Disposition of sanguinarine in the rat.

Sanguinarine is an alkaloid with known antibiotic and anti-inflammatory activity and its pharmacokinetics have been studied in the rat after a single oral dose (10 mg kg(-1) body weight). Alkaloid determination in the plasma and liver was carried out by high-performance liquid chromatography-electrospray ionization mass spectrometry (HPLC/ESI-MS). The pharmacokinetic parameters (t(max), c(max), AUC(0-->t) and AUC(0-->infinity)) were determined for sanguinarine and dihydrosanguinarine, the major components detected in plasma. The first step in sanguinarine metabolism in the rat was the reduction of the iminium bond resulting in formation of the less toxic dihydrosanguinarine. Both compounds were completely eliminated from the plasma and liver after 24 h and not detected in urine. After a single oral dose of (3)H-sanguinarine, more than 42% of the ingested radioactivity was present in gastrointestinal tract. Benz[c]acridine, up to date the only sanguinarine metabolite referred to in the literature, was not detected in the plasma, liver or urine.

Antimicrobial activity of some medicinal barks used in Peruvian Amazon.

The aim of this study was to evaluate the antimicrobial activity of six barks traditionally used in Callería District (Ucayali Department, Peru) for treating conditions likely to be associated with microorganisms. Ethanol extracts of stem barks of Abuta grandifolia (Menispermaceae), Dipteryx micrantha (Leguminosae), Cordia alliodora (Boraginaceae), Naucleopsis glabra (Moraceae), Pterocarpus rohrii (Leguminosae), and root bark of Maytenus macrocarpa (Celastraceae) were tested against nine bacteria and one yeast using the broth microdilution method. All plants possessed significant antimicrobial effect, however, the extract of Naucleopsis glabra exhibited the strongest activity against Gram-positive bacteria (MICs ranging from 62.5 to 125 microg/ml), while the broadest spectrum of action was shown by the extract of Maytenus macrocarpa, which inhibited all the strains tested with MICs ranging from 125 to 250 microg/ml.

Degradation of 2,4,6-trinitrotoluene by selected helophytes.

Four emergent plants (helophytes, synonyms emersion macrophytes, marsh plants, etc.) Phragmites australis, Juncus glaucus, Carex gracillis and Typha latifolia were successfully used for degradation of TNT (2,4,6-trinitrotoluene) under in vitro conditions. The plants took up and transformed more than 90% of TNT from the medium within ten days of cultivation. The most efficient species was Ph. australis which took up 98% of TNT within ten days. The first stable degradation products 4-amino-2,6-dinitrotoluene (4-ADNT) and 2-amino-4,6-dinitrotoluene (2-ADNT) were identified and analysed during the cultivation period. [14C] TNT was used for the detection of TNT degradation products and their compartmentalization in plant tissues after two weeks of cultivation. Forty one percent of 14C was detected as insoluble or bound in cell structures: 34% in roots and 8% in the aerial parts. These results open the perspective of using the above-mentioned plants for the remediation of TNT contaminated waters.

Evaluation of 3-azidiamantane as photoaffinity probe of cytochrome P450.

3-azidiamantane (DIA-N2) has been shown to be a photolabile carbene-generating probe interacting specifically with cytochrome P450 (P450) active centre. To evaluate the modification of P450 by the probe, radiolabelled [9-3H]-3-azidiamantane was prepared by reductive dehalogenation of its precursor, 3-oxo-9-bromodiamantane ethylene ketal. The synthesis was optimized as the proper precursor and reaction conditions were concerned to produce 96% pure product (overall yield 59%). An incorporation efficacy of the probe photoactivated at 366 nm was examined with two different proteins, BSA and rat phenobarbital-inducible P450 2B1, both having hydrophobic binding sites. Under photolysis the photoaffinity probe generated short-lived (> 90%) intermediates binding immediately to the protein. The yield of photoactivated DIA-N2 incorporation was 12% and 11% for BSA and P450, respectively. The presence of reduced glutathione, a scavenger of reactive intermediates, did not affect the probe incorporation markedly. On the other hand, scavengers entering the P450 active centre, methanol and dithiothreitol, reduced the protein labelling by 36% and 42%, respectively. Similarly, at DIA-N2, aminopyrine (substrates), and metyrapone (inhibitor) 50 times molar excess over the probe, prevented its binding by about 40%. In addition, when photoaffinity labelling was carried out with microsomal preparation, the substrate with a high affinity for the P450 2B1, diamantane, (at 20 times molar excess to the probe) caused 47% inhibition of the P450 covalent labelling. These results, suggesting a high specificity of the probe binding, show that it can be applied as a photoaffinity probe for cytochrome P450 2B1 active centre studies.

Heterobifunctional photoaffinity probes for cytochrome P450 2B.

Three heterobifunctional photoaffinity probes, N-(p-azidobenzyl)-N-methyl-p-aminobenzylamine (I), N-(p-azidobenzyl)-N-methyl-p-aminophenethylamine (II), and N-(p-azidophenethyl)-N-methyl-p-aminophenethylamine (III), were synthesized and characterized. These probes, containing a photolabile azido-group and an amino-group on opposite sides of the molecule, were designed for photoaffinty labeling of the cytochrome P450 (CYP) 2B active site cavity differing in distance from the heme iron. Spectroscopic studies proved that probes I and II coordinated with the heme iron via their amino-group in the enzyme active center, whereas probe III did not. This result in conjunction with data from kinetic studies suggests probes I and II are appropriate for photoaffinity labeling of the CYP 2B active center. Thus, probe II was used to identify amino acid residues within a distance of the probe length (about 16.5 A) from the heme. Analysis of a Lys-C digest of the probe II-labeled CYP 2B4 revealed a single labeled hexapeptide corresponding to position 192-197 of the CYP 2B4 sequence. Using postsource decay/matrix-assisted laser desorption ionization-time of flight, Arg197 was identified as a probe II target. The location of the labeled site in three-dimensional structures of bacterial CYPs and in CYP 2B homology models is discussed.