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Langmuir 1996, 12, 2956-2960
Chiral Lipophilic Ligands. 3.† Control of Enantioselectivity in Copper(II)-Catalyzed Cleavage of r-Amino Acid Esters by Aggregate Morphology Marco C. Cleij,‡,§ Paolo Scrimin,| Paolo Tecilla,*,§ and Umberto Tonellato*,§ Department of Chemical Sciences, University of Trieste, Via Giorgieri, 1-34127 Trieste, Italy, and Department of Organic Chemistry and Centro CNR Meccanismi di Reazioni Organiche, University of Padova, Via Marzolo, 1-35131 Padova, Italy Received December 12, 1995. In Final Form: March 19, 1996X The cleavage of the enantiomers of the p-nitrophenyl esters of phenylglicine (PhgPNP) was studied using chiral Cu(II) complexes of ligands 2 [2-N-R1-N-(1-R2-2-hydroxyethyl)aminomethylpyridine] as catalyst. Lipophilic ligands 2a,b (2a, R1 ) n-dodecyl, R2 ) methyl; 2b, R1 ) n-dodecyl, R2 ) isopropyl) were studied in aggregates of nonfunctional surfactants forming micelles (cationic, anionic, nonionic) or vesicles. With respect to the nonmicellar complex 2c‚Cu(II) (2c, R1 ) methyl, R2 ) isopropyl), large rate accelerations (up to 400 times) and moderate (up to 11) enantioselectivities (as rate ratio between the faster and slower enantiomer) were found in cationic micelles. On the contrary, large inhibition was observed in anionic micelles, whereas in the nonionic ones the kinetic effects were negligible. In cationic vesicles the enantioselectivities are strongly influenced by the fluidity of the aggregate bilayer: remarkably large values (up to 26) were observed below the main phase transition temperature, Tc. The results were explained on the basis of different reaction mechanism due to the compartmentalization of the reacting species (a ternary complex ligand/Cu(II)/substrate) in different loci of the aggregate. It is suggested that the more lipophilic diastereomeric complex reacts with the substrate with attack of the Cu(II)-bound alkoxide of the ligand while the more hydrophilic one reacts with attack of a metal-bound hydroxyl of an exogenous water molecule. The first mechanism is both faster and more enantioselective.
Introduction It is well accepted that kinetic effects for reactions in micellar (or vesicular) aggregates can mainly be ascribed to changes of local concentrations (i.e., in the aggregate pseudophase) and compartmentalization of reacting species.1 However, intriguing results of quite impressive modifications of stereoselectivity as the outcome of subtle modifications of aggregate structure due to the change of surfactant composition have been reported.2 It appears that, at least from the point of view of stereoselectivity, the aggregate morphology plays a crucial role in controlling the reaction rates. These morphological changes are the focus of the investigation described in this paper where amphiphilic Cu(II) complexes have been used as catalyst of the hydrolysis of activated esters. We have recently reported3 on the enantioselective cleavage of the p-nitrophenyl esters of several R-amino acids (phenylalanine (PhePNP), phenylglycine (PhgPNP), and leucine (LeuPNP)) catalyzed by metallomicelles made up of complexes of lipophilic chiral ligands 1a and 2a-b with transition metal ions. The results obtained showed two relevant features of the aggregate systems as compared to monomeric and water soluble complexes: (i) higher enantioselectivities and (ii) † Part 2: Scrimin, P.; Tecilla, P.; Tonellato, U. Tetrahedron 1995, 51, 217. ‡ European E.C. Research Fellow (1993-1994). § University of Padova. | University of Trieste. X Abstract published in Advance ACS Abstracts, May 15, 1996.
(1) (a) Bunton, C. A.; Nome, F.; Quina, F. H.; Romsted, L. S. Acc. Chem. Res. 1991, 24, 357. (b) Bunton, C. A.; Savelli, G. Adv. Phys. Org. Chem. 1986, 22, 213. (2) (a) Ueoka, R.; Moss, R. A.; Swarup, S.; Matsumoto, Y.; Strauss, G.; Murakami, Y. J. Am. Chem. Soc. 1985, 107, 2185. (b) Ueoka, R.; Matsumoto, Y.; Yoshino, T.; Watanabe, N.; Omura, K.; Murakami, Y. Chem. Lett. 1986, 1743. (c) Ueoka, R.; Matsumoto, Y.; Moss, R. A.; Swarup, S.; Sugii, A.; Harada, K.; Kikuchi, J.; Murakami, Y. J. Am. Chem. Soc. 1988, 110, 1588. (3) Scrimin, P.; Tecilla, P.; Tonellato, U. J. Org. Chem. 1994, 59, 4194.
S0743-7463(95)01533-2 CCC: $12.00
Chart 1
higher reactivities. Furthermore, the micellar complex is remarkably more efficient while the monomeric complex is less efficient than aquo Cu(II): as a matter of fact, differences in rates exceeding 3 orders of magnitude were observed between micellar and nonmicellar complexes.3,4 The dichotomy between aggregate and nonaggregate systems was explained to be the result of a different mode of action. Evidence was reported that the high catalytic efficiency of the micellar systems containing the lipophilic ligands and Cu(II) was due, as schematically shown in Scheme 1, to the formation of a ternary complex ligand/ metal ion/substrate (A) in which the ligand’s hydroxyl, (4) De Santi, G.; Scrimin, P.; Tonellato, U. Tetrahedron Lett. 1990, 31, 4791.
© 1996 American Chemical Society
Chiral Lipophilic Ligands Scheme 1
activated by the metal ion, acts as a very effective nucleophile to give a transacylation intermediate (B). This eventually undergoes a metal-ion-promoted hydrolysis thus ensuring a relatively efficient turnover of the catalyst. On the other hand several pieces of evidence indicate that outside any aggregate, although the ternary complex is still formed, the cleavage is brought about by a metalion-bound water molecule and not by the hydroxyl; the chiral arm of the ligand plays a distant role and, as a result, both reactivity and enantioselectivity are modest. The remarkable changes in the mode of action and the relevant consequences in terms of reactivity and enantioselectivity determined by the aggregation prompted us to further investigate the effect of the aggregates in order to realize more efficient and selective catalytic systems and shed further light on the source of the enantioselection. In particular we examined the role of charge and head group size in micellar systems and, considering the morphological effects mentioned above, the influence of the change of the hosting aggregate from rather disordered micelles to a more ordered vesicular membrane. It should be also pointed out that recent results obtained in our laboratory4,5 strongly suggest that aggregation of metalloamphiphiles may lead to substantial modification of the coordination sphere of the metal ion. Results and Discussion The Ligands and the Aggregates. The present study was mainly focused on the enantioselective reactivity of Cu(II) complexes of ligands 2a-b. Ligand 2c with a methyl group in the place of the long paraffinic chain, was also investigated to compare the monomeric (nonaggregate) system to its aggregate counterparts. All ligands, except 2c, have been previously described.3 Lipophilic ligands 1 and 2 are little dispersible in neutral aqueous solutions; their Cu(II) complexes are more soluble, although not much above their critical micelle concentrations (cmc), and they are easily taken up by nonfunctional cationic micelles such as those made of CTABr. Indeed, most of the kinetic measurements previously reported were made for comicellar solution using a 10:1 molar excess of CTABr over the complex to ensure the presence of micellar aggregates well above the cmc, once ascertained that the reactivity and enantioselectivity data were not much affected by comicellization. On the basis of this (5) Bunton, C. A.; Scrimin, P.; Tecilla, P. J. Chem. Soc., Perkin 2 1996, 419.
Langmuir, Vol. 12, No. 12, 1996 2957 Table 1. Observed Pseudo-First-Order Rate Constants, kR and kS (s-1) and Enantioselectivity Ratio, ER, for the Cleavage of PhgPNP by Ligands (S)-2a-c with Cu(II)a entry
ligand
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18
2c 2bb 2b 2bc 2b 2b 2b 2b 2a 2a 2a 2bd 2bd,e 2bd 2bd,e 2bd 2bd,e
cosurfactant
kR
kS
ER
CTABr (3a) CTABr (3a) 3b 3c 3d 3e CTABr (3a) Brij-35 SDS 2C16 2C16 2C18 2C18 5 5
0.11 0.095 26.5 13.2 2.6 25.3 32.4 40.0 8.8 7.7 0.35 2.8 × 10-3 6.3 2.26 27.7 6.5 40.7 5.9
0.12 3.0 1.14 1.3 3.0 3.8 4.4 0.95 1.1 0.3 1.2 × 10-3 1.0 0.14 1.4 0.25 2.2 0.41
1.2 8.8 11.6 2.0 8.4 8.5 9.1 9.2 7.0 1.2 2.3 6.3 16.1 19.7 26.0 18.5 14.4
a For conditions see text. b [2b] ) [Cu(II)] ) 7.5 × 10-4 M. c In water/DMSO 2.5:1. d [MES] ) 0.025 M. e [2b] ) 1 × 10-4 M, [cosurfactant] ) 1 × 10-3 M, 2.5 °C.
background and further evidence (vide infra), the present kinetic study was mainly carried out with the catalysts embedded in micellar or vesicular inert matrices. The micellar ones were made of cationic surfactants, 3; nonionic (Brji-35) and anionic (SDS) surfactants were also tested. Vesicles were made of 4a (2C16), 4b (2C18), and 5 and prepared by sonication using standard procedures. The Effect of Micellization. Table 1 shows the kinetic data observed for the cleavage of PhgPNP, in the absence and presence of monomeric, micellar or comicellar, and vesicular Cu(II) complexes of (S)-2, under the following conditions (unless otherwise indicated): [Cu(II)] ) 8.3 × 10-5 M, [ligand] ) 2 × 10-4 M, [cosurfactant] ) 2 × 10-3 M, [PhgPNP] ) (1-2) × 10-5 M, MES (0.05 M) buffer, pH ) 5.5, 25 °C. The kinetic results obtained with ligands 2 are essentially analogous to those obtained with ligands 1 except for the fact that in the case of the nonmicellar complex 2c‚Cu(II) (entry 2) the observed kinetic (and enantioselectivity) effects are quite small at variance with the pronounced inhibition observed in the case of 1b‚Cu(II). Furthermore with 2c‚Cu(II) the faster reacting enantiomer is that with the same absolute configuration of the catalyst while with all the other ligands the faster enantiomers were consistently those of opposite configuration. The data of Table 1 allow a detailed picture of the micellization or comicellization effects: (i) micellization results in a substantial increase of the rate and the ER6 values (see entries 2 and 4) and the effect is reversed on going from aqueous micellar solutions to water/DMSO (entries 4 and 5) where micelles no longer exist; (ii) although a direct comparison of rate data is not available due to the limited solubility of 2b above the cmc, the enantioselectivity is not much affected by comicellization with cationic cosurfactants (entries 3 and 4); (iii) the ER values are not very sensitive to the bulkiness of headgroups of the cationic cosurfactants 3; on increasing their size7 (entries 4 and 6-9) the rate increases but the kinetic benefits are relatively similar for both enantiomers or only slightly larger for the slower one; (iv) in the presence of the nonionic cosurfactants Brij-35 (entry 11) the (6) ER: Enantioselectivity ratio is kR/kS or kS/kR where R and S denote configuration of the substrate and k are observed pseudo-first-order rate constants measured under the conditions reported. (7) Germani, R.; Savelli, G.; Spreti, N.; Cerichelli, G.; Mancini, G.; Bunton, C. A. Langmuir 1993, 9, 61.
2958 Langmuir, Vol. 12, No. 12, 1996
Figure 1. Fluorescence polarization vs temperature profiles for covesicular blends 2b/2C16 (b), 2b/2C18 (9), and 2b/5 (O). [(S)-2b] ) 5 × 10-5 M, [cosurfactant] ) 5 × 10-4 M, [1,6-diphenyl1,3,5-hexatriene] ) 5 × 10-6 M, MES buffer 0.025 M, pH 5.5.
observed rates are virtually similar to those recorded for solutions containing only the free metal ion or those with the monomeric complex with 2c‚Cu(II) (entries 1 and 2), and the enantioselectivity is vanishingly small; (v) in the presence of anionic SDS (entry 12) there is a dramatic drop of reactivity and the ER is small. Micellization with cationic CTABr results, in the case of PhgPNP enantiomers, in a rate increase up to a factor of 10 for the slower reacting enantiomer (S), whereas for the faster one the acceleration is approximately 10 times as large. The large decrease in rate in the case of anionic SDS can be explained by assuming partial depletion of the complex by the cosurfactant and by the drop of the “local” pH at the surface of anionic micelles. Furthermore the competition of SDS with the substrate for the formation of the ternary complex is certainly possible. Somewhat surprisingly, comicellization with the nonionic surfactant does not increase the reactivity relative to that in nonaggregate systems, thus indicating that electrostatic factors play a role in both reactivity and enantioselectivity. The Effect of Vesicles. The rate of cleavage of PhgPNP was investigated in covesicles made up of the ligand (S)-2b and Cu(II) and surfactants 4a-b and 5 employing in each case a 1:10 ratio [ligand]/[surfactant] and prepared following a standard protocol (see Experimental Section). Preliminary to the kinetic study, we characterized the resulting covesicles and determined their (gel-to-liquid crystal) transition phase temperature (Tc) and hydrodynamic diameters in MES buffer, pH 5.5. Dynamic light scattering measurements showed the presence of vesicular aggregates with a rather broad size distribution. The mean hydrodynamic diameters were (Å) 590 (2C16, 4a), 600 (2C18, 4b), and 520 (5). Fluorescence polarization measurements8 in the temperature range 2-50 °C allowed the Tc (°C) to be defined as follows: 15.0 (2C16), 28.4 (2C18), and 42.4 (5). As shown by the plot of the polarization data vs temperature of Figure 1, the phase transition occurs in a remarkably broad temperature interval. The kinetic experiments carried out under the conditions [Cu(II)] ) 8.3 × 10-5 M, [(S)-2b] ) 0.1 mM, [cosurfactant] ) 1 mM, MES (25 mM), and pH ) 5.5, using PhgPNP enantiomers as substrates, showed a rather dramatic effect of the fluidity of the membrane on the reactivity (8) Using 1,6-diphenyl-1,3,5-hexatriene as a probe: (a) Andrich, M. P.; Vanderkooi, J. M. Biochemistry 1976, 15, 1257. (b) Lentz, B. R. Chem. Phys. Lipids 1989, 50, 171.
Cleij et al.
Figure 2. log kψ vs 1/T profiles for the cleavage of (R)-PhgPNP (b) and (S)-PhgPNP (O) by covesicles of (S)-2b/2C16 and Cu(II). The dotted lines indicate the phase transition interval. [(S)2b] ) 1.0 × 10-4 M, [2C16] ) 1 × 10-3 M, [Cu(II)] ) 8.3 × 10-5 M, MES buffer 0.025 M, pH 5.5.
Figure 3. log kψ vs 1/T profiles for the cleavage of (R)-PhgPNP (b) and (S)-PhgPNP (O) by covesicles of (S)-2b/2C18 and Cu(II). The dotted lines indicate the phase transition interval. [(S)2b] ) 1.0 × 10-4 M, [2C18] ) 1 × 10-3 M, [Cu(II)] ) 8.3 × 10-5 M, MES buffer 0.025 M, pH 5.5.
and particularly on the enantioselectivity. Entries 1318 of Table 1 show rate data collected at 25 and 2.5 °C under conditions comparable to those of the other aggregates. At 25 °C the covesicular systems with 2C16 and 2C18 are in the phase transition regime while at 2.5 °C both are in the gel state. The effect on the reactivity is better illustrated by means of the Arrhenius-type plot log kψ vs 1/T shown in Figures 2 and 3 for covesicular blends with 2C16 and 2C18, respectively. In the case of the faster PhgPNP enantiomer (R), the rate increases with temperature up to the lower limit of the phase transition interval and then decreases during phase transition and climbs up again at higher temperature (Figure 2). In the case of the slower enantiomer, the plot shows a simple inflection around Tc. Although, due to the higher Tc, the temperature effects after the transition phase could not be fully appreciated, a similar trend was observed for covesicles with 5. With this blend, as well as with that of 2C18 (Figure 3), the effects are more pronounced and the difference observed for the two enantiomers is much more impressive than in the case of covesicles with 2C16. Figure 4 shows the changes of the ER values for the three
Chiral Lipophilic Ligands
Langmuir, Vol. 12, No. 12, 1996 2959 Chart 2
Figure 4. Dependence of ER with temperature for the cleavage of the two enantiomers of PhgPNP by covesicles of (S)-2b with 2C16 (b), 2C18 (9), and 5 (O). [(S)-2b] ) 1.0 × 10-4 M, [cosurfactant] ) 1 × 10-3 M, [Cu(II)] ) 8.3 × 10-5 M, MES buffer 0.025 M, pH 5.5.
types of covesicles as a function of the temperature. Thus, on decreasing the temperature from 45 °C (above phase transition for all systems) to 2.5 °C (below Tc for all systems), i.e., on going from liquid-crystal to gel-like-type bilayers, the ER values increase from (as approximately evaluated from Figure 4) 3 to 16 (2C16), from 5 to 26 (2C18), and from 7 to 15 (5). Although less important, we observed that the concentration of MES used as buffer influences the rate and particularly the enantioselectivity: thus in the case of covesicles of 2C18 at 25 °C, on increasing [MES] from 25 to 50 mM, the ER value increases from ca. 20 to 27. The highest observed enantioselectivity ratio of approximately 26 is one of the largest values ever recorded in the hydrolytic cleavage of R-amino acid esters employing chiral functional micelles. Higher values of enantioselectivity or diastereoselectivity were only found using lipophilic nucleophiles with several chiral centers embedded in nonfunctional aggregates.9,10 There are no precedents of such impressive changes in the ER values as a function of the order and packing of the aggregates in which the catalyst is embedded.11 Typically only small ER changes have been associated with Tc.12 The Source of Enantioselection. We have ascribed the large difference in reactivity and enantioselectivity observed between the monomeric and aggregate systems to a change in the mode of action within the ternary complex ligand/Cu(II)/substrate.3 The negligible difference between 2-(aminomethyl)pyridine (log KM ) 9.5) and 2-[[N,N-(dihydroxyethyl))amino]methyl]pyridine (log KM ) 9.2) in binding of Cu(II) ions13 suggest that co-ordination of the alcoholic group to the metal center has little (9) (a) Moss, R. A.; Sunshine, W. L. J. Org. Chem. 1974, 39, 1083. (b) Moss, R. A.; Taguchi, T.; Bizzigotti, G. O. Tetrahedron Lett. 1982, 23, 1985. (c) Moss, R. A.; Hendrickson, T. M.; Ueoka, R.; Kim, K. Y.; Weiner, P. K. J. Am. Chem. Soc. 1987, 109, 4363. (10) (a) Ihara, Y.; Igata, K.; Okubo, Y.; Nango, M. J. Chem. Soc., Chem. Commun. 1989, 1900. (b) Ihara, Y. J. Chem. Soc., Perkin Trans. 2 1980, 1483. (c) Ihara, Y.; Okamoto, M.; Kawamura, Y.; Nakanishi, E.; Nango, M.; Koga, J. J. Chem. Soc., Perkin Trans. 2 1987, 607. (d) Ihara, Y.; Asakawa, S.; Igata, K.; Matsumoto, Y.; Ueoka, R. J. Chem. Soc., Perkin Trans. 2 1991, 543. (11) With the exception of the impressive effects cited in the introduction (ref 2) due to a change of aggregate morphology with surfactant composition. (12) See for instance: (a) Weijnen, J. G. J.; Koudijs, A.; Tap, P. G. J. A., Engbersen, J. F. J. Recl. Trav. Chim. Pays-Bas 1993, 112, 525. (b) Murakami, Y.; Nakano, A.; Yoshimatsu, A.; Fukuya, K. J. Am. Chem. Soc. 1981, 103, 728.
thermodynamic relevance.14 Accordingly, in the ternary complex involved as the active species in our systems it is possible that the equilibrium outlined in Chart 2 is controlled by small modification of the environment. Kinetic evidence with monomeric and water soluble ligand 1b shows that its reactivity is not affected by methylation of the hydroxyl while the same modification remarkably affects the reactivity of 1a in aggregates. Hence, in water, complex 6a largely prevails over 6b while in aggregates the opposite situation is achieved. The explanation previously offered3 of a change of the position of the equilibrium between complexes 6a and 6b appears to be supported by the present findings. Reactions in environments favoring 6a are both slower and poorly enantioselective while those in environments favoring 6b are faster and more sensitive to steric effects. Previous studies of the cleavage of carboxylate esters by Cu(II) complexes in metallomicelles15 suggest that a Cu(II)-bound deprotonated water is less reactive than a Cu(II)-bound alkoxy group being part of the ligand. Furthermore, the complex with the unbound chiral arm bearing the alcoholic group is clearly less stereoselective because the hydroxyl is not directly involved in the reaction center. It is conceivable that the two different diastereomeric complexes formed by interaction of the two enantiomeric substrates with the chiral complex differ for the relative contribution of species 6a and 6b, i.e., for the position of the equilibrium indicated in Chart 2. Likely this is due to their different hydrophobicity16 so that they are confined in different loci of the aggregate which are characterized by different hydration. This confinement should be more effective in vesicles in their gel state and less in vesicles in their fluid states17 or in micelles. Accordingly, the difference in reactivity of the two enantiomers would be due to the relative amount of reaction going via the more reactive complex 6b or the less reactive complex 6a. Following this line of reasoning the fact that the rate of the more reactive enantiomer is slowed down by the increase of fluidity of the membrane is due to a shift of the equilibrium to the left (6a) as a consequence of the increased hydration of the bilayer. It is known that permeability of vesicular membranes to water is greatly enhanced above the Tc.18 The previously reported and present results support the above explanation although, admittedly, there is no direct evidence of this mechanistic dichotomy. As a matter of fact, the ternary complex formed is very labile and not amenable to any characterization in the aggregates we have used. (13) Damu, K. V.; Shaikjee, S. M.; Michael, J. P.; Howard, A. S.; Hancock, R. D. Inorg. Chem. 1986, 25, 3879. (14) Hancock, R. D.; Martell, A. E. Chem. Rev. 1989, 89, 1875. (15) Scrimin, P.; Tecilla, P.; Tonellato, U. J. Org. Chem. 1991, 56, 161. (16) Differences in hydrophobicity between diastereomeric Cu(II) complexes of ligands derived from amino acids have been reported: Nakon, R.; Angelici, R. J. J. Am. Chem. Soc. 1974, 96, 4178. Sigel, H.; Martin, R. B. Chem. Rev. 1982, 82, 385. (17) Harada, S.; Takada, Y.; Yasunaga, T. J. Colloid Interface Sci. 1984, 101, 524. (18) (a) Deamer, D. W.; Brannhall, J. Chem. Phys. Lipids 1986, 40, 167. (b) Disalvo, E. A. Adv. Colloid Interface Sci. 1988, 29, 141.
2960 Langmuir, Vol. 12, No. 12, 1996
Conclusion Our anticipation3 that the morphology of the aggregate, in particular the increase of its order and effectiveness in compartmentalization of substrates, would increase ER of amphiphilic Cu(II) complexes in hydrolytic processes has been verified by the present investigation. The charge of nonfunctional surfactants used plays a role in the catalytic efficiency of the systems and cationic aggregates provide the best environment. The difference in reactivity between the two enantiomers in the vesicular system (in the gel state) is such that kinetic separation of a racemic mixture of R-aminoesters appears, at least in principle, feasible. Experiments aimed at this goal are currently pursued in our laboratory. Experimental Section General Methods and Materials. 1H-NMR spectra were recorded on a 250 MHz Brucker AC 250 F spectrometer and chemical shifts in ppm are reported to internal Me4Si. The enantiomeric purity of the ligand synthesized was checked as described.3 Cu(NO3)2 was an analytical grade product and metal ion stock solutions were titrated against EDTA following standard procedures.19 2-Morpholinoethanesulfonic acid (MES), used as the buffer, was a Fluka product used as received. n-Hexadecyltrimethylammonium bromide (CTABr, 3a) and di-n-octadecyldimethylammonium bromide (4b) were analytical grade commercial products. n-Hexadecyltriethylammonium bromide (3b), n-hexadecyltripropylammonium bromide (3c), n-hexadecyltributylammonium bromide (3d) and n-tetradecylquinuclidinium bromide (3e) were a gift of Professor G. Savelli, University of Perugia, Italy. Polyethylene glycol dodecyl ether (Brij-35) and dodecyl sodium sulfate (SDS) were Fluka products. Di-nhexadecyldimethylammonium bromide20 (4a) and 1,2-bis(palmitoyloxy)-3-(trimethylammonium)propyl bromide21 (5) were prepared as described. The synthesis of (S)-2-((N-n-dodecyl-N(1-methyl-2-hydroxyethyl)amino)methyl)pyridine (2a) and (S)2-((N-n-dodecyl-N-(1-isopropyl-2-hydroxyethyl)amino)methyl)pyridine (2b) has been reported.3 The p-nitrophenyl esters of the two enantiomers of phenylglycine (PhgPNP) were prepared using literature methods.22 (S)-2-((N-Methyl-N-(1-isopropyl-2-hydroxyethyl)amino)methyl)pyridine (2c). To a solution of (S)-2-amino-3-methyl1-butanol (1.08 g, 10.5 mmol) in 50 mL of dry CH2Cl2 containing 2 g of activated molecular sieves, a solution of 2-pyridinecarboxaldehyde (1.13 g, 10.5 mmol) in 80 mL of dry CH2Cl2 was added. The reaction mixture, protected from moisture, was stirred at room temperature for 3 h. The molecular sieves were then filtered off, and the organic solvent was evaporated to give a yellow oil. To this latter, cooled in an ice bath, was added a suspension of NaBH4 (1.6 g, 42 mmol) in 100 mL of absolute ethanol, and the reaction mixture, protected with a CaCl2 tube, was stirred at room temperature overnight. The solvent was then evaporated, and the crude was treated with 50 mL of a 10% Na2CO3 solution and extracted with CHCl3 (3 × 70 mL). The organic phase was dried over Na2SO4 and evaporated to afford 1.79 g (87.7%) of (S)-2-((N-(1-isopropyl-2-hydroxyethyl)amino)methyl)pyridine as a yellow oil: [R]D ) +27.9 (c ) 0.7, CHCl3). 1H-NMR δ (CDCl ): 0.92 (d, J ) 6.8 Hz, 3H, CH(CH ) ), 0.99 (d, 3 3 2 J ) 6.8 Hz, 3H, (CH(CH3)2), 1.83 (m, 1H, CH(CH3)2), 2.48 (m, 1H, NCHCH2OH), 3.44 (dd, J ) 10.9 Hz and 7.17 Hz, 1H, CH2OH), 3.67 (dd, J ) 10.9 Hz and 3.9 Hz, 1H, CH2OH), 3.91 (d, J ) 14.8 Hz, 1H, CH2Py), 4.05 (d, J ) 14.8 Hz, 1H, CH2Py), 7.25 (m, 2H, H3,5 Py), 7.64 (dt, J ) 7.62 Hz and 1.81 Hz, 1H, H4 Py), 8.54 (m, 1H, H6 Py). (19) Holzbecher, Z. Handbook of Organic Reagents in Inorganic Analysis; Wiley: Chichester, 1976. (20) Ueoka, R.; Matsumoto, Y. J. Org. Chem. 1984, 49, 3774. (21) Moss, R. A.; Swarup, S. J. Am. Chem. Soc. 1986, 108, 5341. (22) Schnabel, E. Justus Liebigs Ann. Chem. 1964, 673, 171.
Cleij et al. The above amine (1.43 g, 7.4 mmol) was dissolved in 15 mL of EtOH and poured in a screw-top pressure tube. To this solution were added methyl iodide (0.5 mL, 8.0 mmol) and 6.4 mL of ethyldiisopropylamine. The tube was then sealed and heated at 80 °C for 3 days. A white precipitate was filtered off and the EtOH evaporated. The residue was dissolved in 70 mL of CHCl3 and washed with a 10% Na2CO3 solution (3 × 70 mL). The evaporation of the dried organic solvent (Na2SO4) afforded a crude that was purified by column chromatography (neutral alumina, CH2Cl2) yielding 0.34 g (22%) of pure 2c as a yellowish oil: [R]D ) -4.1 (c ) 0.85, CHCl3). 1H-NMR δ (CDCl3): 0.88 (d, J ) 6.58 Hz, 3H, CH(CH3)2), 1.04 (d, J ) 6.58 Hz, 3H, CH(CH3)2), 1.86 (m, 1H, CH(CH3)2), 2.37 (s, 3H, NCH3), 2.16 (m, 1H, NCHCH2OH), 3.39 (t, J ) 10.6 Hz, 1H, CH2OH), 3.67 (dd, J ) 10.96 and 4.75 Hz, 1H, CH2OH), 3.84 (d, J ) 14.6 Hz, 1H, CH2Py), 4.05 (d, J ) 14.6 Hz, 1H, CH2Py), 7.20 (m, 2H, H3,5 Py), 7.63 (t, J ) 7.68 Hz, 1H, H4 Py), 8.55 (m, 1H, H6 Py). Vesicle Preparation and Characterization. The vesicle solutions were prepared in a 25 mL beaker by dissolving the ligand 2b and the proper nonfunctional surfactant (4-5) in the minimum amount of CH2Cl2. The solvent was allowed to evaporate and then the beaker was kept under vacuum for 2 h. The surfactant film was covered with 20 mL of MES buffer (0.025 M, pH 5.5) and sonicated (Branson Sonifier Model 250, immersion probe, 45% of power output) for 20 min at 50 °C. The vesicle solutions were allowed to cool to room temperature and filtered through 0.45 µm Millipore filters before use. Gel-to-liquid crystal phase transition temperatures, Tc, were determined from fluorescence polarization studies8 using covesicallized 1,6diphenyl-1,3,5-hexatriene (DPH) as a probe under the following conditions: [DPH] ) 5 × 10-6 M, [2b] ) 5 × 10-5 M, [surfactant] ) 5 × 10-4 M. Vesicle sizes were determined by dynamic light scattering using a Nicomp 370 autocorrelator equipped with a Spectra-Physics 2016 argon laser. Kinetic Studies. Slower reactions were followed on a PerkinElmer Lambda 5 spectrophotometer equipped with a thermostated cell holder and faster reactions on an Applied Photophysics SF.17MV stopped flow spectrometer. Solutions of the ligands, metal ions, and additives were prepared in MES buffer (pH 5.5, 0.05 or 0.025 M for micellar and vesicular solutions, respectively). Slower reactions were started by addition of 20 µL of a solution (1-2) × 10-3 M of substrate in CH3CN to 2 mL of solution of ligand and additives. Faster reactions were started by mixing equal volumes of a solution (2-4) × 10-5 M of substrate in water at pH 3.5 (HNO3) for micellar solutions or in MES buffer 0.025 M pH 5.5 for vesicular solutions and of the solution containing ligands, additives, and buffer. In each case, no pH changes were observed during the kinetic runs. The final concentration of substrate was (1-2) × 10-5 M and the release of p-nitrophenol was observed at 317 nm. The kinetics follow in each case a firstorder law up to 90% of reaction. The rate constants were obtained by nonlinear regression analysis of the absorbance vs time data (using the software package Enzfitter: Leatherbarrow, R. J. Enzfitter; Elsevier: Amsterdam, 1987, or the software package SF.17MV provided with the stopped flow work station) and the fit error on the rate constant was always less than 1%.
Acknowledgment. The authors are indebted to Drs. G. Cozzi and C. Barbarotto and Ms. C. Nieuwkerk (ERASMUS exchange student from the University of Groningen, The Netherlands) for their involvement at some stages of this research and to Mr. E. Castiglione for technical assistance. A special thank is due to professor G. Savelli for providing a generous amount of the surfactants with bulky headgroups. Financial support by the Ministry of the University and Scientific and Technological Research (MURST) and by the European Economic Community for Dr. M. C. Cleij’s fellowship under the network Science Programme, Contract SC1*-CT920764, is gratefully acknowledged. LA9515332
