Journal of Pharmaceutical and Biomedical Analysis
MiscellaneousAssignment of the absolute configuration at the sulfur atom of thioridazine metabolites by the analysis of their chiroptical properties: The case of thioridazine 2-sulfoxide
Introduction
Thioridazine (THD), a commonly prescribed phenotiazine neuroleptic drug, used for the treatment of schizophrenia and other psychiatric disorders, is extensively biotransformed in the organism producing as main metabolites the sulfoxides at positions 2 and 5, and the sulfone derivate at position 2 [1], [2].
THD is currently administered as a racemate, although significant differences have been observed for the activity of this drug depending on the stereochemistry; the antipsychotic effect of racemic THD is mainly associated with (R)-THD [2]. In the metabolic process a new stereocenter is formed at the sulfur atom in thioridazine-2-sulfoxide (THD-2-SO) and thioridazine-5-sulfoxide (THD-5-SO) metabolites (Fig. 1). So for each of these two compounds two pairs of enantiomers are possible: (Rc, Rs) (Sc, Ss) and (Rc, Ss) (Sc, Rs), the first and second pairs are diastereoisomers. The (R) and (S) configurations of THD and their metabolites are related to the chiral carbon at position 2 in the piperidyl ring (c) and to the sulfur atom of the sulfoxide metabolites (s).
Considering that THD-2-SO and THD-2-SO2 metabolites are also therapeutically active [3], [4], [5], but with undesirable side effects that can be attributed to specific stereoisomers, the assignment of the absolute configuration to each of these streoisomeric forms of the drug and of its metabolites should allow reliable information on the relationship structure/activity in clinical and/or toxicological studies. In addition, THD-5-SO metabolite contributes to the cardio toxicity of the drug [6], [7].
We report here the determination of the absolute configuration at the sulfur atom of THD-2-SO. The diastereoisomers were separated by non-chiral HPLC and referred as fast-eluting (FE) THD-2-SO and slow-eluting (SE) THD-2-SO, based on their chromatographic behaviour [1], [2]. The enantiomers of the fast-eluted couple (FE) have been separated by HPLC on Chiralpack AS, recording the circular dichroism (CD) spectra for the two collected enantiomeric fractions.
To simulate the experimental electronic CD spectrum of THD 2-SO, we arbitrarily assumed S absolute configuration for the sulfur atom and removed the chain linked to the N atom to reduce the numbers of conformers, without neglecting significant chromophores. Two stable conformers have been found in the gas phase: using the above two conformers as input geometries and the TDDFT/B3LYP/6-31G* method [8], [9], [10] the theoretical CD spectrum (after Boltzmann averaging) was obtained, with the simulated curve (S) AC of the sulfur atom that corresponds to the first eluted peak.
Section snippets
Chemicals
rac-Thioridazine, thioridazine-2-sulfone and thioridazine-2-sulfoxide, were kindly supplied by Novartis Pharma AG (Basel, Switzerland). The THD-2-SO standard sample consisted mainly of the racemic mixture of the enantiomers of the fast-eluted (FE) diastereoisomer, according to the separation procedure reported in the literature [1]. Stock standard solutions were prepared in methanol at concentrations of 200 and 400 μg mL−1 and were stored at −20 °C in the absence of light. HPLC-grade hexane,
Stereochemical characterization of THD and of its metabolites by enantioselective HPLC and circular dichroism
Enantioselective HPLC was successfully applied for the stereoisomeric resolution of THD and of its main metabolites. In particular, enantiomeric fractions of THD, THD-2SO2 and of both the diastereoisomers of THD-2SO were obtained at preparative level, in order to carrying out the CD spectra of each fraction, and then to establish the relationship between sign of the CD spectra and absolute configuration. Thus different chiral stationary phases were used in order to have the enantioselectivity
Conclusions
The Sc, Ss and Rc, Rs absolute configuration was assigned to the first and second eluted fractions of THD-2SO, respectively. The satisfactory simulation of the (Sc, Ss)-THD-2SO CD spectrum by the calculated CD spectrum of (Ss)-1, demonstrates that the CD spectrum depends primarily on the absolute configuration at the sulfur atom. This result, which concerns one of the main metabolites of such an important drug, is really relevant for these studies because this information allows a deeper
Acknowledgments
The authors are grateful to Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP), Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq), and to Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) for financial support and for the grant of research fellowships. Thanks are also due to Università di Bologna (C.B.) and to Università della Basilicata (G.M., C.R.) for financial support.
References (22)
- et al.
Determination of the enantiomers of thioridazine, thioridazine 2-sulfone, and of the isomeric pairs of thioridazine 2-sulfoxide and thioridazine 5-sulfoxide in human plasma
J. Chromatogr. B
(1995) - et al.
Antagonism by neuroleptics of neurotrasmitter receptors of normal human brain in vitro
Eur. J. Pharmacol.
(1984) - et al.
Plasma concentrations of thioridazine metabolites and ECG abnormalities
J. Pharm. Sci.
(1978) - et al.
Cardiotoxicity of thioridazine and two stereoisomeric forms of thioridazine 5-sulfoxide in the isolated perfused rat heart
Toxicol. Appl. Pharmacol.
(1986) - et al.
Stereoselective analysis of thioridazine-2-sulfoxide and thioridazine-5-sulfoxide: an investigation of rac-thioridazine biotransformation by some endophytic fungi
J. Pharm. Biomed. Anal.
(2008) - et al.
Synthesis, receptor binding and functional studies of mesoridazine stereoisomers
Bioorg. Med. Chem. Lett.
(2004) - et al.
Plasma levels of the enantiomers of thioridazine, thioridazine 2-sulfoxide, thioridazine 2-sulfone, and thioridazine 5-sulfoxide in poor and extensive metabolizers of dextromethorphan and mephenytoin
Clin. Pharmacol. Ther.
(1996) On the serum concentrations and antipsychotic effects of thioridazine, thioridazine side-chain sulfoxide and thioridazine side-chain sulfone in chronic psychotic patients
Curr. Ther. Res. Clin. Exp.
(1977)Interactions of neuroleptic metabolites with dopaminergic, alpha adrenergic and muscarinic cholinergic receptors
J. Pharmacol. Exp. Ther.
(1981)Renaissance in chiroptical spectroscopic methods for molecular structure determination
Chem. Rec.
(2007)
The current state of ab initio calculations of optical rotation and circular dichroism spectra
J. Phys. Chem. A
Cited by (10)
Exploring the capabilities of TDDFT calculations to explain the induced chirality upon a binding process: A simple case, 3-carboxycoumarin
2013, Journal of Molecular StructureCitation Excerpt :The TDDFT calculations were found suitable for predicting the absolute configuration of chiral organic compounds, many papers and reviews being devoted to this subject [1–6].
TDDFT studies on electronic structures, chiroptical properties and solvent effect on the CD spectra of diphosphonate-functionalized polyoxomolybdates
2013, Journal of Molecular Graphics and ModellingCitation Excerpt :Theoretical approaches are very helpful to explain the CD spectra of chiral isomers [39,40]. Theoretically reproducing and predicting the ECD spectrum are helpful for specifying the absolute molecular conformation in solution and the origins of the ECD spectrum [41–43]. Ziegler and co-workers calculated the ECD spectra of metal complexes and assigned their origins in detail [44].
Assignment of absolute configuration of sulfinyl dilactones: Optical rotations and <sup>1</sup>H NMR experiment and DFT calculations
2011, Journal of Molecular StructureCitation Excerpt :Therefore, ab initio calculations of chiro-optical and spectroscopic features were used to assist in the AC assignment. Density functional theory (DFT) has been extensively exploited to calculate chiro-optical properties such as optical rotation (OR) [9–13] and circular dichroism (CD) [14], and increasingly, consistency has been obtained between theoretical and experimental results [10]. So we used DFT to calculate OR at the sodium D line wavelength and then compared with experimental results.
TDDFT studies on the determination of the absolute configurations and chiroptical properties of strandberg-type polyoxometalates
2013, Journal of Physical Chemistry A