Original article Biomimetic in vitro oxidation of lapachol: A model to predict and analyse the in vivo phase I metabolism of bioactive compounds Michael Niehues a , Valéria Priscila Barros a , Flávio da Silva Emery a , Marcelo Dias-Baruffi a , Marilda das Dores Assis b , Norberto Peporine Lopes a, * a Faculdade de Ciências Farmacêuticas de Ribeirão Preto, Universidade de São Paulo, Av. do Cafe S/N,14040-903 Ribeirao Preto, SP, Brazil b Departamento de Química, Faculdade de Filosofia, Ciências e Letras de Ribeirão Preto, Universidade de São Paulo, SP, Brazil article info Article history: Received 19 January 2012 Received in revised form 20 June 2012 Accepted 21 June 2012 Available online 28 June 2012 Keywords: Lapachol Naphtoquinones Biomimetic oxidation Jacobsen catalyst Drug metabolism Phase I metabolism abstract The bioactive naphtoquinone lapachol was studied in vitro by a biomimetic model with Jacobsen catalyst (manganese(III) salen) and iodosylbenzene as oxidizing agent. Eleven oxidation derivatives were thus identified and two competitive oxidation pathways postulated. Similar to Mn(III) porphyrins, Jacobsen catalyst mainly induced the formation of para-naphtoquinone derivatives of lapachol, but also of two ortho-derivatives. The oxidation products were used to develop a GCeMS (SIM mode) method for the identification of potential phase I metabolites in vivo. Plasma analysis of Wistar rats orally administered with lapachol revealed two metabolites, a-lapachone and dehydro-a-lapachone. Hence, the biomimetic model with a manganese salen complex has evidenced its use as a valuable tool to predict and elucidate the in vivo phase I metabolism of lapachol and possibly also of other bioactive natural compounds. Ó 2012 Elsevier Masson SAS. All rights reserved. 1. Introduction The prediction and elucidation of drug metabolites is a relevant step in drug development. One of the main elimination pathways for drugs, especially non-polar compounds, in the human body consists in the biotransformation of compounds by oxidation reactions catalysed by enzymes of the cytochrome P450 (CYP) superfamily, also known as phase I metabolism [1]. For that scenario several biomimetic enzymatic and non-enzymatic models have been described, although lastly none of these systems was able to adequately mimic the totality of CYP-mediated reactions [2]. However, a widely used chemical model is based on synthetic metalloporphyrins in presence of monooxygen donors, which basically mimic the iron(III) peroxo and iron(IV) oxo intermediates in the catalytic cycle of CYP 450 [3,4]. Several studies have reported potential oxidative metabolites of bioactive compounds, revealing that epoxidation, aliphatic and aromatic hydroxylation or oxidation of the heteroatoms are the most common reactions [5e8]. With regard to lapachol, a recent study on its biomimetic oxidation with a manganese(III) porphyrin complex and H 2 O 2 as oxidizing agent, presented five oxidation products and a selectivity in the catalysis to generate para-naphtoquinones [9]. Other biomimetic models, such as Fenton’s reagent or electrochemical induced oxidation were also shown to be interesting tools for the prediction of phase I metabolites [10,11]. In the past years metal salen complexes such as the Jacobsen catalyst (Mn(salen)), have received increasing atten- tion as biomimetic model compounds for the active site of CYP 450. With respect to their electronic structure and catalytic activities, metallosalen complexes exhibit similar features to metal- loporphyrins, catalysing usually epoxidation reactions [12]. Until now two species could be characterized by tandem mass spec- trometry as the principal oxidation intermediates of the Mn(salen) with PhIO: the oxomanganese(V) complex as the actual oxygen transfer agent in epoxidation reactions, and the dimeric m-oxo bridged with two terminal iodosylbenzene ligands which acts as reservoir species [13,14] Carbamazepine and primidone are two clinically used pharmaceuticals, on which this system was tested and successfully demonstrated as being a useful tool for the prediction of the in vivo drug metabolism [7,15]. Lapachol (1)(Fig. 1), 2-hydroxy-3-(3-methyl-2-butenyl)-1,4- naphthoquinone, is a natural occurring naphtoquinone present in several vegetal species, mainly in ones from the Bignoniaceae family * Corresponding author. Tel.: þ55 16 36024168. E-mail address: npelopes@fcfrp.usp.br (N.P. Lopes). Contents lists available at SciVerse ScienceDirect European Journal of Medicinal Chemistry journal homepage: http://www.elsevier.com/locate/ejmech 0223-5234/$ e see front matter Ó 2012 Elsevier Masson SAS. All rights reserved. http://dx.doi.org/10.1016/j.ejmech.2012.06.042 European Journal of Medicinal Chemistry 54 (2012) 804e812