oil of black cumin seeds, Nigella sativa L., was tested for a possible antioxidant activity. A rapid evaluation for antioxidants, using two TLC screening methods, showed that thymoquinone and the components carvacrol, t-anethole and 4-terpineol demonstrated respectable radical scavenging property. These four constituents and the essential oil possessed variable antioxidant activity when tested in the diphenylpicrylhydracyl assay for non-specific hydrogen atom or electron donating activity. They were also effective.OH radical scavenging agents in the assay for non-enzymatic lipid peroxidation in liposomes and the deoxyribose degradation assay. GC-MS analysis of the essential oil obtained from six different samples of Nigella sativa seeds and from a commercial fixed oil showed that the qualitative composition of the volatile compounds was almost identical. Differences were mainly restricted to the quantitative composition.

Antioxidant activity of various fractions of non-tannin phenolics of canola hulls.

Cyclone canola hulls were extracted with 70% (v/v) acetone. The dried crude extract was dissolved in ethanol and fractionated on a Sephadex LH-20 column using 95% (v/v) ethanol as the mobile phase. Five major fractions were isolated according to the UV absorption. All fractions exhibited marked antioxidant activity in a beta-carotene-linoleate model system. Fractions I and II showed the best preventive effect against the bleaching of beta-carotene. The scavenging effect of fractions I, III, and V, at 1 mg, on alpha, alpha-diphenyl-beta-picrylhydrazyl (DPPH) radical was 67.4%, 80.7%, and 63.3%, respectively. Fractions II and IV showed weak DPPH scavenging effects. The reducing power of phenolics present in fractions IV and V was greater than that of fractions I-III, and the observed data correlated well (r(2) = 0.937; P = 0.007) with the total content of phenolics present in each fraction.

Antioxidant properties of N-(4-hydroxyphenyl)retinamide (fenretinide).

Fenretinide, N-(4-hydroxyphenyl)retinamide (4-HPR), is a cancer chemopreventive and antiproliferative agent whose mechanism of action is unknown. 4-HPR is a potent inducer of apoptosis in HL60 human leukemia cells which generates intracellular reactive oxygen species. The structural similarity of retinoic acid (RA), 4-HPR, and alpha-tocopherol (vitamin E) led us to investigate whether 4-HPR exhibits antioxidant activity. It was found that 4-HPR scavenged alpha,alpha-diphenyl-beta-picrylhydrazyl (DPPH) radicals in a 1:1 ratio in contrast to vitamin E, where a 1:2 ratio relative to DPPH radicals was observed. In addition, linoleic acid peroxidation initiated by hydroxyl radicals was decreased by 4-HPR to the same extent as by vitamin E. Furthermore, lipid peroxidation in rat liver microsomes was reduced by 4-HPR to a greater extent than by vitamin E. Based on these results, 4-HPR appears to be an effective antioxidant that may have clinical utility for diseases treated with vitamin E.

Dietary antioxidants fail in protection against oxidative genetic damage in in vitro evaluation.

Carcinogenesis is believed to be induced through the oxidative damage of DNA, and antioxidants are expected to suppress it. So, the polyphenolic antioxidants in daily foods were investigated to see whether they protect against genetic damage by active oxygen. In the evaluation, we used a bioassay and a chemical determination, a Salmonella mutagenicity test for mutation by a N-hydroxyl radical from one of the dietary carcinogens 3-amino-1-methyl-5H-pyrido[4,3-b]indole and the formation of 8-hydroxyl (8-OHdG) from 2'-deoxyguanosine (2'-dG) in a Fenton OH-radical generating system. Thirty-one antioxidants including flavonoids were compared in terms of radical-trapping activity with bacterial DNA and 2'-dG. Antioxidants inhibited the mutation but the IC50 values were in the mM order. Against 8-OHdG formation, only alpha-tocopherol had a suppressive effect with an IC50 of 1.5 microM. Thus, except alpha-tocopherol, the dietary antioxidants did not scavenge the biological radicals faster than bacterial DNA and intact 2'-dG, indicating that they failed to prevent oxidative gene damage and probably carcinogenesis.

Inhibitory effect of quercetin metabolites and their related derivatives on copper ion-induced lipid peroxidation in human low-density lipoprotein.

To determine the antioxidant activity of dietary quercetin (3,3',4', 5,7-pentahydroxyflavone) in the blood circulation, we measured the inhibitory effect of quercetin metabolites and their related derivatives on copper ion-induced lipid peroxidation of human low-density lipoprotein (LDL). Conjugated quercetin metabolites were prepared from the plasma of rat 1 h after oral administration of quercetin aglycone (40 micromol/rat). The rate of cholesteryl ester hydroperoxide (CE-OOH) accumulation and the rate of alpha-tocopherol consumption in mixtures of LDL solution (0.4 mg/ml) with equal volumes of this preparation were slower than the rates in mixtures of LDL with preparations from control rats. The concentrations of CE-OOH after 2 h oxidation in the mixtures of LDL with preparations of conjugated quercetin metabolites were significantly lower than those in the control preparation. It is therefore confirmed that conjugated quercetin metabolites have an inhibitory effect on copper ion-induced lipid peroxidation in human LDL. Quercetin 7-O-beta-glucopyranoside (Q7G) and rhamnetin (3,3',4', 5-tetrahydroxy-7-methoxyflavone) exerted strong inhibition and their effect continued even after complete consumption, similarly to quercetin aglycone. The effect of quercetin 3-O-beta-glucopyranoside (Q3G) did not continue after its complete consumption, indicating that the antioxidant mechanism of quercetin conjugates lacking a free hydroxyl group at the 3-position is different from that of the other quercetin conjugates. The result that 4'-O-beta-glucopyranoside (Q4'G) and isorhamnetin (3,4',5, 7-tetrahydroxy-3'-methoxyflavone) showed little inhibition implies that introduction of a conjugate group to the position of the dihydroxyl group in the B ring markedly decreases the inhibitory effect. The results of azo radical-induced lipid peroxidation of LDL and the measurement of free radical scavenging capacity using stable free radical, 1,1,-diphenyl-2-picrylhydrazyl, demonstrated that the o-dihydroxyl structure in the B ring is required to exert maximum free radical scavenging activity. It is therefore likely that conjugation occurs at least partly in positions other than the B ring during the process of metabolic conversion so that the inhibitory effect of dietary quercetin is retained in blood plasma after absorption.

Oxidative modification of apolipoprotein E in human very-low-density lipoprotein and its inhibition by glycosaminoglycans.

The mechanism of metal ion-catalyzed oxidative modification of apolipoprotein E (apoE) in human very-low-density lipoprotein (VLDL) and its inhibition by glycosaminoglycan (GAG) was investigated in vitro. The VLDL oxidation catalyzed by Cu2+ led to the lipid peroxidation, the formation of aggregates, and covalent modification of apoE. The modified apoE lost heparin-binding activity. These results suggest that the lipid peroxidation of VLDL and modification of apoE cause impairment of lipid uptake by cells and deposit the oxidized lipids in the tissues. The lipid peroxidation and oxidative modification of apoE in VLDL mediated by Cu2+ and an aqueous radical generator were suppressed by GAG, heparan sulfate, heparin, and chondroitin sulfate A, even though GAGs demonstrated no ability to scavenge alpha,alpha-diphenyl-beta-picrylhydrazyl radical. There were no relationships between inhibitory activity of GAGs in the VLDL oxidation and their number of sulfate groups which possess chelating activity of metal ion. Therefore, it can be considered that the inhibition of VLDL oxidation by GAGs is possibly due to the interaction between GAG and VLDL which bring about the steric hindrance, interference with the reaction between VLDL particle and the reactive oxygen species. These studies suggest that GAGs preserve the biological functions of apoE from oxidative stress.

Oxidation of the flavonol fisetin by polyphenol oxidase.

The present study demonstrates the antiradical efficiency of fisetin, a flavonol widely distributed in fruits and vegetables, by its ability to react with two different free radicals, ABTS; and DPPH;. The polyphenolic nature of fisetin led us to consider whether it might be oxidised by polyphenol oxidase (PPO), and the results reported show that it can be oxidised by PPO extracted and partially purified from broad bean seeds. The reaction was followed by recording spectral changes with time, with maximal spectral changes being observed at 282 nm (increase in absorbance) and at 362 nm (decrease). The presence of two isosbectic points (at 265 and 304 nm) suggested that only one absorbent product was formed. These spectral changes were not observed in the absence of PPO. The oxidation rate varied with the pH, reaching its highest value at pH 5.5. The fisetin oxidation rate increased in the presence of sodium dodecyl sulfate, an activator of polyphenol oxidase. Maximal activity was obtained at 0.87 mM sodium dodecyl sulfate. The following kinetic parameters were determined: Vmax=49 microM/min, Km=0.6 mM, Vmax/Km=8.2x10-2 min-1. Flavonol oxidation was inhibited by selective PPO inhibitors such as cinnamic acid (a classical competitive inhibitor, Ki=1.4 mM) and 4-hexylresorcinol, which behaved as a slow-binding inhibitor. The results reported show that fisetin oxidation was strictly dependent on the presence of polyphenol oxidase.

Structural aspects of the inhibitory effect of glabridin on LDL oxidation.

The inhibitory effects of glabridin, an isoflavan isolated from licorice (Glycyrrhiza glabra) root, and its derivatives on the oxidation of LDL induced by copper ions or mediated by macrophages were studied, in order to evaluate the contribution of the different parts of the isoflavan molecule to its antioxidant activity. The peak potential (E1/2) of the isoflavan derivatives, their radical scavenging capacity toward 1,1-diphenyl-2-picryl-hydrazyl (DPPH) radical and their ability to chelate heavy metals were also analyzed and compared to their inhibitory activity on LDL oxidation. In copper ion-induced LDL oxidation, glabridin (1), 4'-O-methylglabridin (2), hispaglabridin A (3), and hispaglabridin B (4), which have two hydroxyl groups at positions 2' and 4' or one hydroxyl at position 2' on ring B, successfully inhibited the formation of conjugated dienes, thiobarbituric acid reactive substances (TBARS) and lipid peroxides, and inhibited the electrophoretic mobility of LDL under oxidation. Compounds 1-3 exhibited similar activities, whereas compound 4 was less active. In macrophage-mediated LDL oxidation, the TBARS formation was also inhibited by these isoflavans (1-4) at a similar order of activity to that obtained in copper ion-induced LDL oxidation. On the other hand, 2'-O-methylglabridin (5), a synthesized compound, whose hydroxyl at 2'-position is protected and the hydroxyl at 4'-position is free, showed only minor inhibitory activity in both LDL oxidation systems. 2',4'-O-Dimethylglabridin (6), whose hydroxyls at 2'- and 4'-positions are both protected, was inactive. Resorcinol (7), which is identical to the phenolic B ring in glabridin, presented low activity in these oxidation systems. The isoflavene glabrene (8), which contains an additional double bond in the heterocyclic C ring, was the most active compound of the flavonoid derivatives tested in both oxidation systems. The peak potential of compounds 1-5 (300 microM), tested at pH 7.4, was similar (425-530 mV), and that for compound 6 and 8 was 1078 and 80 mV, respectively. Within 30 min of incubation, compounds 1, 2, 3, 4, 8 scavenged 31%, 16%, 74%, 51%, 86%, respectively, of DPPH radical, whereas compounds 5 and 6, which almost did not inhibit LDL oxidation, also failed to scavenge DPPH. None of the isoflavan derivatives nor the isoflavene compound were able to chelate iron, or copper ions. These results suggest that the antioxidant effect of glabridin on LDL oxidation appears to reside mainly in the 2' hydroxyl, and that the hydrophobic moiety of the isoflavan is essential to obtain this effect. It was also shown that the position of the hydroxyl group at B ring significantly affected the inhibitory efficiency of the isoflavan derivatives on LDL oxidation, but did not influence their ability to donate an electron to DPPH or their peak potential values.

Evaluation of the antioxidant potential of natural products.

Since reactive oxygen radicals play an important role in carcinogenesis and other human disease states, antioxidants present in consumable fruits, vegetables, and beverages have received considerable attention as cancer chemopreventive agents. Thus, in order to identify antioxidants in plant extracts, test materials were assessed for potential to scavenge stable 1,2-diphenyl-2-picrylhydrazyl (DPPH) free radicals, reduce TPA-induced free radical formation in cultured HL-60 human leukemia cells, and inhibit responses observed with a xanthine/xanthine oxidase assay system. Approximately 700 plant extracts were evaluated, and 28 were found to be active in the DPPH free radical scavenging assay. Based on secondary analyses performed to assess inhibition of 7,12-dimethylbenz(a)anthracene-induced preneoplastic lesion formation with a mouse mammary organ culture model, Chorizanthe diffusa Benth. (Polygonaceae), Mezoneuron cucullatum Roxb. (Leguminosae), Cerbera manghas L. (Apocynaceae) and Daphniphyllum calycinum Benth. (Daphniphyllaceae) were selected and subjected to bioassay-guided fractionation. 5,7,3',5'-Tetrahydroxy-8,4'-dimethoxyflavonol, 5,8,4'-trihydroxy-7,3'-dimethoxyflavonol, 5,3',4'-trihydroxy-7-methoxyflavonol, and 6,3',4'-trihydroxy-7-methoxyflavonol were identified as active principles from C. diffusa. Piceatannol, trans-resveratrol, apigenin and scirpusin A were found as the active principles of M. cucullatum, olivil, (-)-carinol, and (+)-cycloolivil were active principles from C. manghas, and 5,6,7,4'-tetrahydroxyflavone 3-O-rutinoside and kaempferol 3-O-neohesperidoside were active principles from D. calycinum. Of these substances, the hydroxystilbenes piceatannol and transresveratrol have thus far been shown to inhibit carcinogen-induced preneoplastic lesion formation in the mouse mammary gland organ culture model.

Beta-blockers inhibit the modification of low-density lipoproteins by sodium hypochlorite in vitro.

The effect of beta-blockers (alprenolol, oxprenolol, atenolol, acebutolol) and the non-steroidal anti-inflammatory drug, diclofenac, on modification of low-density lipoproteins (LDL) by sodium hypochlorite (NaOCl) was investigated in vitro. Beta-blockers and diclofenac inhibit the formation of thiobarbituric acid reactive substances in LDL modified by NaOCl. Beta-blockers, but not diclofenac, inhibit the hypochlorite-induced aggregation of LDL which was determined by photon correlation spectroscopy. The intracellular accumulation of cholesterol esters in J774 macrophages is inhibited by addition of beta-blockers, but not diclofenac, to LDL prior to the addition of NaOCl. The modification inhibiting effect of beta-blockers is inversely correlated to the binding capabilities of these substances to LDL which were assessed by laser electrophoresis. Inhibition of LDL modification in vivo by beta-blockers may reduce the risk of atherosclerosis and, therefore, compensate for the cholesterol-raising effect of these drugs in human plasma.