Langmuir 1998, 14, 975-978
975
Kinetic Amplification of the Enantioselective Cleavage of r-Amino Acid Esters by Metallomicelles Federica Bertoncin,† Fabrizio Mancin,† Paolo Scrimin,‡ Paolo Tecilla,*,† and Umberto Tonellato*,† Department of Organic Chemistry and Centro CNR Meccanismi di Reazioni Organiche, University of Padova, Via Marzolo 1, 35131 Padova, Italy, and Department of Chemical Sciences, University of Trieste, Via Giorgieri 1, 34127 Trieste, Italy Received October 10, 1997. In Final Form: December 5, 1997 The enantioselective cleavages of p-nitrophenyl esters of phenylalanine (PhePNP), phenylglycine (PhgPNP), and leucine (LeuPNP) catalyzed by Cu(II) complexes of homochiral ligands have been investigated in conditions of very fast change of pH (“pH-jump”) at the beginning of the reaction. Using lipophilic ligands comicellized with hexadecyltrimethylammonium bromide and decreasing the pH from 9 to 5, a remarkable amplification of the enantioselectivity has been observed when compared to the same reaction performed at constant pH (from 24 to 58 for PhgPNP, as the ratios of the rate constants measured for the faster and slower reacting enantiomers). This has been correlated with the changes in the ligand/Cu(II) complex concentration induced by the change of pH. In fact, it has been established that under the conditions employed the faster enantiomer of the substrate reacts with a higher concentration of catalyst than the slower one: in the first case the reaction occurs before decomplexation takes place and in the second case after decomplexation is virtually complete.
Introduction Surfactant aggregates can influence chemical rates and equilibria.1 For example, in the case of ester cleavage large rate accelerations have been obtained in water using cationic surfactants bearing a nucleophilic function2 or, more recently, metalloaggregates made by lipophilic transition-metal complexes.3 Typically, the aggregate exerts its role on the basis of two main effects: hydrophobic interactions concentrate the reactants in the small aggregate volume and charge interactions shift to a more favorable position the dissociation equilibria of the nucleophile.3g,4 In both cases equilibria are involved and, consequently, the type of control exerted by the aggregate is purely thermodynamic. In some cases such control can be very sophisticated. For example, we have reported5 that a different partition between water and aggregate of the two diastereomeric reacting species (a ternary complex comprising ligand, Cu(II), and substrate) is at the basis of the enantiomeric discrimination (26-fold rate difference) in the cleavage of † ‡
University of Padova. University of Trieste.
(1) (a) Fendler, J. Membrane Mimetic Chemistry; Wiley: New York, 1982. (b) Fendler, J. H.; Fendler, J. E. Catalysis in Micellar and Macromolecular Systems; Academic Press: New York, 1975. (2) Bunton, C. A.; Savelli, G. Adv. Phys. Org. Chem. 1986, 22, 213. (3) (a) Fornasier, R.; Milani, D.; Scrimin, P.; Tonellato, U. Gazz. Chim. Ital. 1986, 116, 55. (b) Gellman, S. H.; Petter, R.; Breslow, R. J. Am. Chem. Soc. 1986, 108, 2388. (c) Menger, F. M.; Gan, L. H.; Johnson, E.; Durst, D. H. J. Am. Chem. Soc. 1987, 109, 2800. (d) Weijnen, J. G. J.; Koudijs, A.; Engbersen, J. F. J. J. Org. Chem. 1992, 57, 7258. (e) Ogino, K.; Kashihara, N.; Ueda, T.; Isaka, T.; Yoshida, T.; Tagaki, W. Bull. Chem. Soc. Jpn. 1992, 65, 373. (f) Scrimin, P.; Tecilla, P.; Tonellato, U. J. Org. Chem. 1994, 59, 18. (g) Bunton, C. A.; Scrimin, P.; Tecilla, P. J. Chem. Soc., Perkin Trans. 2 1996, 419. (h) Kriste, A. G.; Vizitiu, D.; Thatcher, R. J. Chem. Commun. 1996, 913. (4) (a) Fornasier, R.; Tonellato, U. J. Chem. Soc., Faraday Trans. 1980, 76, 1301. (b) Bunton, C. A.; Nome, F.; Quina, F. H.; Romsted, L. Acc. Chem. Res. 1991, 24, 357. (c) Menger, F. M. Angew. Chem., Int. Ed. Engl. 1991, 30, 1086. (5) (a) Cleij, M. C.; Tecilla, P.; Tonellato, U.; Scrimin, P. Gazz. Chim. Ital. 1996, 126, 827. (b) Cleij, M. C.; Tecilla, P.; Tonellato, U.; Scrimin, P. Langmuir 1996, 12, 2956. (c) Scrimin, P.; Tecilla, P.; Tonellato, U. J. Org. Chem. 1994, 59, 4194.
the enantiomers of an R-amino acid ester. In the absence of the aggregate, no enantioselection is observed. On the other hand, it is known that the association of lipophilic metal complexes with cationic aggregates may result not only in a smaller formation constant of the complex but also in a slower complexation and decomplexation of the metal ion.6 In some cases, depending on the complex and on the operative conditions, the rate of formation and disruption of the complexes can be so slow that it becomes comparable with the rate of cleavage of a given substrate. Furthermore, with ligands involved in acid-base equilibria, the pH plays a key role in determining the strength of the complex: at low pH the competition with protons may become so unfavorable that no complex is formed.7 It occurred to us that, by decreasing abruptly the pH of the solution from a high value, where the complex is totally formed, to a lower value, where the complex is only partially formed, we could realize a system in which the rate of the decomplexation process lies between the rate of cleavage of two enantiomers, leading to amplification of enantioselectivity. In such a system one enantiomer reacts so fast that the effective catalyst (the metal ion/ ligand complex) has no time to decomplex while the other enantiomer reacts only after the decomplexation has taken place. We are pleased to report here the first successful example of a kinetically controlled amplification of enantioselectivity by metallomicellar aggregates. The reaction here investigated is the cleavage of the p-nitrophenyl esters of phenylglycine (PhgPNP), phenylalanine (PhePNP), and leucine (LeuPNP) by the Cu(II) complexes of the ligands shown in Chart 1. While ligands 1a and 2a, lacking the long paraffinic chain, are water soluble, the more lipophilic 1b and 2b are not soluble in water even as Cu(II) complexes and were studied as comicelles with hexadecyltrimethylammonium bromide (CTABr) in a 1:10 ratio. Previous work8 has shown that (6) Cierpiszewski, R.; Hebrant, M.; Szymanowski, J.; Tondre, C. J. Chem. Soc., Faraday Trans. 1996, 249 and references therein. (7) Martell, A. E.; Motekaitis, R. J. Determination and Use of Stability Constants; VCH Publishers: New York, 1992.
S0743-7463(97)01112-8 CCC: $15.00 © 1998 American Chemical Society Published on Web 02/04/1998
976 Langmuir, Vol. 14, No. 5, 1998 Chart 1
the Cu(II) complexes of 1b and 2b, comicellized with CTABr, accelerate the rate of cleavage of PhePNP and PhgPNP and the process is remarkably enantioselective. Ligand 2b was found to be the most enantioselective catalyst: in the case of the two enantiomers of PhgPNP and PhePNP, the observed enantioselectivity ratios (ER, the ratio between the pseudo-first-order rate constants measured for the cleavage of the two enantiomers of the substrate) were 24 and 14, respectively. In the present study we have investigated the reactivity of the ligands under the condition of very fast change of the pH (9 f 5 and 9 f 7.5) of the reacting mixture at the beginning of the kinetic run. We have also investigated ligands 1a and 2a to compare the aggregate system with the monomeric one. Experimental Section General Methods and Materials. Melting points are uncorrected. 1H NMR spectra were recorded on a Bruker AC 250F spectrometer operating at 250 MHz, and chemical shifts are reported relative to internal Me4Si. Elemental analyses were performed by the Laboratorio di Microanalisi of our Department. Cu(NO3)2 was an analytical-grade product, and metal ion stock solutions were titrated against EDTA following standard procedures.9 The buffers10 2-morpholinoethanesulfonic acid (MES; Fluka) and 4-(2-hydroxyethyl)-1-piperazinethanesulfonic acid (HEPES; Sigma) were used as received. n-Hexadecyltrimethylammonium bromide (CTABr) was an analytical-grade commercial product. The p-nitrophenyl esters of the R-amino acids used as substrates were prepared according to literature methods.11 The syntheses of (S)-1,2-diamino-[N-tetradecyl-N ′-(1-benzyl-2-hydroxyethyl)]ethane (1b) and (S)-1,2-diamino-[N-tetradecyl-N ′((S)-1-benzyl-2-hydroxyethyl)]-1-methylethane (2b) have been reported.8 The ligands (S)-1,2-diamino-[N-methyl-N ′-(1-benzyl2-hydroxyethyl)]ethane (1a) and (S)-1,2-diamino-[N-ethyl-N ′((S)-1-benzyl-2-hydroxyethyl)]-1-methylethane (2a) were synthesized using the same procedure as described for lipophilic analogues 1b and 2b. In the case of 1a, the starting material was BOC- protected N-methylglycine.12 (S)-1,2-Diamino-[N-methyl-N ′-(1-benzyl-2-hydroxyethyl)]ethane‚2HCl (1a‚2HCl). Mp: 167-168 °C. [R]25D ) - 8.2 (c ) 1.0; MeOH). 1H NMR (CD3OD): 2.84 (s, 3 H, NCH3) 3.1 (m, 2 H, CH2C6H5); 3.45 (m, 6 H, NCH2CH2N and CH2OH); 3.81 (m, 1 H, CH); 7.37 (m, 5 H, C6H5). Anal. Calcd for C12H22N2OCl2: C, 51.25; H, 7.89; N, 9.96. Found: C, 50.97; H, 7.81; N, 9.85. (S)-1,2-Diamino-[N-ethyl-N ′-((S)-1-benzyl-2-hydroxyethyl)]-1-methylethane (2a). The dihydrochloride salt of this compound was an oil. The free diamine was purified by column chromatography (silica; CHCl3/MeOH, 20:1), giving a clear oil. [R]20D ) +1.2 (c ) 2; CHCl3).1H NMR (CDCl3): 1.07 (d, J ) 6.8 Hz, 3 H, CHCH3); 1.15 (t, J ) 7.2 Hz, 3 H, CH2CH3); 2.55 (m, 3 H, CH3CH2N and CH(CH3)); 2.85 (m, 4 H, CH(CH3)CH2N and CH2C6H5); 2.95 (m, 1 H, NCH(CH2C6H5)CH2OH); 3.55 (AB system, 2 H, CH2OH); 7.25 (m, 5 H, C6H5). Anal. Calcd for C14H24N2O: C, 71.14; H, 10.23; N,11.85. Found: C, 70.86; H, 10.19; N, 11.79. (8) Cleij, M. C.; Mancin, F.; Scrimin, P.; Tecilla, P.; Tonellato, U. Tetrahedron 1996, 53, 357. (9) Holzbecher, Z. Handbook of Organic Reagents in Inorganic Analysis; Wiley: Chichester, U. K., 1976. (10) Good, N. E.; Winget, G. D.; Winter, W.; Connolly, T. N.; Izawa, S.; Singh, R. M. M. Biochemistry 1966, 5, 467. (11) Schnabel, E. Ann. 1964, 673, 171. (12) Perseo, G.; Piani, S.; De Castiglione, R. Int. J. Peptide Protein Res. 1983, 21, 227.
Letters Table 1. Apparent Formation Constants (Kf, M-1), Molar Extinction Coefficients (E, M-1 cm-1), and Rate Constants of Decomplexation (k-1, s-1) for the Different Ligand/ Cu(II) Complexes at pH ) 5a entry
ligand
log Kfb (log Kf, pH ) 9)c
(λ) [M-1 cm-1 (nm)]
k-1d (s-1)
1 2 3 4 5
1a 1be 2a 2be 2be
4.7 (10.3) 4.2 (9.8) 4.0 (9.6) 3.7 (9.3) 3.7 (9.3)
1980 (280) 2370 (280) 1460 (280) 3086 (280) 1305 (317)
3.05 1.26 6.40 1.10 1.10
a MES buffer, 0.05 M, T ) 25 °C. b The error on formation constants is (0.1 logarithmic units. c See ref 14. d Measured from pH-jump experiments, see text for details. e In the presence of 10fold excess CTABr.
Binding Constants Determination. The apparent formation constants for the ligand/Cu(II) complexes at pH 5 were determined by UV titration using a Perkin-Elmer Lamda 16 spectrophotometer equipped with a thermostated cell holder. A solution (2 mL) of ligand (0.5 × 10-4-1.0 × 10-4 M) and CTABr in the case of 1b and 2b (0.5 × 10-3-1.0 × 10-3 M) in 0.05 M MES at pH 5 was placed in the measurement cuvette and balanced with a reference cuvette containing the same volume of buffer and CTABr solution. To the two cuvettes, thermostated at 25 ( 0.1 °C, were added equal volumes of (1) a solution containing ligand, CTABr, and MES at the same concentrations as above and Cu(II) (1.0 × 10-3-4.77 × 10-3 M) to the measurements cuvette and (2) a solution containing the same amounts of CTABr, MES, and Cu(II), but without ligand, to the reference cuvette. After each addition the absorbance at 260, 280, and 317 nm was recorded, and the data were fitted using the HOSTEST 5 program.13 “pH-Jump” Kinetics. The correct amount of ligand to obtain a concentration of 2.0 × 10-4 M was dissolved in water containing in the case of 1b and 2b CTABr (2.0 × 10-3 M). To this solution was added an aliquot of 5.0 × 10-2 M Cu(NO3)2 to give a 2.0 × 10-4 M concentration of Cu(II), and the pH value was adjusted to 9 with 0.1 M NaOH (solution 1). The buffer solution (0.1 M) was prepared using MES or HEPES at pH 5 or 7.5, respectively (solution 2). In the case of ester cleavage, the substrate (2 × 10-5 M) was added to solution 2 just before the kinetic run. The kinetic measurements were performed with an Applied Photophysics SF.17MV stopped-flow spectrophotometer. In the stopped-flow apparatus equal amounts (0.1 mL) of solutions 1 and 2 were mixed. The decomplexation of ligand/Cu(II) species was monitored as the decrease of absorption at 280 and 317 nm, and the release of 4-nitrophenol was followed by the increase of absorption at 317 (pH ) 5) and 400 (pH ) 7.5) nm. The absorbance vs time data were treated as described in the text, and the rate constants were calculated as the average value of 6 repeated runs using the software package provided with the stopped-flow workstation.
Results and Discussion Apparent formation constants (Kf) of the different ligand/Cu(II) complexes measured at pH 5 are reported in Table 1 along with those estimated14 at pH 9. The data were obtained by UV titration of a ligand (and CTABr 1:10 for 1b and 2b) solution in 0.05 M morpholinoethanesulfonic acid (MES) buffer with a Cu(II) solution in the same buffer. The absorbance vs concentration data were nicely fitted for a 1:1 stoichiometry of the complex using the HOSTEST 5 program.13 From the data of Table 1 it is possible to evaluate the concentration of the complexes at different pH values. For example, for a solution containing 2b (1 × 10-4 M), CTABr (1 × 10-3 M), and Cu(II) (1 × 10-4 M), the concentration (10-5 M) of the (13) Wilcox, C. S. In Frontiers in Supramolecular Organic Chemistry and Photochemistry; Schneider, H.-J., Du¨rr, H., Eds.; VCH Publishers: New York, 1991. (14) Estimated using the pKa values reported for the amino groups of N-(2-hydroxyethyl)ethylendiamine: Smith, R. M.; Martell, A. E. Critical Stability Constants; Plenum Press: New York, 1975; Vol. 2.
Letters
Langmuir, Vol. 14, No. 5, 1998 977 Table 2. Pseudo-First-Order Rate Constants, kR and kS (s-1), and the Enantioselectivity Ratio, ER ) kS/kR, for the Cleavage of PhgPNP in the Presence of Different Ligand/Cu(II) Complexes at pH 9 f 5 and 9 f 7.5a 9f5 entry ligandc 1 2 3 4 5 6
1a 1bd 2a 2bd 2bd,e 2bd,f
7.5b
9 f 7.5
kS
kR
ER
kS
0.047 9.3 0.024 23.9 26.9 2.86
0.020 0.40 0.017 0.41 0.84 0.19
2.3 23.2 1.4 58.3 32.0 15.0
40.81 169.6 27.54 340.4 479.9 242.9
kR
ER
ER
11.64 3.5 18.38 9.2 6.43 4.3 12.3 27.7 24.1 29.1 16.5 14.2 22.9 10.6
a Data obtained from “pH-jump” experiments; see text for details. Data from ref 8. c [Ligand] ) [Cu(II)] ) 1 × 10-4 M. d In the presence of 10-fold excess CTABr. e Using PhePNP as substrate. f Using LeuPNP as substrate. b
Figure 1. Change of absorbance vs time profiles for the cleavage of S-PhgPNP (A) and R-PhgPNP (B) in the presence of 1 × 10-4 M 2b, 1 × 10-4 M Cu(II), and 1 × 10-3 M CTABr in MES buffer (0.05 M, pH 5). The profiles have been obtained by mixing an unbuffered solution of the copper complex at pH 9 with a buffered solution (pH ) 5) containing (curve a) or not containing (curve b) the substrate. Curve c is the difference between curves a and b.
complex 2b/Cu(II) on moving from pH 9 to 7.5 to 5 decreases from 10 to 9.9 to 2.7, respectively. The rate constants of the decomplexation process (k-1) are also reported in Table 1. These were measured with the following procedure here referred to as “pH-jump”: an unbuffered solution at pH 9 of ligand, CTABr when necessary, and Cu(II) of concentrations 2 × 10-4, 2 × 10-3, and 2 × 10-4 M, respectively, was mixed with an equal volume of a pH 5 solution of 0.1 M MES buffer, with the aid of a stopped-flow apparatus. During the mixing time (
