Ionization of beta;-carbolines in concentrated hydroxide solutions

摘要

J. CHEM. SOC. PERKIN TRANS. II 1986 Ionization of p-Carbolines in Concentrated Hydroxide Solutions M.A. MuAoz Pbrez, M. C. Carmona Guzmhn, J. Hidalgo Toledo, and M. Bal6n Almeida Departamento de Fisicoquimica Aplicada, Facultad de Farmacia, Universidad de Sevilla, Sevilla 4 10 12, Spain ~~~~~ ~~ ~ ~~ The ionization of several P-carbolines (norharman or P-carboline, harman, harmine, harmaline, and ~ tetra hydroharmine) in concentrated potassium hydroxide solutions have been examined. The pK, values, calculated using the H-acidity function, indicate that these compounds are stronger acids than indole and their acidities increase with the degree of aromaticity of the compounds in the order tetrahydro dihydro fully aromatic P-carbolines. This sequence can be interpreted in terms of resonance stabilization of the indolate anion.Because of their pharmacological properties, P-carbolines (I)-(HI) have been the subject of extensive chemical and bio- chemical investigation. lP6,tInterest in P-carboline chemistry has recently been increased by reports of their inhibitory in- fluence toward monoamine oxidase (MAO),' serotonin uptake by synaptosomas,8 and the sodium pump.' The acid-base behaviour of P-carbolines has long been of interest and much recent work has focussed on this sub- ject.I0-l5 Owing to the acidic indole NH group and the basic non-indole nitrogen atom on the P-carboline skeleton, these compounds can exist in aqueous solutions, at least, as three differently charged species, whose equilibria, exemplified for P-carboline (Ia) as the model compound, are depicted in the Scheme.Whereas the protonation equilibria on the non-indole nitrogen atom have been extensively investigated and pKa values are available,' O-' information about the dissociation of the indole NH group is very scanty. It must be noted that this group is feebly acidic 16-1 'and dissociates only in concen- trated hydroxide solution and, therefore, an acidity function must be used to provide the pK, values. In view of the paucity of data, it was of interest to examine the dissociation equilibria of P-carbolines in concentrated hydroxide solution. This paper presents pKa data on the H-acidity scale of several P-carboline derivatives of biochemical and pharmacological interest.In particular the influence of the degree of aromaticity was investigated to provide information of this effect on acidity. Experimental P-Carboline (Ia), harman (Ib), harmine hydrochloride' hydrate (Ic), and harmaline hydrochloride di hydrate (II), were pur- chased from EGA Chemie and were used as received. These compounds were pure by t.1.c. Tetrahydroharmine (111) was prepared from harmaline by reduction with sodium boro- hydride and purified by recrystallization from methanol. Stock solutions of these compounds were prepared in methanol and stored in the dark to avoid photo-oxidation. The hydroxide solutions were prepared from Merck A.R. potassium hydroxide as described by Yagil l6 and their concentrations were deter- mined by titration with standardized sulphuric acid solutions.The ionization constants were determined by the spectro- photometric method. * The necessary solutions were prepared by pipetting portions of each alkaloid solution (5amp;100 PI) into hydroxide solution (5 cm3) in sufficient amounts to give accurately measurable absorbances (final t fl-Carboline is named systematically as pyrido3,4-bindole; trivial names are used throughout. concentrations ca. 1 x ~PM).After thorough mixing, the final solutions were kept in a constant temperature bath at 25.0 f0.1 "C. In some cases where turbid solutions were formed, warming in hot water was necessary to bring about complete dissolution. Absorption spectra and absorbance measurements were made with a Lambda-5 spectrophotometer whose cell com- partment was also thermostatted at 25.0 amp; 0.1 "C.Usually, a hydroxide solution of the same concentration to the test solution was used as reference.Restilts ad Discussion As expected, deprotonation of P-carbolines significantly affects their electronic absorption spectra, since the indole NH group is part of a conjugated system. The first two columns in the Table show the position of the absorption maxima, A,,,,,., of the neutral species and their conjugated bases. As can be seen in going from dilute to concentrated hydroxide solution, the intensities of the bands of the acid forms of fully aromatic P-carbolines decreased and new absorp- tion bands or a shoulder (harmine) appeared at ca.360-380 nm where the acid forms did not absorb. These systems showed good isosbestic points. Partially reduced P-carbolines behaved differently to the other members of the series. Thus, the intense absorption band of harmaline in its acid form at 330 nm decreased upon proton- ation but no new band appeared. On the other hand, the positions of the absorption maxima of tetrahydroharmine were unaffected by changes in hydroxide concentration although a considerable increase in absorbance occurs at 3amp;340 nm upon conversion into base. However, it should be noted that the spectrum of the base form of this latter compound could not be obtained because of the turbidity of its more concentrated hydroxide solution.The ionization ratios for all the other compounds were calculated from equation (1) where dB-and dBHare the absorb- ances at the chosen wavelengths of the fully ionized species and the non-ionized species, respectively and d is the absorbance at the wavelength monitored for various intermediate hydroxide concentrations. In obtaining these data we used points corres- ponding to log 1 values within k0.75,because clear deviations from parallel behaviour were observed beyond these limits. The effect of varying the wavelength at which ionization ratio measurements were taken was checked in the case of harman. In other cases, the calculation of I was performed at only one wavelength, as listed in the Table. 1574 J. CHEM. SOC.PERKIN TRANS. II 1986 Table. Absorption characteristics and ionization data for P-carbolines in KOH at 25 "C Compound BH, hmax,/nm B-7 hmax./nm h/nm a -d log I/d H-rc PK, 338, 348 380 380 1.00 k0.02 0.991(7) 14.49 amp; 0.07 334, 346 375 334 0.93 k 0.16 0.992(6) 14.50 amp; 0.07 375 0.98 f0.03 0.986(7) 14.45 amp; 0.09 298 298, 362 (sh) 362 1.02 f0.06 0.997(9) 14.43 F 0.02 330 330 330 1.02 amp; 0.2 0.996(5) 15.34 amp; 0.03 270, 296 270, 296 276, 304 15.6 amp; 0.2 a Wavelength employed for measurement. I.e.slopes of plots of log I against H-. Correlation coefficients of these plots. The figures in parentheses are the number of points taken. (I 1 a; R' = ~2 = H (11) R' =CH3, R2 =OCH3 (111) R' = CH3 , R2 =OCH3 b; R' =CH3 , R2= H C; R' = CH, , R~ =OCH, (la) Scheme. Ionization equilibria of P-carboline In examining the ionization behaviour of P-carbolines it seemed appropriate to consider the H-acidity function stablished by Yagil16 to describe the ionization of indoles in concentrated hydroxide solution.Plots of log 1 against H-gave straight lines whose slopes and correlation coefficients are reported in the Table. It can be noted that the values of these slopes were sufficiently close to unity to permit the conclusion that the ionization of these compounds follows the H-acidity function. Hence, the pK, values were calculated using equation (2) and the data are reported in the Table. As can be seen, the pK, value of harman is fairly independent of the wavelength used for the ionization ratio determination. On the other hand, it is in very good agreement with the value of 14.6 previously reporkd.' In the case of tetrahydroharmine, since a spectrophotometric titration curve was impossible to obtain because of the afore- mentioned solubility problems, we have employed the method of Marioni and Calmon l9 to estimate its pK, value.According to this method, which only requires a part of the titration curve, the pK, value can be found, by using equation (3), from the slope of the plot of d against (dBH-d) h-, where h-= antilog (-H-). Since the error in this method may be important, absorbance measurements were carried out at two different wavelengths, the pK, of tetrahydroharmine reported in the Table being the average of these results.Inspection of the pK, data in the Table reveals that p-carbolines are stronger acids than indole (pKa 16.97). Thus, annelation of the pyridine ring to indole to give p-carboline causes an increase of the acidity by 2.5 pK, units. The higher acid strength of P-carboline compared with indole follows the trend of the five-membered heterocycles pyrrole, imidazole, and pyrazole and their benzologues indole, benzimidazole, and indazole; the extension of conjugation raises the acidity due to resonance stabilization of the anions.' However, the increase of acidity in going from indole to 0-carboline compared with the increase of 0.5 pKa units observed between pyrrole and indole seems exceptionally high.Unfor- tunately, the different effects of benzene and pyridine annelation cannot be discussed because the acid strength of the parent compound 6-azaindole is unknown. By supposing that annel- ation effects of benzene and pyridine rings are additive, a pK, value of ca. 15 could be predicted for 6-azaindole. It should be also noted that the acidities of P-carbolines increase with the degree of aromaticity of the p-carboline skeleton in the order tetrahydro dihydro fully aromatic P-carbolines as would be expected for the resonance stabiliz- ation of the indolate anions. Finally, we can briefly comment on the rather small influence of methoxy and methyl substitution at C-7 and C-1 of the p-carboline ring, because these substituents do not make possible additional resonance forms for the anion.In contrast, an exceptionally high increase in acidity occurred with the sub-stitution of a methyl group on the pyridine nitrogen atom, as illustrated by 2-methyl-P-carboline, whose pK, is 10.9.'' In this case it is evident that the positively charged quaternary nitrogen atom is an additional factor for stabilization of the anions. J. CHEM. SOC. PERKIN TRANS. II 1986 Acknowledgements We gratefully acknowledge financial support from the Cornision Asesora de Investigacion Cientifica y Tecnica of Spain. References 1 R.A. Abramovitch and I. D. Spencer, Ado. Heterocycl. Chem., 1964, 3, 79. 2 K. Stuart and R. Woo-Ming, Heterocycles, 1975, 3, 223. 3 R.H.F. Manske, lsquo;The Alkaloids,rsquo; Academic Press, New York, 1965, vol. VIII, pp. 47-53. 4 V. Snieckus, lsquo;The Alkaloids,rsquo; ed. R.H. F. Manske, Academic Press, New York, 1968, vol. XI, pp. 1-40. 5 D. H. Aarons, G. V. Rossi, and R. F. Orzechowski, J. Pharm. Sci., 1977,66, 1244. 6 R. G. Rahwan, Toxicol. Appl. Pharmacol., 1975, 34, 3. 7 B. T. Ho, W. M.McIsaac, K. E. Walker, and V. Estevez, J. Pharm. Sci., 1968, 57, 269, 1368. 8 N. S. Buckholtz and W. 0.Boggan, LiJe Sci., 1977, 20, 2093. 9 M. Canessa, E. Jaimovich, and M. de la Fuente, J. Memhr. Bio(., 1973, 13, 263. 10 R.Sakuros and K. P. Ghiggino, J. Photochem., 1982, 18, I. 11 0.S. Wolfbeis and E. Furlinger, Monatsh. Chem., 1982, 113, 509. 12 0.S. Wolfbeis and E. Furlinger, 2. Phys. Chem. Neue Folge, 1982, 129, 171. 13 K. T. Douglas, R. K. Sharma, J. F. Walmsley, and R.C. Hider, Mnl. Pharmacol., 1983, 23, 614. 14 F. Tomas Vert, I. Zabala Sanchez, and A. Olba Torrent, J. Photochem., 1983, 23, 355. 15 F. Tomas Vert, I. Zabala Sanchez, and A. Olba Torrent, J. Photachem., 1984, 26, 285. 16 G. Yagil, J. Phys. Chem., 1967, 71, 1034. 17 G. Yagil, Tetrahedron, 1967, 23, 2855. 18 A. Albert and E. P. Serjeant, lsquo;The Determination of Ionization Constants,rsquo; Chapman and Hall, London, 1971, 2nd edn. 19 P. Maroni and J. P. Calmon, Bull. Sac. Chim. Fr., 1964, 519. Received 6th November 1985; Paper 51 1950