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In vitro metabolism of citalopram by monoamine oxidase B in human blood

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Abstract

The metabolism of the antidepressant citalopram (CIT) by monoamine oxidase B (MAO-B) was studied in vitro. In incubations with blood of nine healthy volunteers R-(P=0.015) and S-(P=0.0034) CIT propionic acid (CITPROP) production was correlated with the number of platelets. S-CITPROP production was 5.6 times higher than R-CITPROP production and in incubations containing the MAO-B inhibitor deprenyl, racemic CITPROP production was diminished to 9.1%. To our knowledge, this is the first time that MAO-B activity in blood is shown with an antidepressant as substrate. As MAO is strongly expressed in human brain, this observation suggests that this enzymatic system may be implicated in drug metabolism in the CNS.

Introduction

The antidepressant citalopram (CIT), which belongs to the group of the selective serotonin reuptake inhibitors (SSRI), undergoes, in the liver, stereoselective biotransformation catalysed by cytochrome P-450 (EC 1.14.14.1, CYP) and monoamine oxidase (MAO, EC 1.4.3.4). The pharmacological activity of CIT resides primarily in its S-enantiomer (Hyttel et al., 1992). The main metabolic pathway of CIT in liver, N-demethylation, is catalysed by CYP2C19, CYP3A4 and CYP2D6, and leads to the stereoselective formation of demethylcitalopram (DCIT) and didemethylcitalopram (DDCIT) (Rochat et al., 1997, Olesen and Linnet, 1999). By a minor pathway, MAO catalyses the transformation of CIT to the hypothetical intermediate citalopram-aldehyde (CITALD). CITALD is then dehydrogenated, presumably by aldehyde oxidase (EC 1.2.3.1, AO), to yield a CIT propionic acid derivative, CITPROP (Rochat et al., 1998). In a detailed study about the metabolism of CIT, 75% of labelled CIT was recovered after 17 days in the urine of volunteers. Most of this radioactivity (75%) was made up of CIT, D-CIT and DD-CIT and/or their glucuroconjugated metabolites. Twelve percent was found to be the glucuronidated form of CITPROP and 7% was CIT-N-oxide (Dalgaard and Larsen, 1999). In one study, after a single dose of intravenous CIT in human volunteers, CITPROP appeared to be the main metabolite (Seifritz et al., 1996).

In human blood, only platelets contain MAO and only the MAO-B isoenzyme is expressed. Platelet MAO-B has the same amino acid sequence as human brain MAO-B (Chen et al., 1993). As in the brain, MAO is implicated in the metabolism of monoamine neurotransmitters such as dopamine, serotonin and noradrenaline, MAO activity in platelets was extensively studied as a potential biological marker in psychiatric disease (Oreland and Hallman, 1995, Wirz-Justice, 1988). MAO has only for a short time been considered as a drug and a xenobiotic metabolising enzyme (Testa, 1995, Strolin Benedetti and Tipton, 1998).

In the present study, we investigated the hypothesis of the presence of CIT metabolism by MAO-B in human blood. In an assay using kynuramine as substrate, MAO activity measured in whole blood reflected the activity measured in purified platelets (van Kempen et al., 1985). Preliminary experiments in our laboratory (not shown) yielded no evidence for a biotransformation of CIT to CITPROP in isolated platelets, even if in control experiments, kynuramine was found to be metabolised by MAO-B. Therefore, the in vitro production of R- and S-CITPROP in whole blood was measured. The production of the presumed metabolite CITALD was not investigated. Produced aldehyde metabolites are usually unstable and not detected in in vitro assays, as the biological samples often contain the aldehyde metabolising enzymes (Beedham et al., 1995).

Section snippets

Experimental procedures

The following drugs were gifts: the enantiomers of CIT (S-CIT and R-CIT), the racemates of CIT, DCIT DDCIT and CITPROP (Lundbeck AS, Copenhagen, Denmark), the internal standard S-flurbiprofen (Boots, Nottingham, UK).

The following drugs and chemicals were obtained by commercial sources: R(−)-deprenyl (selegiline) (RBI, Natick, MA, USA); clorgyline, (Sigma, Buchs, Switzerland); methyliodide (Fluka, Buchs, Switzerland).

Intravenous blood was collected from nine volunteers in EDTA containing

Results and discussion

CITPROP production in whole blood incubated with CIT was linear from 3 h to at least 9 h and at a final concentration of CIT varying from 67.5 to 500 μM. At concentrations above 1 mM CIT, haemolysis occurred and measurement of CITPROP was therefore impossible.

In Fig. 1, R- and S-CITPROP production in blood from nine subjects is shown as a function of the platelet count. Spearman correlation statistics were performed. R-CITPROP production was significantly correlated with the platelet count (R2

Acknowledgments

This study was supported by the Swiss National Science Foundation (Grants 32-42076.94 and 32-53717.98). We are grateful to Mrs K. Powell Golay for the preparation of the manuscript.

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