In vitro metabolism of haloperidol and sila-haloperidol: new metabolic pathways resulting from carbon/silicon exchange.

The neurotoxic side effects observed for the neuroleptic agent haloperidol have been associated with its pyridinium metabolite. In a previous study, a silicon analog of haloperidol (sila-haloperidol) was synthesized, which contains a silicon atom instead of the carbon atom in the 4-position of the piperidine ring. In the present study, the phase I metabolism of sila-haloperidol and haloperidol was studied in rat and human liver microsomes. The phase II metabolism was studied in rat, dog, and human hepatocytes and also in liver microsomes supplemented with UDP-glucuronic acid (UDPGA). A major metabolite of haloperidol, the pyridinium metabolite, was not formed in the microsomal incubations with sila-haloperidol. For sila-haloperidol, three metabolites originating from opening of the piperidine ring were observed, a mechanism that has not been observed for haloperidol. One of the significant phase II metabolites of haloperidol was the glucuronide of the hydroxy group bound to the piperidine ring. For sila-haloperidol, the analogous metabolite was not observed in the hepatocytes or in the liver microsomal incubations containing UDPGA. If silanol (SiOH) groups are not glucuronidated, introducing silanol groups in drug molecules could provide an opportunity to enhance the hydrophilicity without allowing for direct phase II metabolism. To provide further support for the observed differences in metabolic pathways between haloperidol and sila-haloperidol, the metabolism of another pair of C/Si analogs was studied, namely, trifluperidol and sila-trifluperidol. These studies showed the same differences in metabolic pathways as between sila-haloperidol and haloperidol.

Genetic and serological typing of European infectious haematopoietic necrosis virus (IHNV) isolates.

Infectious haematopoietic necrosis virus (IHNV) causes the lethal disease infectious haematopoietic necrosis (IHN) in juvenile salmon and trout. The nucleocapsid (N) protein gene and partial glycoprotein (G) gene (nucleotides 457 to 1061) of the European isolates IT-217A, FR-32/87, DE-DF 13/98 11621, DE-DF 4/99-8/99, AU-9695338 and RU-FR1 were sequenced and compared with IHNV isolates from the North American genogroups U, M and L. In phylogenetic studies the N gene of the Italian, French, German and Austrian isolates clustered in the M genogroup, though in a different subgroup than the isolates from the USA. Analyses of the partial G gene of these European isolates clustered them in the M genogroup close to the root while the Russian isolate clustered in the U genogroup. The European isolates together with US-WRAC and US-Col-80 were also tested in an enzyme-linked immunosorbent assay (ELISA) using monoclonal antibodies (MAbs) against the N protein. MAbs 136-1 and 136-3 reacted equally at all concentrations with the isolates tested, indicating that these antibodies identify a common epitope. MAb 34D3 separated the M and L genogroup isolates from the U genogroup isolate. MAb 1DW14D divided the European isolates into 2 groups. MAb 1DW14D reacted more strongly with DE-DF 13/98 11621 and RU-FR1 than with IT-217A, FR-32/87, DE-DF 4/99-8/99 and AU-9695338. In the phylogenetic studies, the Italian, French, German and Austrian isolates clustered in the M genogroup, whereas in the serological studies using MAbs, the European M genogroup isolates could not be placed in the same specific group. These results indicate that genotypic and serotypic classification do not correlate.

Novel metabolites of amodiaquine formed by CYP1A1 and CYP1B1: structure elucidation using electrochemistry, mass spectrometry, and NMR.

An aldehyde metabolite of amodiaquine and desethylamodiaquine has been identified. The aldehyde was the major metabolite formed in incubations with two recombinantly expressed human cytochromes P450 (rP450s), namely, CYP1A1 and CYP1B1. The aldehyde metabolite was also formed, to a lesser extent, in both human and rat liver microsomes. When comparing results from incubations with liver microsomes from 3-methylcholanthrene-treated rats (inducing CYP1A1 and CYP1B1) with those from noninduced rats, a 6-fold increase of the aldehyde metabolite was observed in the rat liver microsomes after 3-methylcholanthrene treatment. The metabolic oxidation was mimicked by the electrochemical system, and the electrochemical oxidation product was matched with the metabolite from the in vitro incubations. The electrochemical generation of the aldehyde metabolite was repeated on a preparative scale, and the proposed structure was confirmed by NMR. Trapping of the aldehyde metabolite was done with methoxyl amine. Trapping experiments with N-acetyl cysteine revealed that the aldehyde was further oxidized to an aldehyde quinoneimine species, both in the rP450 incubations and in the electrochemical system. Three additional new metabolites of amodiaquine and desethylamodiaquine were formed via rCYP1A1 and rCYP1B1. Trace amounts of these metabolites were also observed in incubations with liver microsomes from 3-methylcholanthrene-treated rats. Tentative structures of the metabolites and adducts were assigned based on liquid chromatography/tandem mass spectrometry in combination with accurate mass measurements.

Mimicry of phase I drug metabolism--novel methods for metabolite characterization and synthesis.

The extent to which electrochemical oxidation, electrochemically assisted Fenton chemistry and synthetic metalloporphines can be used to mimic cytochrome P450 catalyzed oxidations has been investigated for a large range of metabolic reactions. Most relevant metabolic oxidations can be mimicked by at least one of the three investigated systems. The EC oxidation system successfully mimics benzylic hydroxylation, hydroxylation of aromatic rings containing electron-donating groups, N-dealkylation, S-oxidation, dehydrogenation and less efficiently N-oxidation and O-dealkylation. The EC-Fenton system is able to mimic aliphatic hydroxylation, benzylic hydroxylation, aromatic hydroxylation, N-dealkylation, N-oxidation, O-dealkylation, S-oxidation and dehydrogenation. The porphine system mimics all types of reactions although the yields are low for some reactions. In conclusion, these three complementary systems can be used during the drug discovery and development of new drugs to elucidate the structure of metabolites that are difficult to characterize in biological matrices. Moreover, such techniques can replace the classical chemistry strategy, especially when synthesis is complicated or too time-consuming in order to access metabolites for further testing.

A novel fish rhabdovirus from sweden is closely related to the Finnish rhabdovirus 903/87.

A novel rhabdovirus, preliminary designated as the Sea trout rhabdovirus 28/97 (STRV 28/97), was isolated from sea trout (Salmo trutta trutta) in Sweden in 1996. The fish showed central nervous symptoms, and at the autopsy petechial bleedings in the mesenteric fat were visible. STRV 28/97 was shown to be serologically related to the vesiculotype rhabdovirus 903/87 isolated from brown trout (Salmo trutta lacustris) in Finland [1,3]. The sequences for the nucleocapsid protein, phosphoprotein, matrix protein, glycoprotein and beginning of the polymerase protein of STRV 28/97 were determined. At the amino acid level the genes were over 97% similar to virus 903/87. The nucleocapsid proteins, glycoproteins and beginning of the polymerase protein of STRV 28/97 and virus 903/87 were clustered with the vesiculoviruses and the phosphoproteins close to the vesiculoviruses in protein parsimony analysis. The matrix proteins formed a distinct clade in protein parsimony analysis.