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Journal of the Chilean Chemical Society

versión On-line ISSN 0717-9707

J. Chil. Chem. Soc. v.51 n.2 Concepción jun. 2006

http://dx.doi.org/10.4067/S0717-97072006000200016 

 

J. Chil. Chem. Soc., 51, Nº 2 (2006) , pags: 919-922

 

SYNTHESIS AND STRUCTURAL CHARACTERIZATION OF N-BENZOYL-TRYPTAMINE AND ITS NEW ANALOGUE N-SALICYLOYLTRYPTAMINE, A POTENTIAL ANTICONVULSANT AGENT

 

FRANCISCO DE ASSIS OLIVEIRA, DAVI ANTAS E SILVA, LUCINDO JOSÉ QUINTANS-JÚNIOR, STANLEY JUAN CHAVEZ GUTIERREZ, REINALDO NÓBREGA DE ALMEIDA, JOSÉ MARIA BARBOSA-FILHO AND MARIA DE FÁTIMA VANDERLEI DE SOUZA*

Laboratório de Tecnologia Farmacêutica "Delby Fernandes de Medeiros", Universidade Federal da Paraíba, Cx. Postal 5009, 58051-970, João Pessoa, PB, Brazil


ABSTRACT

N-Benzoyltryptamine (3), previously isolated from Myrtopsis myrtoidea and a new analogue, N-salicyloyltryptamine (5), were prepared by coupling tryptamine with benzoylchloride and methyl salicylate, respectively. Their structural elucidation was made by the usual spectroscopic methods and two-dimentional NMR techniques COSY, COBY and HMBC. N-salicyloyltryptamine (5) was submitted to pharmacological screening behavior, showing depressant effects.

Keywords: N-benzoyltryptamine, N-salicyloyltryptamine, two-dimentional NMR spectroscopy, anti-convulsant activity


INTRODUCTION

The isolation of reserpine in 1952 and its synthesis [1] in 1956 motivated the study of indole derivatives and their therapeutic uses. The indole nucleus is found in many bioactive substances with psychobiological and medicinal activity [2]. They include N-N-dimethyltryptamine and 5-methoxy-N-N-dimethyltryptamine, which possess psychomimetic activity [3].

N-Benzoyltryptamine (3) is a natural product, isolated for the first time from Myrtopsis myrtoidea, a plant of the Rutaceae family [4]. N-Salicyloyltryptamine (5) is a new analogue. In this work both compounds were prepared by coupling tryptamine with benzoylchloride and methyl salicylate, respectively.

In a preliminary behavioral screening in our laboratory, only (5) showed depressant effects on the Central Nervous System (CNS) in mice [5]. Therefore, we attempted to investigate the effect of (5) on seizures induced by pentylenetetrazole (PTZ).

EXPERIMENTAL

General Methods

UV spectra were recorded on a VANKEL 50 UV-Vis spectrophotometer in MeOH as solvent. IR spectra were obtained on a BOMEN MICHELSON spectrophotometer using KBr. 1H and 13C NMR spectra were recorded in C5D5N on a BRUKER AMX 400 spectrometer at 400 MHz for 1H and 100 MHz for 13C, using 1D and 2D techniques. EIMS were collected on a HP 5988A at 70 eV. For the analytical TLC, silica gel PF254 MERCK was used with a 0.25 mm layer. For Column Chromatography, silica gel MERCK 7733 was used. Tryptamine, benzoyl chloride, methyl salicylate and triethylamine were purchased from SIGMA-ALDRICH.

Preparation of N-benzoyltryptamine (3)

Tryptamine (1) (3 g) was dissolved in a solution containing 3 ml of triethylamine in 60 ml of chloroform. Then 3 ml of benzoyl chloride (2) dissolved in 30 ml of CHCl3 were added dropwise. The solution was stirred for 30 min and then the reaction mixture was washed with 2% aqueous HCl, then with water, dehydrated with anhydrous sodium sulphate, yielding an oily residue (2,5 g). This residue was chromatographed on a silica gel column, and eluted with hexane, CHCl3 and MeOH with increasing polarity and the product was crystallized from hexane-CHCl3 (9,5:0,5), affording 80% of N-benzoyltryptamine (3), white crystals, mp=131 °C (Hexane-CHCl3).


 
Figure 1. Preparation of N-benzoyltryptamine

Preparation of N-salicyloyltryptamine (5)

Tryptamine (1) (3 g) and 3 ml of methyl salicylate (4) were kept under reflux with magnetic stirring in 60 ml of methanol during 6 hours. Stirring was continued for further 24 hours at room temperature. The excess reagent was removed at reduced pressure and the oily residue (6.3 g) was chromatographed on silica gel using hexane, CHCl3 and MeOH with increasing polarity. Fractions 13-20, containing the desired product was recrystallized from CHCl3:MeOH (8:2), giving 55% of N-salicyloyltryptamine (5), which formed a white powder with mp=153 °C (CHCl3-MeOH).


 
Figure 2. Preparation of N-salicyloyltryptamine

Animals

Male Swiss mice (25-30 g) were used throughout this study. The animals were randomly housed in appropriate cages at 25 ± 2 oC on a 12 h light/dark cycle (lights on 06:00-18:00) with free access to food (Purina) and water. They were used in groups of ten animals each. Experimental protocols and procedures were approved by the Laboratório de Tecnologia Farmacêutica Animal Care and Use Committee.

Drugs

Pentylenetetrazole (PTZ), polyoxyethylenesorbitan monooleate (Tween 80) and cremophor were purchased from Sigma (USA) and Diazepam (DZP) from CEME (Brazil). Agents were injected intraperitoneally (i.p.) with a dose volume of 1ml/100g.

Anticonvulsant activity

PTZ-induced convulsions. PTZ (60 mg/kg ip) was used to induce clonic convulsions [6,7]. Mice were divided into 4 groups (n = 10). The first group was used as control and received Tween 80 (0.2%) (vehicle), while to the second group diazepam (DZP, 4 mg/kg ip) was given. The remaining groups received an injection of (5) at doses of 100 and 200 mg/kg. After 60 min of drug administration, the mice were treated with intraperitoneal PTZ at a dose of 60 mg/kg and observed for at least 30 min to detect the occurrence of the first episode of forelimb clonus. The incidence of mortality was noted until 24 h after the injection of PTZ.

Statistical analysis

The data obtained were evaluated by one-way analysis of variance (ANOVA) followed by Dunnett's t test. The incidence (%) of clonic or tonic convulsions as well as the mortality were evaluated by Fisher's Exact Test. Differences were considered to be statistically significant when p < 0.05.

RESULTS AND DISCUSSION

N-Benzoyltryptamine (3). The mass spectrum of N-Benzoyltryptamine (3) showed the molecular ion peak at m/z 264, consistent with the molecular formula C17H16N2O. The IR spectrum nmax (KBr) cm-1 exhibited the characteristic band of N-H and amide at 3407 and 3308 cm-1, respectively. The presence of the amide group was further confirmed by the band at 1649 cm-1 (C=O) and 1484 cm-1 (N-H). The UV spectrum lmax MeOH showed absorptions at 220, 277 and 288 nm, characteristic of the indole nucleus. The mass spectrum showed a fragment at 143 m/z consistent with indol-3-ethylene group and others at m/z 121 and 105 suggesting a benzamide group in the molecule.

The two-dimensional NMR spectrum (1H x 1H-COSY), confirmed the indole nucleus in the molecule exhibiting coupling between H-2 (7.39 d) and N-H (11.08 d), as well as the correlations d 7.92/7.24, consistent with aromatic hydrogens.

By heteronuclear correlated two-dimensional spectroscopy NMR 1J 1H x 13C-COBY it was possible to assign the coupling H-4'/C-4', H-2/C-2 and H-3/C-3, besides H-8 and H-7 by its coupling with C-8 (26.91 d) and C-7 (42.08 d), respectively.

The heteronuclear multiple bond correlation spectrum (2J, 3J 1H x 13C-HMBC supported absorption for H-7 (4.10 d) through interaction with C-3 and H-8 (3.35 d) which showed interactions with C-3 (2J) and C-3a (3J) (Table 1). The structure elucidation was also confirmed by comparison with literature data [4, 5, 8]. When this compound was first isolated, only one dimensional methods were performed. In the present work, all the hydrogen and carbon assignments were made unambiguously based on two-dimensional techniques.


Table 1. NMR spectral data of N-benzoyltryptamine (3) (C5D5N, 400 MHz, d, J = Hz)

The IR spectrum of N-salicyloyltryptamine (5) showed peaks at 3125 and 1661 cm-1 for the stretching of the hydrogen bond O-H and C=O, respectively. This was also supported by the mass spectrum which showed a fragment at m/e 137 related to the existence of an orthohydroxybenzamide moiety (C7H7NO2). A peak at m/z 280 refers to the molecular ion, consistent with the formula C17H16N2O2 . More details of structural fragmentation can be seen in Figure 3.


 
Figure 3. Mass fragmentation of N-salicyloyltryptamine (5)

The 1H NMR spectrum supported the attribution of orthohydroxy benzamide substituted at C-2' due to the absence of a multiplet at 8.03 (H-2') which could be seen in 3. Indeed, it shows a large singlet at 13.75 d related to a hydroxyl group, deshielded due to the formation of an intramolecular hydrogen bond with carboxyl from the amide group localized in the ortho position in the orthohydroxybenzamide moiety.

Through the two-dimensional homonuclear correlated NMR spectrum (1H x 1H-COSY 45°) it was possible to assign magnetic interaction between the indole hydrogen N-H at 11.79 d with H-2 at 7.23 d. The two-dimensional heteronuclear correlation NMR 1J 1H x 13C COBY technique showed the correlations between the indole hydrogen and its respective carbons. Interaction between H and C through 2J and 3J shown in the HMBC spectrum, allowed the unambiguously assignments of hydrogens and carbons (Table 2).


Table 3 shows in the control group that PTZ consistently induced clonic seizures in 100% of 10 mice. However, administration of 200 mg/kg (ip) of 5 increased the latency of clonic convulsions significantly different from control (p < 0.05; Dunnett's test). Similarly, the group which received DZP, the latencies significantly were prolonged and was observed a reduction of percentage for clonic seizures induced by PTZ.


The results showed that acute administration of 5 (200 mg/kg ip) on PTZ-induced convulsive seizures in mice significantly reduced (P < 0,05) the incidence of clonic seizures and mortality (as shown in Table 3). The PTZ-induced seizures, is considered as an experimental model for "generalized absence seizure" [6]. PTZ may cause convulsions by inhibiting chloride ion channels associated with GABAA receptors. However, the PTZ test is particularly sensitive to GABA mimetic drugs [7].

ACKNOWLEDGEMENTS

The authors are grateful to CNPq for financial support and Alexander I. Gray of Glasgow University for the acquisition of NMR spectra.

 

REFERENCES

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7. Almeida, R. N.; Navarro, D. S.; Assis, T. S.; Medeiros, I. A.; Thomas, G. J. Ethnopharmacol., 1998, 63 (3): 247-252.         [ Links ]

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e-mail: mfuanderlei@lrf.ufpb.br
 

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