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The zwitterion structure of imidazol-1-ylacetic acids in the solid state and in solution
Journal of
MOLECULAR STRUCTURE
ELSEVIER
Journal
of Molecular
Structure
377 (1996) 105-l 12
The zwitterion structure of imidazol- 1-ylacetic acids in the solid state and in solution Pilar
L6peza, Paula Zaderenko”, Jose Luis Balcazarb, Isabel Fonsecac, F&lix Hernhdez Cane’, Paloma Ballesterosa,*
‘Departamenio de Quimica Orgcinica y Biologia, Facultad de Ciencias, UNED, Senda de1 Rey s/n, 28040-Madrid, Spain bDepartamento de Ciencias Analiiicas, Facultad de Ciencias, UNED, Senda del Rey s/n, 28040-Madrid, Spain ‘Departamento de Cristalografin. Insrituto de Quimica Fisica ‘Rocasolano’, CSIC, Serrano 119, 28OO&Madrid, Spain
Received 7 June 1995; accepted in final form 20 September 1995
Abstract The crystal zwitterion structures of imidazol-1-ylacetic acid 1 and (*)-3-ethoxycarbonyl-2-imidazol-1-ylpropionic acid 2 have been determined by X-ray diffraction analysis. Spectroscopic 13C NMR studies in the solid state, by the CP/MAS technique, and in solution have revealed the presence of the zwitterion structure in Hz0 and D20 solutions. Results have been complemented with “0 NMR and IR data. Keywords: X-ray crystallography;
Solid state NMR
spectroscopy;
1. Introduction
Zwitterion
acid
0
1996 Elsevier
Science
B.V. All rights
COpEt
I
C02H
C02Me
HOzC 2
1
3
N
I \ C>N < coat-Bu
author.
SSDZ 0022-2860(95)09118-l
l-5
I ‘: C>N
4 0022-2860/96/$15.00
Imidazol-I-ylacetic
by ‘H NMR [4]. Among them, compounds and the related 6 were prepared
The analgesic and hypnotic activity of imidazol1-ylacetic acid 1 was formerly related to the presence of an intramolecular hydrogen bond between the carboxylic group and the nitrogen atom at position 3 of the imidazole ring [l]. Later and considering the ratio of ionic molecules to neutral ones obtained from Ebert’s equation, a zwitterion structure was proposed [2]. Also pioneer ‘H NMR work suggested a similar zwitterion structure in D20 solution [3]. We have described recently a series of imidazol1-ylalkanoic acids as new probes to determine the intracellular and extracellular pH and cell volume
* Corresponding
structure;
reserved
COpH COzH
HO& 5
6
106 Table 1 Crystal data, data collectiona
P. Ldpez et al./Journal of Molecular Structure 377 (1996) 105-112
and structure
refinementb
of compounds
1 and 2 Compound
Compound
Formula Crystal size (mm) Symmetry Unit cell determination
CsH6NzG2 0.38 x 0.32 x 0.21 Monoclinic, P2, /n Least-squares fit from 58 reflexions (10 < 20 < 90”) 14.328(l), 8580(l), 4.618(l) A 90.0”, 98.777(5)“, 90.0” 561.1(l), 4 1.4930, 126.115, 264.0 9.575
CsH,>NzQ 0.38 x 0.32 x 0.21 Monoclinic, C2/c Least-squares fit from 64 reflexions (12 < 20 < 90”) 26.927(2), 5.426(l), 14.555(l) .& 90.0”, 104.043(l)“, 90.0” 2063.0(4), 8 1.3664, 212.205, 896 8.801
958 862 (I > 2u (I) criterion) -16/16 O/l1 O/6 Fourier syntheses, Coord. parameters refined 151 0.002 0.041, 0.049 0.18eA-3
1738 1699 (I > 2~ (I) criterion) -31/31 o/7 o/17 Fourier synthesis, Coord. and thermal parameters refined 256 0.07 0.047, 0.058 0.24 e Am3
Unit cell dimensions Packing: V (A3), Z D, (gcme3), M, F (000) p (cm-’ ) Number of reflexions: Measured Observed Range of hkl H atoms Parameters (A/o) max Final R and R, Ap max
1
2
Parameter
and thermal
a Technique: Four circle diffractometer, Philips PW 1100, Bisecting geometry, Graphite oriented monochromator, Cu Ka 1.5418 A, correction, None. Two standard reflexions, frequency, 90 min. w/20 scan. Scanning range for 19: 2 < 0 < 65. Absorption b Solution: Direct methods. Refinements: L.S. on Fobs. w-Scheme: Empirical as to give no trends in < wA2F > vs. < ]Fs] > and < sin Q/x >. Computer and programs: Vax 1 l/750, MLJLTANBO [8], XRAYBO [9], PESOS [lo], PARST [1 11. Scattering factors and anomalous dispersion: Int. Tables for X-Ray Crystallog [12].
The knowledge of the precise ionization state of these new probes in solution is essential to the determination of their membrane permeability properties and to the interpretation of their biological behaviour. In this report we have established unambiguously the zwitterion structures of 1 and 2 in the solid state by X-ray diffraction analysis and the relation with those observed in solution by combined studies of t3C NMR in solid state and in solution.
2. Experimental Imidazol- 1-ylacetic acid (1) [4], (&)-3-ethoxycarbonyl-2-imidazol-1-ylpropionic acid (2) [4], methyl imidazol-1-ylacetate (3) [4], tert-butyl imidazol- I-ylacetate (4) [5], (f)-2-imidazol- l-ylsuccinic acid (5) [6], pyrazol-1-ylacetic acid (6) 173,were prepared according to literature procedures.
2.1. Crystal structure determination
Crystals of compounds 1 and 2 were obtained from aqueous ethanol and absolute ethanol respectively. Crystal and experimental data are given in Tables 1 and 2. The data were collected on a Philips PW 1100 diffractometer, with graphitemonochromated radiation, bisecting geometry, and 1.5” scan width in the w/28 mode. Two reflections were monitored every 90 min, showing no significant variation in the crystals nor in the experimental conditions. The structures were solved by direct methods and refined by leastsquares based on Fobs only. All hydrogen atoms were unambiguously obtained in a difference synthesis and included isotropically in the final cycles of refinement. Weights were chosen to give no trends in < wA2/F > versus < /Fobs1> and < sin 0/X >, by functions of the w = K/[f(F&]-[g(sin e/x)] type, K being a scale factor to ensure that < wA2F > N 1. Final fractional
P. Ldpez et al./Journal of Molecular Structure 377 (1996) 105-112
107
08
(4
(b) Fig. 1. (a) A ORTEP [13] view of compound 1 showing the atomic numbering; displacement ellipsoids are drawn at the 50% probability level; (b) packing of the molecules of compound 1 viewed down the c axis.
atomic coordinates, the full list of bond lengths and angles, and the list of thermal parameters of compounds 1 and 2 are deposited with the B.L.L.D. as Supplementary Publication No. SUP2656 1 (48 pages). 2.2. Infrared measurements IR spectra were recorded in KBr on a BomemDA3 FT-IR spectrophotometer. 2.3. NMR determinations 13C NMR
spectra in solution were recorded
at 50.33 MHz using a Bruker AC200 spectrometer at 25°C. Samples were dissolved in D20, H20 or DMSO-d6 and the concentrations were 0.5 to 1 M. Chemical shifts are expressed in p.p.m. from tetramethylsilane using DMSO-d6 as external or internal reference. Typical conditions were as follows: pulse width, 3 ps (ca. 45”); relaxation delay, 1 s, spectral width, 10 KHz; data points, 32768. 170 NMR spectra were taken at 42.82 MHz on a Bruker AM-360. The enriched compounds 1, 2 and 5 were obtained by partial and total hydrolysis of the corresponding esters with enriched “0 (28%) “0 (32%) water, and spectra conditions were the same as those
108
P. Ldpez et al./Journal of Molecular Structure 377 (1996) 105-112 08
(4
Fig. 2. (a) A ORTEP [13] view of compound 2 showing the atomic numbering; Displacement level; (b) packing of the molecules of compound 2 viewed down the b axis.
ellipsoids
are drawn at the 50% probability
P. Ldpez et al./Journal of Molecular Structure 377 (1996) 105-112 Table 2 Geometric
parameters
Bond lengths
Bond Compound NlLC2 Nl-C5 NlLC6 C2-N3 N3-C4 c44c5 C6-C7 C7p08 c7-09
(A, deg) for compounds (A)
109
1 and 2 Angle
(deg)
1.324(2) 1.379(3) 1.459(2) 1.325(3) 1.373(3) 1.351(3) 1.529(3) 1.259(2) 1.231(2)
C5-Nl-C6 C2-Nl -C6 C2-Nl C5 Nl-C2-N3 C2-N3-C4 N3-C4-C5 N 1-C5-C4 Nl-C66C7 C6-C7-09 C6-C7-08 08-C7-09
125.6(2) 125.4(2) 108.9(l) 108.4(2) 109.0(2) 107.0(2) 106.7(2) 113.4(l) 119.6(l) 113.9(l) 126.4(2)
1.335(2) 1.378(2) 1.468(2) 1.314(2)
C5-NlLC6 C2-Nl C6 C2-Nl-C5 Nl-C2-N3 C2pN3-C4 N3-C4-C5 Nl-C5-C4 Nl-C66ClO Nl-C6-C7 C7-C66ClO C6-C7-09 C6-C7708 088C7-09 C6-CIO-Cl1 ClO-Cll-013 ClO-Cl I-012 012-CllLO13 Cll-013-Cl4 013-c14-Cl5
124.2(l) 128.3(l) 107.5(l) 109.7(l) 108.2(l) 107.6(l) 107.0(l) 111.6(l) 113.1(l) 113.3(l) 111.6(l) 120.5(l) 127.8(l) 114.0(l) 110.4(l) 125.8(2) 123.8(2) 116.4(2) 106.2(3)
H...A 1.66(2) 1.57(3)
D...A 2.645(2) 2.550(2)
1
Compound 2 NlLC2 NlLC5 Nl-C6 C2-N3 N3-C4 c44c5 C66C7 C6-Cl0 C7-08 c7709 cto-Cl1 C11&012 Cl l-013 013-Cl4 c14pc15
Hydrogen
bonds
Compound Compound
1 2
1.370(2) 1.350(2) I .544(2) 1.525(2) 1.229(2) 1.257(2) 1.502(2) 1.202(2) 1.332(2) 1.456(3) 1.496(5)
D-H...A N3-H3 N3-H3
Neighbouring chain interactions Compound 2 012,..H142 2.69(3) 012...H151 2.82(6) 3.01(4) Ol3...H142 3.04(7) O13...H151 2.69(4) O13...Hl52 a +x - l/2, -y + 112 + 1, +z - l/2. b +x, -y, +z - l/2. Symmetry code.
08a 09b
(-x + 1, -y, --z + 1) (-x + 1, -y + 1, -z + 1) (-x+1,-y+l,-z+l) (-x+1,-y+l,-z+l) (-x + 1, y. -z + l/2 + 1)
D-H...A 172(2) 169(2)
P. Lbpez et al./Journal of Molecular Structure 377 (1996) 105-112
110 Table 3 13C NMR chemical
shifts (6, ppm)a of compounds
1-6 at 50.33 MHz
Compound
pD or pH
13CNMR
c2
c4
G
1
5.60 4.70
D2O
134.4
H20
135.2
121.9 122.6
118.5 119.1
CP/MAS DzO
135.2 134.3 133.9
123.5 120.6 120.1
119.9 118.6 118.2
136.4
D2O
137.8 138.4 138.1 138.5 134.3
121.4 126.7 130.0 120.9 127.6 121.6
121.4 120.3 121.0 120.5 121.9 118.4
CP/MAS
133.7
122.9
116.2
169.8
105.0 106.5
131.0 129.5
174.5 174.4
2
3.57 3.33
H20
CP/MAS 3
D2O
4
CP/MAS DMSO-d; CP/MAS
5
3.29
6
139.9 140.7
D20
CP/MAS a Imidazole ring assignments b Very insoluble in D20. ’ Performed in H:‘O.
have been made considering
the data described
C02H
C02R
171.5 172.1 170:271.1’ 171.4 171.5 170.8 170:257.0’ 172.9
in the literature
171.8
CH
CH2
50.8 51.4
171.0 170.4 171.1 170.0 172.0 167.5 169.1 173.2 “0: 267.1’ (R=H) 171.8 (R=H)
59.5 58.9 59.7
51.2 36.4 35.9
59.6
38.8 46.8 47.7 47.7 48.0 36.6
60.0
37.9 53.6 56.2
[21].
previously reported [5]. 13C Cross-polarization magic angle spinning (CP/MAS) NMR spectra
3. Results and discussion
were obtained at 50.33 MHz
Structural study by X-ray diffraction analysis of compounds 1 and 2 (Table 1) revealed that both acids existed in zwitterion form in the solid state (Figs. 1 and 2). The proton from carboxylic oxygen migrates to the N3 in the imidazole ring forming the zwitterion. The charge was delocalized along Nl-C2-N3, and the imidazole ring was planar with maxima deviations of this plane of -0.0008(20) (C5 compound 1) and -0.0016(19) (C4 compound 2). Bond distances and angles (Table 2) were very similar in the two compounds and were on the same range of values as those of the imidazolium cation in imidazolium sulphate dihydrate [14]. Both zwitterion structures were also stabilized by strong hydrogen bonds (distances N3 . . ..O = 2.645(2) A for compound 1 and 2.549(2) A for compound 2) (Table 2). In this situation, both compounds formed infinite chains along the a axis (see Figs. 1 and 2) similar to those observed in some salts of hydroxybenzoic acids
spectrometer.
Samples
2.5 kHz in a 7 mm time was
were
on a Bruker CP-200 rotated
at
3.5 or
Zr02 rotor; the initial contact
1 ms; 90” pulse lengths
for
protons
were 7 ps; 4 s delay between scans and 200 transients were used. Chemical shifts were measured using glycine
(176.1 ppm)
as external
reference. 2.4. Determination
of pKH1 of compounds
1 and 2
The pK, of the carboxylic group of compounds 1 and 2 in Hz0 (pKH’) were determined by 13C NMR (50.33 MHz) using DMSO as external reference. pH titrations (25°C) were performed using 0.8 M solutions of the appropriate compound in H20 adjusting the pH with the addition of HCl. The dependence of chemical shifts of carbons N-CH2 of compound 1 and N-CH of compound 2 vs pH were obtained to calculated the pK,.
P. Ldpez et al./Journal of Molecular Structure 377 (1996) 105-112
[15]. Besides, in compound 2 there are some interactions of note between neighbouring chains (Table 2). Moreover, compound 2 was further linked into antiparallel pairs through ring inte:actions (distance between centroids of 3.52 A) [16], and by a short contact between the centroid of the ring and Cl5 (3.686 A), and H152 (3.38 A). Infrared data in the solid state confirmed the zwitterion structure of compounds 1 and 2. Protonation of N3 was observed by the two broad stretching absorptions in the regions at ca 2500 and ca 1900 cm-‘. These bands corresponded to +NH stretching linked by strong hydrogen bonding as it has been found previously for similar compounds [17-191. As it has been stated, NMR solid-state chemical shifts generally are very similar to those found in solution. Thus, the CP/MAS data can be used as a ‘bridge’ between solid-state structures determined by diffraction techniques and those which exist in solution [20]. 13C NMR chemical shifts values of compounds l-6 in the solid state and in solution are depicted in Table 3. Although it has been reported previously [3], that compound 1 presented a zwitterion structure in D,O, it must be taken into account that the pH of the solution is decisive in the formation of the zwitterion structure, as is observed in amino acids. We have recently reported that compound 1 presented a pKD2 (N+) value of 7.23 [4,5]. Considering the pKD (pK, in D20) and pKH (pK, in H20) relationship [22] pKD = 1.018 pKH + 0.43; pKH2 of compound 1 is 6.68. On the other hand, the pKH’ (COOH) calculated here is 2.1. If we consider compound 1 as an amino acid, these values corresponded to an isoelectric point (Ip) of 4.4 The value of the pH (4.70) obtained when compound 1 was dissolved in Hz0 suggested that the zwitterion form is a maximum in Hz0 solution. This fact is confirmed because of the values of the chemical shifts of all carbons fitted well with those found in solid state (Table 3). Furthermore, the 170 chemical shift value (27 1.1 ppm) of the carboxylic group, is also in agreement with those found in other amino acids in zwitterionic form [23]. The described pKD2 value of compound 2 was 6.86 [4] which corresponds with a pKH2 of 6.32. The carboxylic group has a pKH’ of 1.3.
111
These values give an Ip of 3.8. This value suggest that at 3.30, pH of the solution of compound 2 in Hz0 the contribution of the zwitterion form is slightly lower than that observed in compound 1. This fact is also corroborated by the data obtained in the solid state and the I70 NMR chemical shift value (257.0 ppm) which corresponds well with those values reported for other amino acids with protonated carboxyl O-atoms [23]. In the absence of X-ray data it could be inferred that IR data of diacid 5 in KBr suggested that the (r-COOH (1597 cm-‘) is ionized in the solid state but not the /3-COOH, as has been described for other dicarboxylic mono-amino acids [24]. The 13C NMR data in the solid state and in solution also suggested a similar behaviour to compound 2 in solution. Finally, we can conclude that this work unambiguously proves that imidazol- 1-ylacetic acids behave as amino acids and present zwitterion structures in the solid state and in solution within a certain pH range. In the solid state the packing of ions is governed by strong hydrogen bonds which play a significant role in the structural organization. The presence of rigid hydrogenbonded aggregate structures in these acids makes them especially attractive from the crystal engineering point of view [25].
Acknowledgements This work was supported in part by D.G.I.C.Y.T. (PM-92-01 1, PB-93-0037, and PB-93-0125) and Community of Madrid (AE00219/94). P.L. and P.Z. received F.P.U. fellowships from the Spanish Ministry of Education and Science. We are indebted to Dr C. L6pez for her valuable technical assistance in the performance of CP/MAS spectra.
References [l] E.
Roberts and D.G. Simonsen, Biochem. Pharmacol., I5 (1966) 1875. [2] V. Sunjic, F. Kajfez and P. Mildner, Croat. Chem. Acta, 41 (1969) 107.
112
P. Ldprz et al./Journul
qf Moleculur Structure 377 (1996) 105-112
[3] E.O. Bishop and R.E. Richards, Biochem. .I.. 86 (1963) 277. [4] (a) MS. Gil, F. Cruz, S. Cerdln and P. Ballesteros, Bioorg. Med. Chem. Lett., 2 (1992) 1717. (b) M.S. Gil, P. Zaderenko, F. Cruz, S. Cerdin and P. Ballesteros, Bioorg. Med. Chem., 2 (1994) 305. [5] P. Zaderenko, M.S. Gil, P. Ballesteros and S. Cerdan, J. Org. Chem., 59 (1994) 6268. [6] A. Mroczkiewicz, Acta Pol. Pharm., 41 (1984) 425. [7] R.G. Jones, M.J. Mann and K.C. McLaughlin, J. Org. Chem., 19 (1954) 1428. [8] P. Main, S.J. Fiske, S.E. Hull, L. Lessinger. G. Germain, J.P. Declercq and M.M. Woolfson, MULTANXO. A System of Computer Programs for the Automatic Solution of Crystal Structures from X-Ray Diffraction Data. Univs. of York, England and Lovain, Belgium, 1980. [9] J.M. Stewart, F.A. Kundell and J.C. Baldwin, The xRAYx0 System. Computer Science Center. University of Maryland, College Park, Maryland, USA, 1980. [IO] M. Martinez-Rip011 and F.H. Cano, PESOS. A Computer Program for the Automatic Treatment of Weighting Schemes. Instituto Rocasolano. CSIC. Madrid. Spain, 1971. [l I] M. Nardelh, PARST. Comput. Chem., 7 (1983) 95. [12] International Tables for X-ray crystallography. Vol. 4., Birmingham, Kynoch, 1974. [13] S.R. Hall and J.M. Stewart (Eds), Xtal 3.0 Reference Manual. Univs. of Western Australia, Australia and Maryland, USA, 1990. [14] H.C. Freeman, F. Huq, J.M. Rosalky and I.F. Taylor. Acta Cryst., B31 (1975) 2833.
[15] C.B. Aakeriiy, G.S. Bahra, P.B. Hitchcock, Y. Pate11 and K.R. Seddon, J. Chem. Sot., Chem. Commun. (1993) 152. [16] A. Albert and F.H. Cano, CONTACTOS. A Program for Systematic Study of Aromatic Rings Interactions. Instituto Rocasolano. CSIC, Madrid. Personal communication, 1990. [I71 M.F. Claydon and N. Sheppard, Chem. Comm. (1969) 1431. [18] M. Cordes and J.L. Walter, Spectrochimica Acta, 24A (1968) 237. [19] (a) J. Bellanato. C. Avendaiio, P. Ballesteros and M. Martinez, Spectrochimica Acta, 35A (1979) 807. (b) J. Bellanato, C. Avendaiio, P. Ballesteros, E. de la Cuesta, E. Santos and G.G. Trigo, Spectrochimica Acta, 37A (1981) 965. [20] C.A. Fyfe, Solid State NMR for Chemists, C.F.C. Press, Ontario, Canada, 1983. [21] (a) M.R. Grimmett, Imidazoles and their Benzo Derivatives. In Comprehensive Heterocyclic Chemistry, Vol. 5, Pergamon, Oxford, 1984, p. 354. (b) M. Begtrup, J. Elguero, R. Faure, P. Camps, C. Estopi, D. Ilavsky, A. Fruchier, C. Marzin and J. Mendoza, Magn. Reson. Chem., 26 (1988) 134. [22] H.J.C. Yeh, K.L. Kirk, L.A. Cohen and J.S. Cohen, J. Chem. Sot. Perkin Trans 2 (1975) 928. [23] I.P. Gerothanassis, R. Huston and J. Lauterwein, Helv. Chim. Acta, 65 (1982) 1764. [24] L.J. Bellamy, The Infrared Spectra of Complex Molecules, Vol. I. Chapman and Hall, New York, 1975, p. 263. [25] C.B. Aakeroy and K.R. Seddon, Chem. Sot. Rev., 22 ( 1993) 397.
MOLECULAR STRUCTURE
ELSEVIER
Journal
of Molecular
Structure
377 (1996) 105-l 12
The zwitterion structure of imidazol- 1-ylacetic acids in the solid state and in solution Pilar
L6peza, Paula Zaderenko”, Jose Luis Balcazarb, Isabel Fonsecac, F&lix Hernhdez Cane’, Paloma Ballesterosa,*
‘Departamenio de Quimica Orgcinica y Biologia, Facultad de Ciencias, UNED, Senda de1 Rey s/n, 28040-Madrid, Spain bDepartamento de Ciencias Analiiicas, Facultad de Ciencias, UNED, Senda del Rey s/n, 28040-Madrid, Spain ‘Departamento de Cristalografin. Insrituto de Quimica Fisica ‘Rocasolano’, CSIC, Serrano 119, 28OO&Madrid, Spain
Received 7 June 1995; accepted in final form 20 September 1995
Abstract The crystal zwitterion structures of imidazol-1-ylacetic acid 1 and (*)-3-ethoxycarbonyl-2-imidazol-1-ylpropionic acid 2 have been determined by X-ray diffraction analysis. Spectroscopic 13C NMR studies in the solid state, by the CP/MAS technique, and in solution have revealed the presence of the zwitterion structure in Hz0 and D20 solutions. Results have been complemented with “0 NMR and IR data. Keywords: X-ray crystallography;
Solid state NMR
spectroscopy;
1. Introduction
Zwitterion
acid
0
1996 Elsevier
Science
B.V. All rights
COpEt
I
C02H
C02Me
HOzC 2
1
3
N
I \ C>N < coat-Bu
author.
SSDZ 0022-2860(95)09118-l
l-5
I ‘: C>N
4 0022-2860/96/$15.00
Imidazol-I-ylacetic
by ‘H NMR [4]. Among them, compounds and the related 6 were prepared
The analgesic and hypnotic activity of imidazol1-ylacetic acid 1 was formerly related to the presence of an intramolecular hydrogen bond between the carboxylic group and the nitrogen atom at position 3 of the imidazole ring [l]. Later and considering the ratio of ionic molecules to neutral ones obtained from Ebert’s equation, a zwitterion structure was proposed [2]. Also pioneer ‘H NMR work suggested a similar zwitterion structure in D20 solution [3]. We have described recently a series of imidazol1-ylalkanoic acids as new probes to determine the intracellular and extracellular pH and cell volume
* Corresponding
structure;
reserved
COpH COzH
HO& 5
6
106 Table 1 Crystal data, data collectiona
P. Ldpez et al./Journal of Molecular Structure 377 (1996) 105-112
and structure
refinementb
of compounds
1 and 2 Compound
Compound
Formula Crystal size (mm) Symmetry Unit cell determination
CsH6NzG2 0.38 x 0.32 x 0.21 Monoclinic, P2, /n Least-squares fit from 58 reflexions (10 < 20 < 90”) 14.328(l), 8580(l), 4.618(l) A 90.0”, 98.777(5)“, 90.0” 561.1(l), 4 1.4930, 126.115, 264.0 9.575
CsH,>NzQ 0.38 x 0.32 x 0.21 Monoclinic, C2/c Least-squares fit from 64 reflexions (12 < 20 < 90”) 26.927(2), 5.426(l), 14.555(l) .& 90.0”, 104.043(l)“, 90.0” 2063.0(4), 8 1.3664, 212.205, 896 8.801
958 862 (I > 2u (I) criterion) -16/16 O/l1 O/6 Fourier syntheses, Coord. parameters refined 151 0.002 0.041, 0.049 0.18eA-3
1738 1699 (I > 2~ (I) criterion) -31/31 o/7 o/17 Fourier synthesis, Coord. and thermal parameters refined 256 0.07 0.047, 0.058 0.24 e Am3
Unit cell dimensions Packing: V (A3), Z D, (gcme3), M, F (000) p (cm-’ ) Number of reflexions: Measured Observed Range of hkl H atoms Parameters (A/o) max Final R and R, Ap max
1
2
Parameter
and thermal
a Technique: Four circle diffractometer, Philips PW 1100, Bisecting geometry, Graphite oriented monochromator, Cu Ka 1.5418 A, correction, None. Two standard reflexions, frequency, 90 min. w/20 scan. Scanning range for 19: 2 < 0 < 65. Absorption b Solution: Direct methods. Refinements: L.S. on Fobs. w-Scheme: Empirical as to give no trends in < wA2F > vs. < ]Fs] > and < sin Q/x >. Computer and programs: Vax 1 l/750, MLJLTANBO [8], XRAYBO [9], PESOS [lo], PARST [1 11. Scattering factors and anomalous dispersion: Int. Tables for X-Ray Crystallog [12].
The knowledge of the precise ionization state of these new probes in solution is essential to the determination of their membrane permeability properties and to the interpretation of their biological behaviour. In this report we have established unambiguously the zwitterion structures of 1 and 2 in the solid state by X-ray diffraction analysis and the relation with those observed in solution by combined studies of t3C NMR in solid state and in solution.
2. Experimental Imidazol- 1-ylacetic acid (1) [4], (&)-3-ethoxycarbonyl-2-imidazol-1-ylpropionic acid (2) [4], methyl imidazol-1-ylacetate (3) [4], tert-butyl imidazol- I-ylacetate (4) [5], (f)-2-imidazol- l-ylsuccinic acid (5) [6], pyrazol-1-ylacetic acid (6) 173,were prepared according to literature procedures.
2.1. Crystal structure determination
Crystals of compounds 1 and 2 were obtained from aqueous ethanol and absolute ethanol respectively. Crystal and experimental data are given in Tables 1 and 2. The data were collected on a Philips PW 1100 diffractometer, with graphitemonochromated radiation, bisecting geometry, and 1.5” scan width in the w/28 mode. Two reflections were monitored every 90 min, showing no significant variation in the crystals nor in the experimental conditions. The structures were solved by direct methods and refined by leastsquares based on Fobs only. All hydrogen atoms were unambiguously obtained in a difference synthesis and included isotropically in the final cycles of refinement. Weights were chosen to give no trends in < wA2/F > versus < /Fobs1> and < sin 0/X >, by functions of the w = K/[f(F&]-[g(sin e/x)] type, K being a scale factor to ensure that < wA2F > N 1. Final fractional
P. Ldpez et al./Journal of Molecular Structure 377 (1996) 105-112
107
08
(4
(b) Fig. 1. (a) A ORTEP [13] view of compound 1 showing the atomic numbering; displacement ellipsoids are drawn at the 50% probability level; (b) packing of the molecules of compound 1 viewed down the c axis.
atomic coordinates, the full list of bond lengths and angles, and the list of thermal parameters of compounds 1 and 2 are deposited with the B.L.L.D. as Supplementary Publication No. SUP2656 1 (48 pages). 2.2. Infrared measurements IR spectra were recorded in KBr on a BomemDA3 FT-IR spectrophotometer. 2.3. NMR determinations 13C NMR
spectra in solution were recorded
at 50.33 MHz using a Bruker AC200 spectrometer at 25°C. Samples were dissolved in D20, H20 or DMSO-d6 and the concentrations were 0.5 to 1 M. Chemical shifts are expressed in p.p.m. from tetramethylsilane using DMSO-d6 as external or internal reference. Typical conditions were as follows: pulse width, 3 ps (ca. 45”); relaxation delay, 1 s, spectral width, 10 KHz; data points, 32768. 170 NMR spectra were taken at 42.82 MHz on a Bruker AM-360. The enriched compounds 1, 2 and 5 were obtained by partial and total hydrolysis of the corresponding esters with enriched “0 (28%) “0 (32%) water, and spectra conditions were the same as those
108
P. Ldpez et al./Journal of Molecular Structure 377 (1996) 105-112 08
(4
Fig. 2. (a) A ORTEP [13] view of compound 2 showing the atomic numbering; Displacement level; (b) packing of the molecules of compound 2 viewed down the b axis.
ellipsoids
are drawn at the 50% probability
P. Ldpez et al./Journal of Molecular Structure 377 (1996) 105-112 Table 2 Geometric
parameters
Bond lengths
Bond Compound NlLC2 Nl-C5 NlLC6 C2-N3 N3-C4 c44c5 C6-C7 C7p08 c7-09
(A, deg) for compounds (A)
109
1 and 2 Angle
(deg)
1.324(2) 1.379(3) 1.459(2) 1.325(3) 1.373(3) 1.351(3) 1.529(3) 1.259(2) 1.231(2)
C5-Nl-C6 C2-Nl -C6 C2-Nl C5 Nl-C2-N3 C2-N3-C4 N3-C4-C5 N 1-C5-C4 Nl-C66C7 C6-C7-09 C6-C7-08 08-C7-09
125.6(2) 125.4(2) 108.9(l) 108.4(2) 109.0(2) 107.0(2) 106.7(2) 113.4(l) 119.6(l) 113.9(l) 126.4(2)
1.335(2) 1.378(2) 1.468(2) 1.314(2)
C5-NlLC6 C2-Nl C6 C2-Nl-C5 Nl-C2-N3 C2pN3-C4 N3-C4-C5 Nl-C5-C4 Nl-C66ClO Nl-C6-C7 C7-C66ClO C6-C7-09 C6-C7708 088C7-09 C6-CIO-Cl1 ClO-Cll-013 ClO-Cl I-012 012-CllLO13 Cll-013-Cl4 013-c14-Cl5
124.2(l) 128.3(l) 107.5(l) 109.7(l) 108.2(l) 107.6(l) 107.0(l) 111.6(l) 113.1(l) 113.3(l) 111.6(l) 120.5(l) 127.8(l) 114.0(l) 110.4(l) 125.8(2) 123.8(2) 116.4(2) 106.2(3)
H...A 1.66(2) 1.57(3)
D...A 2.645(2) 2.550(2)
1
Compound 2 NlLC2 NlLC5 Nl-C6 C2-N3 N3-C4 c44c5 C66C7 C6-Cl0 C7-08 c7709 cto-Cl1 C11&012 Cl l-013 013-Cl4 c14pc15
Hydrogen
bonds
Compound Compound
1 2
1.370(2) 1.350(2) I .544(2) 1.525(2) 1.229(2) 1.257(2) 1.502(2) 1.202(2) 1.332(2) 1.456(3) 1.496(5)
D-H...A N3-H3 N3-H3
Neighbouring chain interactions Compound 2 012,..H142 2.69(3) 012...H151 2.82(6) 3.01(4) Ol3...H142 3.04(7) O13...H151 2.69(4) O13...Hl52 a +x - l/2, -y + 112 + 1, +z - l/2. b +x, -y, +z - l/2. Symmetry code.
08a 09b
(-x + 1, -y, --z + 1) (-x + 1, -y + 1, -z + 1) (-x+1,-y+l,-z+l) (-x+1,-y+l,-z+l) (-x + 1, y. -z + l/2 + 1)
D-H...A 172(2) 169(2)
P. Lbpez et al./Journal of Molecular Structure 377 (1996) 105-112
110 Table 3 13C NMR chemical
shifts (6, ppm)a of compounds
1-6 at 50.33 MHz
Compound
pD or pH
13CNMR
c2
c4
G
1
5.60 4.70
D2O
134.4
H20
135.2
121.9 122.6
118.5 119.1
CP/MAS DzO
135.2 134.3 133.9
123.5 120.6 120.1
119.9 118.6 118.2
136.4
D2O
137.8 138.4 138.1 138.5 134.3
121.4 126.7 130.0 120.9 127.6 121.6
121.4 120.3 121.0 120.5 121.9 118.4
CP/MAS
133.7
122.9
116.2
169.8
105.0 106.5
131.0 129.5
174.5 174.4
2
3.57 3.33
H20
CP/MAS 3
D2O
4
CP/MAS DMSO-d; CP/MAS
5
3.29
6
139.9 140.7
D20
CP/MAS a Imidazole ring assignments b Very insoluble in D20. ’ Performed in H:‘O.
have been made considering
the data described
C02H
C02R
171.5 172.1 170:271.1’ 171.4 171.5 170.8 170:257.0’ 172.9
in the literature
171.8
CH
CH2
50.8 51.4
171.0 170.4 171.1 170.0 172.0 167.5 169.1 173.2 “0: 267.1’ (R=H) 171.8 (R=H)
59.5 58.9 59.7
51.2 36.4 35.9
59.6
38.8 46.8 47.7 47.7 48.0 36.6
60.0
37.9 53.6 56.2
[21].
previously reported [5]. 13C Cross-polarization magic angle spinning (CP/MAS) NMR spectra
3. Results and discussion
were obtained at 50.33 MHz
Structural study by X-ray diffraction analysis of compounds 1 and 2 (Table 1) revealed that both acids existed in zwitterion form in the solid state (Figs. 1 and 2). The proton from carboxylic oxygen migrates to the N3 in the imidazole ring forming the zwitterion. The charge was delocalized along Nl-C2-N3, and the imidazole ring was planar with maxima deviations of this plane of -0.0008(20) (C5 compound 1) and -0.0016(19) (C4 compound 2). Bond distances and angles (Table 2) were very similar in the two compounds and were on the same range of values as those of the imidazolium cation in imidazolium sulphate dihydrate [14]. Both zwitterion structures were also stabilized by strong hydrogen bonds (distances N3 . . ..O = 2.645(2) A for compound 1 and 2.549(2) A for compound 2) (Table 2). In this situation, both compounds formed infinite chains along the a axis (see Figs. 1 and 2) similar to those observed in some salts of hydroxybenzoic acids
spectrometer.
Samples
2.5 kHz in a 7 mm time was
were
on a Bruker CP-200 rotated
at
3.5 or
Zr02 rotor; the initial contact
1 ms; 90” pulse lengths
for
protons
were 7 ps; 4 s delay between scans and 200 transients were used. Chemical shifts were measured using glycine
(176.1 ppm)
as external
reference. 2.4. Determination
of pKH1 of compounds
1 and 2
The pK, of the carboxylic group of compounds 1 and 2 in Hz0 (pKH’) were determined by 13C NMR (50.33 MHz) using DMSO as external reference. pH titrations (25°C) were performed using 0.8 M solutions of the appropriate compound in H20 adjusting the pH with the addition of HCl. The dependence of chemical shifts of carbons N-CH2 of compound 1 and N-CH of compound 2 vs pH were obtained to calculated the pK,.
P. Ldpez et al./Journal of Molecular Structure 377 (1996) 105-112
[15]. Besides, in compound 2 there are some interactions of note between neighbouring chains (Table 2). Moreover, compound 2 was further linked into antiparallel pairs through ring inte:actions (distance between centroids of 3.52 A) [16], and by a short contact between the centroid of the ring and Cl5 (3.686 A), and H152 (3.38 A). Infrared data in the solid state confirmed the zwitterion structure of compounds 1 and 2. Protonation of N3 was observed by the two broad stretching absorptions in the regions at ca 2500 and ca 1900 cm-‘. These bands corresponded to +NH stretching linked by strong hydrogen bonding as it has been found previously for similar compounds [17-191. As it has been stated, NMR solid-state chemical shifts generally are very similar to those found in solution. Thus, the CP/MAS data can be used as a ‘bridge’ between solid-state structures determined by diffraction techniques and those which exist in solution [20]. 13C NMR chemical shifts values of compounds l-6 in the solid state and in solution are depicted in Table 3. Although it has been reported previously [3], that compound 1 presented a zwitterion structure in D,O, it must be taken into account that the pH of the solution is decisive in the formation of the zwitterion structure, as is observed in amino acids. We have recently reported that compound 1 presented a pKD2 (N+) value of 7.23 [4,5]. Considering the pKD (pK, in D20) and pKH (pK, in H20) relationship [22] pKD = 1.018 pKH + 0.43; pKH2 of compound 1 is 6.68. On the other hand, the pKH’ (COOH) calculated here is 2.1. If we consider compound 1 as an amino acid, these values corresponded to an isoelectric point (Ip) of 4.4 The value of the pH (4.70) obtained when compound 1 was dissolved in Hz0 suggested that the zwitterion form is a maximum in Hz0 solution. This fact is confirmed because of the values of the chemical shifts of all carbons fitted well with those found in solid state (Table 3). Furthermore, the 170 chemical shift value (27 1.1 ppm) of the carboxylic group, is also in agreement with those found in other amino acids in zwitterionic form [23]. The described pKD2 value of compound 2 was 6.86 [4] which corresponds with a pKH2 of 6.32. The carboxylic group has a pKH’ of 1.3.
111
These values give an Ip of 3.8. This value suggest that at 3.30, pH of the solution of compound 2 in Hz0 the contribution of the zwitterion form is slightly lower than that observed in compound 1. This fact is also corroborated by the data obtained in the solid state and the I70 NMR chemical shift value (257.0 ppm) which corresponds well with those values reported for other amino acids with protonated carboxyl O-atoms [23]. In the absence of X-ray data it could be inferred that IR data of diacid 5 in KBr suggested that the (r-COOH (1597 cm-‘) is ionized in the solid state but not the /3-COOH, as has been described for other dicarboxylic mono-amino acids [24]. The 13C NMR data in the solid state and in solution also suggested a similar behaviour to compound 2 in solution. Finally, we can conclude that this work unambiguously proves that imidazol- 1-ylacetic acids behave as amino acids and present zwitterion structures in the solid state and in solution within a certain pH range. In the solid state the packing of ions is governed by strong hydrogen bonds which play a significant role in the structural organization. The presence of rigid hydrogenbonded aggregate structures in these acids makes them especially attractive from the crystal engineering point of view [25].
Acknowledgements This work was supported in part by D.G.I.C.Y.T. (PM-92-01 1, PB-93-0037, and PB-93-0125) and Community of Madrid (AE00219/94). P.L. and P.Z. received F.P.U. fellowships from the Spanish Ministry of Education and Science. We are indebted to Dr C. L6pez for her valuable technical assistance in the performance of CP/MAS spectra.
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