Anti-Helicobacter pylori flavonoids from licorice extract

Licorice is the most used crude drug in Kampo medicines (traditional Chinese medicines modified in Japan). The extract of the medicinal plant is also used as the basis of anti-ulcer medicines for treatment of peptic ulcer. Among the chemical constituents of the plant, glabridin and glabrene (components of Glycyrrhiza glabra), licochalcone A (G. inflata), licoricidin and licoisoflavone B (G. uralensis) exhibited inhibitory activity against the growth of Helicobacter pylori in vitro. These flavonoids also showed anti-H. pylori activity against a clarithromycin (CLAR) and amoxicillin (AMOX)-resistant strain. We also investigated the methanol extract of G. uralensis. From the extract, three new isoflavonoids (3-arylcoumarin, pterocarpan, and isoflavan) with a pyran ring, gancaonols A[bond]C, were isolated together with 15 known flavonoids. Among these compounds, vestitol, licoricone, 1-methoxyphaseollidin and gancaonol C exhibited anti-H. pylori activity against the CLAR and AMOX-resistant strain as well as four CLAR (AMOX)-sensitive strains. Glycyrin, formononetin, isolicoflavonol, glyasperin D, 6,8-diprenylorobol, gancaonin I, dihydrolicoisoflavone A, and gancaonol B possessed weaker anti-H. pylori activity. These compounds may be useful chemopreventive agents for peptic ulcer or gastric cancer in H. pylori-infected individuals.     

Anti-Helicobacter pylori activities of six Iranian plants

BACKGROUND: Helicobacter pylori is the major worldwide cause of bacterial gastrointestinal infections in adults and children. Antibiotic therapy and a combination of two or three drugs have been widely used to eradicate these infections. However, development of drug resistance in bacteria calls for new sources of drugs, and plants seem to be a logical source of new antibacterial compounds. METHODS: The anti-H. pylori activities of six native Iranian plants (Glycyrrhiza aspera, Juglans regia, Ligustrum vulgare, Thymus kotschyanus, Trachyspermum copticum and Xanthium brasilicum) and seven antibiotics were determined against 70 clinical isolates from children using the disk susceptibility assay. Minimum inhibitory concentrations were also measured for the biologically active extracts. One extract with the best anti-H. pylori activity was fractionated by silica gel and thin layer chromatography and the active compounds were identified by hydrogen nuclear magnetic resonance ((1)HNMR) spectroscopy. RESULTS: All plant extracts showed anti-H. pylori activity by the disk sensitivity method, but the most active extracts were those from X. brasilicum and T. copticum. In fact, the anti-H. pylori activities of the two extracts were superior to the disk antibiotic susceptibility profile. Minimum inhibitory concentrations were within the range of 31.25-250 micro g/ml. Fractionation and chemical identification of the extract from X. brasilicum showed the presence of two substances, a flavonoid and a xanthanolide. CONCLUSIONS: Due to the rise in antibiotic resistance, new sources of anti-H. pylori drugs are needed. The use of medicinal plants and/or their chemical components may have potential benefit in eradicating such problems.     

Anti-Helicobacter pylori activity of Plumbago zeylanica L

It has been shown that the presence of infection by Helicobacter pylori is strongly associated with gastric cancer and peptic ulceration. In western medicine, a 3-fold therapeutic regimen, emphasizing the use of antibiotics, is typically used to suppress H. pylori activity. However, antibiotic drug resistance frequently develops as a consequence of such treatment. In our previous study, 50 Taiwanese folk medicinal plants were screened for their anti-H. pylori activities. The results revealed that Plumbago zeylanica L. had the highest inhibitory effects against H. pylori. In this study, therefore, we have focused on establishing the anti-H. pylori activities of P. zeylanica L. Water and the organic solvents ethanol, ethyl acetate and acetone were used for P. zeylanica L. extraction, obtaining yields of 1.66-6.84% (w/w). Excluding the water extract, higher anti-H. pylori activity was demonstrated for all the extracts, both using the agar diffusion and dilution methods. The ethyl acetate extract exhibited the lowest minimum inhibitory concentrations against five H. pylori strains, of which ranged from 0.32 to 1.28 mg ml-1, followed, in ascending order, by the acetone, ethanol and water analogs. Bactericidal activity was determined for P. zeylanica L. extracts, with the lowest minimum bactericidal concentrations (5.12-20.48 mg ml-1) demonstrated for the ethyl acetate, followed, in ascending order, by the acetone and ethanol analogs. Bactericidal activity appeared to be in a dose-dependent manner. Through a broad pH range (2-7), bactericidal activity was not affected when extract concentrations were greater than or equal to the minimum bactericidal concentration. High stability was demonstrated for the ethyl acetate P. zeylanica L. extract within pH range of 1-7, exhibiting all pH treatments bactericidal activity.     

Comparing binding of histamine and H2-antagonist with their effects on gastric acid secretion

A microsomal fraction from isolated frog gastric mucosa was used to study the binding of labeled histamine, labeled metiamide (a histamine H2-antagonist), and competition between labeled histamine and unlabeled metiamide. The separation of free from bound ligand was done by gel chromatography. The acid secretion was studied in frog gastric mucosa in vitro by a pH-stat method. The binding data could be interpreted in terms of two independent binding sites for both histamine and metiamide. However, the competition between histamine and metiamide does not support the independence of the sites. Moreover, the dissociation kinetics of labeled metiamide in the presence of unlabeled metiamide is non-monotone and, thus, indicates cooperativity. In the physiological studies, the dependence of the rate of acid secretion on histamine stimulation occurs within very narrow limits, which is the result of characteristics other than related to binding. However, the total amount of acid secreted caused by a pulse of histamine does indicate two sites, of which the high-affinity site is the more effective. Metiamide inhibition of acid secretion can be interpreted as an interaction between high-affinity sites of histamine and metiamide. Overall, studies involving physiological effects provide less precise data than the direct binding studies.     

In vitro histamine H2-antagonist activity of the novel compound HUK 978

Histamine stimulated adenylate cyclase from guinea-pig fundic mucosa and 3H-tiotidine binding in guinea-pig cerebral cortex were used to assess the in-vitro histamine H2-activity of the novel H2-antagonist HUK 978. The results showed that HUK 978 was a more potent H2-antagonist than either cimetidine or ranitidine. HUK 978 was also shown to be devoid of activity at the histamine H1-receptor, the muscarinic receptor and the alpha and beta-adrenergic receptors.     

Anti-Helicobacter pylori effects of IgY from egg york of immunized hens

Effects of egg york containing IgY specific for Helicobacter pylori on the bacterial growth and intragastric infection were investigated in comparison with a proton-pump inhibitor pantoprazole. For in vitro anti-bacterial activity test, H. pylori (1×10(8) CFU/mL) was incubated with a serially diluted IgY for 3 days. As a result, IgY fully inhibited the bacterial growth at 16 mg/mL, which was determined to a minimal inhibitory concentration. In vivo elimination study, male C57BL/6 mice were infected with the bacteria by intragastric inoculation (1×10(8) CFU/mouse) 3 times at 2-day intervals, and 2 weeks later, orally treated twice a day with 50, 100, 200 or 500 mg/kg IgY for 18 days. After the final administration, biopsy sample of the gastric mucosa was assayed for the bacterial identification via urease, oxidase, catalase, nitrate reduction and H(2)S tests in addition to microscopic examination for mucosal inflammation. In CLO kit test, 75, 50, 12.5 and 12.5% of the animals revealed positive reaction following treatment with 50, 100, 200 and 500 mg/kg IgY, respectively, resulting in a superior efficacy at 200 mg/kg than 30 mg/kg pantoprazole that displayed 75% elimination. The CLO test results were confirmed by bacterial identification. Microscopic examination revealed that H. pylori infection caused severe gastric mucosal inflammation, which were not observed in the CLO-negative mice following treatment with IgY or pantoprazole. Taken together, IgY inhibited the growth of H. pylori, and improved gastritis and villi injuries by eliminating the bacteria from the stomach. The results indicate that IgY could be a good candidate overcoming tolerance of antibiotics for the treatment of H. pylori-mediated gastric ulcers.     

Anti-Helicobacter pylori activity of Lactobacillus delbrueckii subsp. bulgaricus strains: preliminary report

Aims:  To evaluate the activities of six Lactobacillus delbrueckii subsp. bulgaricus (LB) strains against 30 Helicobacter pylori strains by agar-well diffusion method.
Methods and Results:  LB cultures [4 × 10⁸–4 × 10⁹ CFU ml⁻¹) either were prepared in milk at their native pH, 3·8–5·0, or were adjusted to pH 6·4–7·7. At low and neutralized pH, LB strains inhibited the growth by 40–86·7% and 16·7–66·7% of H. pylori strains, respectively. LB activity was strain-dependent. At low and neutralized pH, one and five H. pylori strains, respectively, were not inhibited by any LB strain. LB2 and LB3, taken together, were active against most metronidazole and clarithromycin resistant strains.
Conclusions:  All LB strains inhibited a number of H. pylori strains, including also antibiotic resistant strains. LB activity was strain-dependent and better at low pH. At low pH values, the most active LB strains were LB1, LB2 and LB3, inhibiting 86·7% of H. pylori strains, while at neutralized pH values, the most active LB strains were LB2 and LB3, inhibiting 53·3 and 66·7% of H. pylori strains, respectively.
Significance and Impact of the Study:  LB could be utilized in the treatment or prophylaxis of H. pylori infection and warrants clinical investigations.     

Anti-Helicobacter pylori activity of the methanolic extract of Geum iranicum and its main compounds

Geum iranicum Khatamsaz, belonging to the Rosaceae family, is an endemic plant of Iran. The methanol extract of the roots of this plant showed significant activity against one of the clinical isolates of Helicobacter pylori which was resistant to metronidazole. The aim of this study was the isolation and evaluation of the major compounds of G. iranicum effective against H. pylori. The compounds were isolated using various chromatographic methods and identified by spectroscopic data (1H and 13C NMR, HMQC, HMBC, EI-MS). An antimicrobial susceptibility test was performed employing the disk diffusion method against clinical isolates of H. pylori and a micro dilution method against several Gram-positive and Gram-negative bacteria; additionally the inhibition zone diameters (IZD) and minimum inhibitory concentrations (MIC) values were recorded. Nine compounds were isolated: two triterpenoids, uvaol and niga-ichigoside F1, three sterols, beta-sitosterol, beta-sitosteryl acetate, and beta-sitosteryl linoleate, one phenyl propanoid, eugenol, one phenolic glycoside, gein, one flavanol, (+)-catechin, and sucrose. The aqueous fraction, obtained by partitioning the MeOH extract with water and chloroform, was the most effective fraction of the extract against all clinical isolates of H. pylori. Further investigation of the isolated compounds showed that eugenol was effective against H. pylori but gein, diglycosidic eugenol, did not exhibit any activity against H. pylori. The subfraction D4 was the effective fraction which contained tannins. It appeared that tannins were probably the active compounds responsible for the anti-H. pylori activity of G. iranicum. The aqueous fraction showed a moderate inhibitory activity against both Gram-positive and Gram-negative bacteria. The MIC values indicated that Gram-positive bacteria including Staphylococcus aureus, Staphylococcus epidermidis, and Bacillus subtilis are more susceptible than Gram-neagative bacteria including Escherichia coli and Pseudomonas aeruginosa.     

Carnitine and acetyl-carnitine,natural substances endowed with interesting pharmacological properties.

The inotropic and anti-fatigue effects of d, l-carnitine, l-carnitine and d, l-acetyl-carnitine in laboratory animals were studied. In the isolated rabbit heart these substances at the concentration level of 10^?5 g/l increase significantly the contractile force depressed by anoxia, without changes in ECG, heart rate and coronary flow. The following rank order of activity was observed: d, l-acetyl-carnitine > l-carnitine > d, l-carnitine. Sodium acetate + d, l-carnitine mixture exerts the same inotropic effect as d, l-carnitine alone, showing the important role of the acetyl-bond. In rats submitted to a fatigue test (rotarod) only d, l-acetyl-carnitine induces a significant anti-fatigue effect. Since endogenous carnitine depletion is linked with many pathological states, particularly of the heart, exogenous carnitine and especially its acetyl-derivative might represent an important drug treatment with high tolerance advantages.     

Synthesis,Docking Studies and Evaluation of Chalcones as Anti-Helicobacter pylori and antitumoral Agents

Helicobacter pylori colonizes the gastric epithelium of 50 % of world population and it is the main etiological agent of human chronic gastritis, peptic ulcer, and gastric cancer. In this study, we synthesized and characterized a series of 14 chalcones and evaluated their anti-H. pylori, NO inhibition (in vitro and in silico), and AGS cells cytotoxic effects. Compounds 3b and 3h showed MIC of 8 μg/mL. We observed structure-activity relationships, mainly related to the influence of methoxy substituent at C-2 ( 3b ) and the nitro group at C-4 ( 3h ) in chalcone scaffold. The fourteen chalcones inhibited the NO production in LPS-stimulated macrophages and showed potential for interaction on the active site of the iNOS enzyme. Finally, 3b and 3h showed the highest selectivity to the AGS cell lines. Thus, ours results suggest 3b and 3h as potential candidates for design of new and effective agents against H. pylori and related diseases.     

One drug for two targets: Biological evaluation of antiretroviral agents endowed with antiproliferative activity

AIDS-related cancer diseases are malignancies with low incidence on healthy people that affect mostly subjects already immunocompromised. The connection between HIV/AIDS and these cancers has not been established yet, but a weakened immune system is certainly the main cause. We envisaged the possibility to screen a small library of compounds synthesized in our laboratory against opportunistic tumors mainly due to HIV infection like Burkitt’s Lymphoma. From cellular assays and gene expression analysis we identified two promising compounds. These derivatives have the dual action required inhibiting HIV replication in human TZM-bl cells infected with HIV-1 NL4.3 and showing cytotoxic activity on human colon HT-29 and breast adenocarcinoma MCF-7 cells. In addition, preclinical in vitro adsorption, distribution, metabolism, and excretion studies highlighted a satisfactory pharmacokinetic profile.     

Histamine paw edema of mice was increased and became H2-antagonist sensitive by co-injection of nitric oxide forming agents,but serotonin paw edema was decreased

Nitric oxide (NO) suprisingly caused the opposite effect on histamine and serotonin edema. The local injection of acidified nitrite (0.3–30 μg /paw which correspond to 10 μg−1mg/kg) increased histamine edema of mice up to 45±4% and suppressed serotonin edema to 90±3%. Other NO-generators (nitroprusside sodium and hydroxylamine) showed similar effects. These results were in accordance with our previous data on endogenous NO. Methylene blue (MB, 30ng/paw which corresponds to 1 μg/kg) suppressed histamine edema (62±3%) and increased serotonin edema (43±3%) in normal mice, being reversed by acidified nitrite. This suggests the involvement o of guanosine 3′, 5′ -cyclic monophosphate (cGMP) formation for the action of NO. Histamine edema became sensitive to H₂-antagonist, cimetidine, by co-injection of 30 μg/paw (which corresponds to 1mg/kg) acidified nitrite (ED₅₀=30 μg/kg versus ⪢ 1mg/kg). NO seemed to modify the histamine receptor(s) or tautomeric form of histamine. NO, O₂⁻ and other oxyradicals might finely control the vascular permeability together with inflammatory mediators.     

Plasminogen endowed with magnetic property

Plasminogen was modified with an activated magnetic modifier, which consists of magnetite (Fe₃O₄) and a polyethylene glycol derivative. The magnetic plasminogen was activated with urokinase and became a magnetic derivative of plasmin. The magnetic plasmin exhibited specific activity which is 32% of the native plasmin; 7.0 versus 22 casein units mg⁻¹ protein. It was readily attracted by a magnet (250 Oe). It is particularly worth mentioning that the magnetic plasmin was more resistant to plasmin inhibitor(s) in plasma than plasmin. By applying magnetic force, the magnetic plasmin was attracted at the site of fibrin clot and local fibrinolysis was achieved in saline-circulating system.     

Stereoselective disposition of flurbiprofen from a mutual prodrug with a histamine H2-antagonist to reduce gastrointestinal lesions in the rat

The in vitro and in vivo stereoselective hydrolysis characteristics of the mutual prodrug FP-PPA, which is a conjugate of flurbiprofen (FP) with the histamine H₂-antagonist PPA, to reduce gastrointestinal lesions induced by FP were investigated and compared with those of FP methyl ester (rac-FP-Me) and FP ethyleneglycol ester (rac-FP-EG). The rac-FP derivatives were hydrolyzed preferentially to the (+)-S-isomer in plasma and to the (−)-R-isomer in liver and small intestinal mucosa. Interestingly, in the gastric mucosa, the stereoselectivity of hydrolysis of (−)-R-FP-PPA was opposite from that of rac-FP-Me and rac-FP-EG, which suggested that the stereoselective hydrolysis of FP-PPA was helpful in reducing gastric damage induced by (+)-S-FP. However, hydrolysis of all rac-FP derivatives was found to be catalyzed by carboxylesterases in the gastric mucosa. The stereoselective disposition of FP enantiomers early after intravenous administration of rac-FP-PPA could be explained by the stereoselective formation of (−)-R-FP from rac-FP-PPA in the liver. (−)-R-FP-PPA was completely hydrolyzed to form (−)-R-FP in vivo, while 78% of (+)-S-FP-PPA was hydrolyzed to (+)-S-FP, with a corresponding decrease in the area under the curve. Twenty-five percent of (+)-S-FP-PPA might be eliminated as the intact prodrug or its metabolites other than FP. The most important bioconversion of FP-PPA occurred in plasma, and additional hydrolysis of the R-enantiomer in liver resulted in the stereoselectivity observed following both i.v. and p.o. administration. © 1996 Wiley-Liss, Inc.     

5'-Modified G-quadruplex forming oligonucleotides endowed with anti-HIV activity: synthesis and biophysical properties

Oligodeoxyribonucleotides of sequence d(5'TGGGAG3') carrying bulky aromatic groups at the 5' end were found to exhibit potent anti-HIV activity [Hotoda, H., et al. (1998) J. Med. Chem. 41, 3655-3663 and references therein]. Structure-activity relationship investigations indicated that G-quadruplex formation, as well as the presence of large aromatic substituents at the 5'-end, were both essential for their antiviral activity. In this work, we synthesized some representative examples of the anti-HIV active Hotoda's 6-mers and analyzed the resulting G-quadruplexes by CD, DSC, and molecular modeling studies, in comparison with the unmodified oligonucleotide. In the case of the sequence carrying the 3,4-dibenzyloxybenzyl (DBB) group, identified as the best candidate for further drug optimization, we developed an alternative protocol to synthesize the 5'-DBB-thymidine phosphoramidite building block in higher yields. The thermodynamic and kinetic parameters for the association/dissociation processes of the 5'-conjugated quadruplexes, determined with respect to the unmodified one, were discussed in light of the molecular modeling studies. The aromatic groups at the 5' position of d(5'TGGGAG3') dramatically enhance both the equilibrium and the rate of formation of the quadruplex complexes. The overall stability of the investigated quadruplexes was found to correlate with the reported IC50 values, thus furnishing quantitative evidence for the hypothesis that the G-quadruplex structures are the ultimate active species, effectively responsible for the biological activity.     

Anti-secretory activity of the novel H2-antagonist, HUK 978, in rat, guinea-pig and dog

The anti-secretory activity of the competitive H2-antagonist HUK 978 was determined in rat, guinea-pig and dog. In all systems examined, HUK 978 was more potent than cimetidine and ranitidine both intravenously and orally. In addition, the compound at approximately equipotent doses as these established H2-antagonists exhibited a significantly longer inhibitory profile following oral and systemic administration. Data from these pharmacological studies and the in vitro investigations previously reported, suggest that HUK 978 is a highly specific H2-antagonist and inhibits acid secretion for longer periods than other competitive compounds.     

Cardiovascular hybrid drugs: new benzazepinone derivatives as bradycardic agents endowed with selective beta1-non-competitive antagonism

The synthesis and pharmacological profile of some hybrid compounds bearing both the benzazepinone moiety present in Zatebradine and typical beta-blocker aryloxypropanolamine groups are described. The new compounds proved to be endowed with negative chronotropic and inotropic activity and are weak vasorelaxant agents. The cardiodepressant action is probably due to selective beta(1)-noncompetitive reversible antagonism. Both enantiomers of the most active compound 5c were synthesized and they showed a different cardiovascular profile, that is (+)-(R)-enantiomer displays affinity for cardiac beta(1)-adrenoceptors, while (-)-(S)-enantiomer shows specificity for vessel smooth muscle.     

Studies on anti-Helicobacter pylori agents. Part 2: new cephem derivatives

The synthesis and optimization of the anti-Helicobacter pylori activity of a novel series of cephem derivatives are described. Introduction of thio-heterocyclic groups containing N- and S-atoms to the 3-position and phenyl or thienyl acetamido groups to the 7-position of the cephem nucleus dramatically improved the activity. From this series of derivatives, compound 13i was found to have extremely potent in vitro anti-H. pylori activity, superior therapeutic efficacy compared to AMPC and CAM, no cross-resistance between CAM or MNZ and low potential for causing diarrhea due to instability to beta-lactamase.