Antimicrobial Activity of 23 Endemic Plants in Madagascar

Purpose: To screen the crude methanol extracts obtained from 23 endemic plants in Madagascar for antimicrobial activity. Methods: In order to assess the antimicrobial properties of the extracts; their minimum inhibitory concentrations (MICs) were obtained using the broth microdilution method. The six test pathogenic species used were Bacillus subtilis; Staphylococcus aureus; Escherichia coli; Salmonella typhi; Pseudomonas aeruginosa and Candida albicans. Bioautography agar overlay test and phytochemical screening were also performed on the most active extracts. Results: From the 23 plants tested; 16 of which are used in traditional medicine; Poivrea phaneropetala Combretaceae); Koehneria madagascariensis (Lythraceae) and Rhopalopilia perrieri (Opiliaceae) exhibited the broad spectrum of activity; being active against all the test organisms; while Monoporus clusiifolius (Myrsinaceae) showed the strongest antifungal activity against Candida albicans with a minimal inhibitory concentration of 0.250 mg/ml. Bioautography and phytochemical analysis of the five active extracts against bacterial strains and of one active extract against C. albicans indicate that the active compounds responsible for antimicrobial activity may be mainly flavonoids and/or terpenes. Conclusion: These preliminary results are the first antimicrobial studies on these of some of them in traditional medicine

Comparative effects of antifungal agents on zidovudine glucuronidation by human liver microsomes.

Zidovudine (ZDV) is extensively metabolised by the liver to an inactive glucuronide (GZDV). Since ZDV is often administered with antimycotic drugs, we studied the effect of six systemic antifungal agents on the in vitro glucuronidation of ZDV by human liver microsomes. 5-fluorocytosine and itraconazole had no inhibitory effect whereas amphotericine B, ketoconazole, miconazole and fluconazole inhibited in vitro GZDV formation (Ki values were 0.13, 0.08, 0.18 and 1.4 mM respectively).

Effect of anticancer drugs on the glucuronidation of 3'-azido-3'-deoxythymidine in human liver microsomes.

Because zidovudine (3'-azido-3'-deoxythymidine or AZT) is frequently used in combination with other drugs for the treatment of acquired immune deficiency syndrome (AIDS) or AIDS-related complex diseases, drug interaction studies are required to improve the efficiency or decrease the toxicity of this antiviral drug. Although AZT is extensively metabolized as 5'-O-glucuronide (GAZT), we have recently demonstrated that many drugs that are or are not glucuronidated could be involved in relevant interactions. In this article, we screened the effect of 16 anticancer drugs on the glucuronidation of AZT by human liver microsome. Our results demonstrate that six anticancer drugs inhibit the in vitro formation of GAZT. Cyclophosphamide, ifosfamide, methotrexate, and etoposide are competitive inhibitors, whereas navelbine and vinblastine are noncompetitive inhibitors of AZT glucuronidation. Their estimated apparent Ki values ranged from 0.3 mM for navelbine to 9.8 mM for methotrexate. For compounds that competitively inhibit the in vitro formation of GAZT, theoretical percentages of inhibition obtainable in vivo at clinically relevant plasma concentrations of the coadministered drugs were determined. By considering these parameters, the rank order of these drugs with respect to their potential inhibition is cyclophosphamide >> ifosfamide > methotrexate = etoposide. Because the peak physiological concentrations (usual expected plasma levels) of ifosfamide, methotrexate, and etoposide are considerably less than their Ki values, only cyclophosphamide should inhibit the in vivo hepatic glucuronidation of AZT. Complementary clinical pharmacokinetic studies should be useful to confirm these findings.

[In vitro metabolism of zidovudine in man]. / Métabolisme in vitro de la zidovudine (AZT) chez l'homme.

The metabolism of zidovudine (AZT) and its modulating factors have been studied in human liver microsomes. In a first step, we demonstrated the involvement of UDP-glucoronosyltransferase (UDPGT) 2 form in AZT glucuronidation. In a second step, in order to predict drug interactions, we screened the effect of 52 drugs, representative of 17 different therapeutic classes, on AZT glucuronidation. We demonstrated that about twenty molecules glucuronidated or not are able to inhibit AZT glucuronidation. Finally, the NADPH-dependent reductive metabolism of AZT which produced a toxic metabolite, 3'-amino-3'-deoxythymidine (AMT) has been studied. Our studies demonstrated that AMT was formed only under anaerobic conditions and that its formation is catalysed by the NADPH-cytochrome P450 reductase.

UDP-glucuronosyltransferase activity toward digitoxigenin monodigitoxoside in human liver microsomes.

The properties of UDP-glucuronosyltransferase (UDPGT) toward digitoxigenin-monodigitoxoside (DT1) have been studied in human liver microsomes. The enzyme activity determined in nonactivated microsomes was very low (20 pmol/min/mg protein) compared with previously published values in rats (104 pmol/min/mg protein) or mice (379 pmol/min/mg protein) DT1-UDPGT activity was increased (180 to 220% of control activity) by Lubrol PX, Triton X-100, or (3-[3-cholamidopropyl]dimethylammonio)-1-propane sulfonic acid. The rate of DT1 glucuronidation determined for 29 different human liver microsomes was variable (18 to 87 pmol/min/mg protein). The KM found was approximately 4.5 microM. DT1-UDPGT activity was tentatively correlated with other known UDPGT activities. No significant correlations were found between DT1 and p-nitrophenol or 4-hydroxybiphenyl UDPGT activities. On the other hand, a strong correlation (r = 0.64, p < 0.05) was observed between DT1-UDPGT activity and digoxigenin-monodigitoxoside (DG1, another cardiac glycoside) glucuronidation rate.

3'-azido-3'-deoxythymidine drug interactions. Screening for inhibitors in human liver microsomes.

Zidovudine is a widely used antiretroviral drug active against human immunodeficiency virus. The drug interactions of this compound, which are primarily eliminated as a glucuronide, have not yet been extensively studied. Because zidovudine is frequently combined with other drugs, complete knowledge of interactions is essential to optimize AIDS therapy. We therefore screened the effect of 55 molecules, representative of 20 different therapeutic classes, on 3'-azido-3'-deoxythymidine (AZT) glucuronidation by human liver microsomes. We demonstrate that many drugs caused more than 15% inhibition of AZT glucuronidation in vitro, whereas major antibiotics (ceftazidine, isoniazid, aminoglycosides, macrolides, and sulfamides), antivirals (2',3'-dideoxycytidine, 2',3'-dideoxyinosine, and acyclovir), flucytosine, metronidazole, acetaminophen, and ranitidine had no effect. For compounds that appeared to inhibit AZT glucuronidation, extrapolation to the clinical situation must take into account both the in vitro apparent Ki values and the usual expected plasma level for the coadministered drug. By considering these parameters, this work indicates that clinically relevant inhibition of AZT glucuronidation may be observed with the following drugs: cefoperazone, penicillin G, amoxicilin, piperacillin, chloramphenicol, vancomycin, miconazole, rifampicin, phenobarbital, carbamazepine, phenytoin, valproic acid, quinidine, phenylbutazone, ketoprofen, probenecid, and propofol. Complementary clinical and pharmacokinetic studies should be performed to validate these assumptions.

In vitro glucuronidation of 3'-azido-3'-deoxythymidine by human liver. Role of UDP-glucuronosyltransferase 2 form.

The glucuronidation of 3'-azido-3'-deoxythymidine (AZT) by human liver microsomes and human hepatocytes in culture has been studied in vitro to determine the UDP-glucuronosyltransferase (UDPGT) form conceivably involved in the AZT biotransformation process. The glucuronide of AZT was preliminarily identified through hydrolysis by beta-D-glucuronidase. Brij 58 was shown to be the best activator of AZT glucuronidation by human liver microsomes, as it increased the rate of glucuronide formation 3-fold. The UDPGT activities toward AZT measured in 29 different microsomal fractions was slightly variable among samples (79 to 268 nmol/hr/mg protein). The apparent KM value for AZT glucuronidation was about 5 mM. We sought to determine if various known UDPGT activities (i.e. p-nitrophenol UDPGT, 4-hydroxybiphenyl UDPGT, and DT1-UDPGT) in 18 microsomal samples were correlated with AZT-UDPGT activity. Experiments revealed that only 4-hydroxybiphenyl UDPGT activity was strongly correlated (r = 0.815, p less than 0.001) with AZT-UDPGT activity, whereas no correlation was found for the other UDPGT activities. To determine the isozyme conceivably involved in AZT glucuronidation, we studied the effect of various compounds on AZT glucuronidation. AZT glucuronidation was inhibited by numerous substrates of the UDPGT2, form: morphine (Ki = 1.8 mM), 4-hydroxybiphenyl (Ki = 0.92 mM), and ketoprofen (Ki = 0.75 mM), but also oxazepam, codeine, and chloramphenicol. p-Nitrophenol appeared to be an inhibitor, whereas acetaminophen had no effect. Bilirubin, aspirin, cimetidine, and acyclovir did not inhibit AZT glucuronidation. Since all the inhibitors tested except p-nitrophenol are known to be glucuronidated by the UDPGT2 form, our results strongly suggest the involvement of this isozyme in AZT glucuronidation.(ABSTRACT TRUNCATED AT 250 WORDS)

Use of a human liver microsome bank in drug glucuronidation studies.

A bank of readily available, well characterized human hepatic microsomal fractions (29 samples from different livers) has been established. The enzymatic characteristics of each microsome sample were determined using three specific UDP-glucuronosyltransferase (UDPGT) substrates (p-nitrophenol for UDPGT(1), 4-hydroxybiphenyl for UDPGT(2)A and monodigitoxoside digitoxigenin (DT(1)) for UDPGT(7)). After characterization, the bank was used to study the phase II biotransformation processes of two drugs: an antiviral, zidovudine (AZT), and an intermediate metabolite of digoxin, monodigitoxoside digoxigenin (DG(1)). Results showed a wide interindividual variability for the glucuronidation rate of 4-hydroxybiphenyl, DT(1), AZT and DG(1). Surprisingly, this variability was lower for p-nitrophenol. No significant correlation was found between the various activities except between DG(1) and DT(1) UDPGT activities and 4-hydroxybiphenyl and AZT UDPGT activities. For AZT, further experiments showed that glucuronidation of this antiviral compound was inhibited by 4-hydroxybiphenyl and chloramphenicol but not by morphine, p-nitrophenol, acetaminophen, or bilirubin. Finally, the results strongly suggest the respective involvement of UDPGT(2)A and UDPGT(7) in the glucuronidation of AZT and DG(1).