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1-Cyclopentyluracils: Synthesis and conformational analysis by X-ray crystallography and AM1 theoretical calculations
Journal of
MOLECULAR STRUCTURE ELSEVIER
Journal of Molecular Structure 448 (1998) 69-75
1-Cyclopentyluracils: synthesis and conformational analysis by X-ray crystallography and AM1 theoretical calculations C. Tenin a, M. Teijeira b, L. Santana b'*, E. Uriarte b, A. Castifieiras c aDepartment of Physico-Chemistry and Organic Chemistry, University of Vigo, 36200, Vigo, Spain bDepartment of Organic Chemistry, Faculty of Pharmacy, University of Santiago de Compostela, 15706, Santiago de Compostela, Spain CDepartmentof Inorganic Chemistry, Faculty of Pharmacy, University of Santiago de Compostela, 15706, Santiago de Compostela, Spain
Received 5 January 1998; revised 23 February 1998; accepted23 February 1998
Abstract A series of 1-cyclopentyl 5-substituted pyrimidines was synthesized and their energy-minimized structures were determined by AM1 and compared with the crystal structure of the parent compound, 1-cyclopentyluracil (2). As has been reported for other nucleoside analogues, the energy-minimized conformations were similar for all the compounds, but were not fully in agreement with the X-ray crystal structure obtained for 2. © 1998 Elsevier Science B.V. All rights reserved Keywords: Cyclopentyluracils; Synthesis; X-ray crystallography; AM1 theoretical calculations
I. Introduction Intense research in the nucleoside field has produced several 2',3'-dideoxy nucleosides and carbocyclic nucleosides with potent antiviral and/or antitumour activity, some of which are now in clinical use. However, relatively little attention has been paid to simpler analogues also lacking the 5'-hydroxymethyl group, the only exception being tegafur ( 1 ) - - a well known prodrug of the antitumour agent 5-fluorouracil [1].
On the basis of published work [2-5] and our own theoretical studies [6,7] of the conformations of analogues of pyrimidine nucleosides, we decided that these simple nucleoside analogues merited further study. Accordingly, in this work we synthesized a series of analogues with parent compound 1-cyclopentyluracil (2), determined their energy-minimized structures by AM1, and compared these structures with the crystal structure of 2. Special attention was focused on the relative orientation of the pyrimidine and cyclopentane rings, since this can be crucial to biological activity [2,8,9].
2. Experimental 2.1. Synthesis and characterisation
* Corresponding author
Melting points: Reichert Kofler or Biichi 510 apparatus, uncorrected. 1H and 13C spectra: Bruker
0022-2860/98/$19.00 © 1998 Elsevier Science B.V. All fights reserved PH S0022-2860(98)00367-6
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C. Terdn et al./Journal of Molecular Structure 448 (1998) 69-75
AMX (300 MHz), TMS as internal standard (6/ppm, J/Hz). IR spectra: Perkin-Elmer 1640FT spectrometer (KBr discs, v/cm-1). Mass spectra: Hewlett Packard 5988A spectrometer. Microanalyses: Perkin-Elmer 240B instrument (___0.4% of the calculated values). Flash chromatography (FC): Merck silica gel 60 (230-400 mesh). Analytical thin-layer chromatography (TLC): precoated silica gel plates Merck 60 F254 (0.25 mm).
2.2. Condensation of pyrimidine bases with cyclopentyl bromide. General procedure [10] A mixture of the pyrimidine (1.53 mmol) and a catalytic amount of ammonium sulphate in hexamethyldisilazane (12ml) was heated at 135°C for 12 h under an Ar atmosphere. The resulting clear solution was concentrated in vacuo under anhydrous conditions to yield the silylated pyrimidine as a colourless oil. This oil was immediately dissolved in dry 1,2-dichloroethane (4 ml), and a solution of cyclopentyl bromide (1.53 mmol) in the same solvent (4ml) was added. This mixture was heated to reflux under an inert atmosphere (for 48 h for compounds 2 and 3, and for 1 week for compound 4) and then cooled, treated with 6:1 methanol/water and filtered. The filtrate was extracted with CH2C12, and the solution obtained was dried over anhydrous Na2SO4. The drying agent was filtered out and the solvent was evaporated in vacuo; the residue was purified by FC using 3:1 hexane/ethyl acetate as eluent.
2.2.1. 1-Cyclopentyluracil 2 Prepared from uracil in 11% yield. M.p. 170172°C. Rf = 0.2 (3:1 ethyl acetate/hexane). 1H NMR (CDC13): 8.13 (s, 1H, NH), 7.22 (d, 1H, H-6, J = 8.10), 5.72 (dd, 1H, H-6, J = 8.10, 2.30), 4.90 (m, 1H, H-I'), 2.12 (m, 2H, 1H-2' + 1H-5'), 1.90-1.50 (m, 6H, 1H-2' + H-3' + H-4' + 1H-5'). 13C NMR (CDC13): 163.4, 151.3, 141.3, 102.8, 57.1, 31.8, 24.5. IR: 3000, 2875, 2826, 1678, 1613, 1470,1421, 1383, 1267. MS, m/z (%): 180 (M +, 19), 113 (JUra + 1] +, 100), 112 (Ura +, 16), 69 (11). Anal. (C9H12N202) C, H, N. 2.2.2. 1-Cyclopentyl-5-methyluracil 3 Prepared from 5-methyluracil in 10% yield. M.p. 1 7 4 - 1 7 6 ° C . R f = 0.11 (3:1 hexane/ethyl acetate).
1H NMR (CDC13): 9.10 (s, 1H, NH), 7.01 (q, 1H, H-6, J = 1.09), 4.91 (m, 1H, H-I'), 2.10 (m, 2H, 1H-2' + 1H-5'), 1.93 (d, 3H, CH3, J = 1.09), 1.831.55 (m, 6H, 1H-2' + H-3' + H-4' + 1H-5'). 13C NMR (CDC13): 164.1, 151.5, 137.1,111.3, 56.7, 31.6, 24.5. IR: 3174, 3038, 2956, 1690, 1659, 1474, 1269. MS, m~ z (%): 195 ([M + 1] +, 6), 127 ([Thy + 1] +, 63), 126 (Thy +, 100), 83 (16). Anal. (Ca0H14N202) C, H, N.
2.2.3. 1-Cyclopentyl-5-fluorouracil 4 Prepared from 5-fluorouracil in 13% yield. M.p. 1 7 8 - 1 8 0 ° C . R e = 0.68 (1:3 hexane/ethyl acetate). 1H NMR (CDC13): 9.45 (s, 1H, NH), 7.27 (d, 1H, H-6, J = 6.31), 4.93 (m, 1H, H-I'), 2.10 (m, 2H, 1H-2' + 1H-5'), 1.85-1.50 (m, 6H, 1H-2' + H-3' + H-4' + 1H-5'). 13C NMR (CDC13): 157.4 and 157.1 (d, Ca, J = 27), 150.3 (C2), 142.6 and 139.5 (d, C5, J = 237), 125.8 and 125.3 (d, C6, J = 32), 57.4, 31.7, 24.4. IR: 3169, 3048, 1710, 1670, 1659, 1261. MS, m/z (%): 199 ([M + 1] +, 4), 198 (M +, 25), 131 ([FUra + 1] +, 100), 130 (FUra +, 30). Anal. (C9H11FN202) C, H, N. 2.3. Halogenation of 2 using N-halosuccinimide. General procedure [11] To a solution of 2 (100 mg, 0.55 mmol) in acetic acid (3 ml) was added a solution of N-halosuccinimide (0.60 mmol) in acetic acid (6 ml), and the mixture was heated to reflux (for 48 h for compound 5, and for 6 h for compound 6). After the reaction mixture had cooled, the solvent was evaporated and the residue was purified by FC using 4:1 hexane/ethyl acetate as eluent.
2.3.1. 5-Chloro-l-cyclopentyluracil 5 Prepared from 2 and N-chlorosuccinimide in 68% yield. M.p. 196-198°C. R f = 0.60 (1:1 hexane/ethyl acetate). IH NMR (CDC13): 8.90 (s, 1H, NH), 7.45 (s, 1H, H-6), 4.90 (m, 1H, H-I'), 2.15 (m, 2H, 1H-2' + ill-5'), 1.95-1.55 (m, 6H, IH-2' +H-3' +H-4' + 1H5'). 13C NMR (CDC13): 159.4, 150.8, 138.5, 109.3, 57.8, 31.9, 24.4. IR: 3167, 3031, 2967, 1717, 1661, 1613, 1471, 1316. MS, m/z (%): 216 ([M + 2] +, 10), 214 (M +, 30), 149 ([C1Ura + 2] +, 35), 148 ([CIUra + 1] +, 30), 147 ([C1Ura] +, 100), 146 ([CIUra - 1] ÷, 72), 103 (25), 69([M - C1Ura]+,12), 68 ([M - C1Ura - 1] +, 10), 67 ( [ M - C1Ura- 2] +, 15). Anal. (C9H11C1N202) C, H, N.
C. TerSn et al./Journal of Molecular Structure 448 (1998) 69-75
2.3.2. 5-Bromo-l-cyclopentyluracil 6 Prepared from 2 and N-bromosuccinimide in 93% yield. M.p. 200-202°C. R f = 0.55 (1:1 hexane/ethyl acetate). 1H NMR (CDCI3): 8.82 (s, 1H, NH), 7.52 (s, 1H, H-6), 4.90 (m, 1H, H-I'), 2.14 (m, 2H, 1H-2' + IH-5'), 1.90-1.50 (m, 6H,1H-2' + H-3' + H-4' + 1H-5'). 13C NMR (CDC13): 159.6, 151.1, 141.1, 97.1, 57.9, 31.9, 24.4. IR: 3160, 3028, 2965, 2840, 1718, 1654, 1606, 1466, 1272. MS, m/z (%): 260 ([M + 2] +, 36), 258 (M +, 36), 193 ([BrUra + 3] +, 100), 192 ([BrUra + 2] +, 91), 191 ([BrUra + l] +, 98), 190 (BrUra +, 84), 149 (33), 147 (30), 69 ([M - BrUra] +, 13), 68 ([M - BrUra - 1]+, 11), 67 ([M - BrUra - 2] +, 16), 53 (13). Anal. ( C 9 H I I B r N 2 0 2 ) C, H , N. 2.3.3. 5-Iodo-l-cyclopentyluracil 7 A mixture of 2 (93mg, 0.52mmol), iodine (260mg, 1.04mmol) and 0.75M nitric acid (0.68 ml) in dioxane (8 ml) was stirred for 2 h at 100°C. After the reaction mixture had cooled, the solvent was evaporated in vacuo and the residue was purified by FC using 4:1 hexane/ethyl acetate as eluent which gave 7 (148mg, 94% yield). M.p. 220-222°C. Rf = 0.72 (1:3 hexane/ethyl acetate).
1H NMR (CDC13): 9.09 (s, 1H, NH); 7.61 (s, 1H, H-6), 4.88 (m, 1H, H-I'), 2.13 (m, 2H, 1H-2' + 1H5'), 1.90-1.53 (m, 6H, 1H-2' + H-3' + H-4' + 1H-5').
13C NMR (CDC13): 160.3, 151.1, 146.2, 68.3, 57.9, 31.9, 24.4. IR: 3148, 1955, 1839, 1699, 1664, 1599, 1427, 1271. MS, m/z (%): 306 (M +, 43), 239 ([IUra + 1]+, 48), 238 (IUra +, 100), 195 (34). Anal. ( C 9 H I I I N z O 2 ) C, H, N. 2.4. Structural study 2.4.1. X-ray collection data collection and data reduction A colourless prismatic crystal of 1-cyclopentyluridine (2) was mounted on a glass fibre in an Enraf-Nonius CAD4 automatic diffractometer [12] and used for data collection. Cell constants and an orientation matrix for data collection were obtained by least-squares refinement of the diffraction data for 25 reflections in the range of 23.04 < 0 < 42.83 ° . Data were collected at 293 K using Cu Ka radiation (X = 1.54184,~) with the co scan technique and were corrected for Lorentz and polarization effects [13]. A semi-empirical absorption correction was also
Table 1 Collection of crystal data and refinement of structure of 1-cyclopentyluridine Empirical formula Formula weight Temperature Crystal system, space group Unit cell dimensions
Volume z, Calculated density Absorption coefficient F(000) Crystal size 0 range for data collection Index ranges Observed unique reflections Completeness to 20 = 74.31 Max. and min. transmission Data/restraints/parameters Goodness-of-fit on F 2 Final R indices [I > 2a(/)] R indices (all data) Absolute structure parameter Extinction coefficient Max. peak-trough difference
71
CgH12N202 180.21 292(2) K Tetragonal, P412 t2 a = 7.1343(7) A b = 7.1343(5) ,~ c = 37.031(2) ,~ 1884.8(2) ~3 8, 1.270 Mg m 0.752 mm -1 768 0.35 × 0.25 × 0.10 mm 3 2.39-74.31 °
O<~h<~8,0<~k<~8,-46<~l<~O 2280/1917 (Ri,t = 0.0137) 100.0% 0,972 and 0,893 1917/0/138 1.032 RI = 0.0442, wR2 = 0,1151 Rt =0.0790, wR2 =0,1339 0.00(1) 0.0035(5) 0.161 and -0.142 e- ~ 3
a = 9o.oo(-)° /3 = 90.00(-) ° ,y = 90.00(-) °
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C. Terdn et al./Journal of Molecular Structure 448 (1998) 69-75
Table 2 Atomic coordinates ( x 104)and equivalent isotropic displacement parameters (.~ 2x 103)for 1-cyclopentyluridine. Ueqis defined as one-third of the trace of the orthogonalized Uij tensor
O(1) 0(2) N(3) N(4) C(5) C(6) C(7) C(8) C(9) C(10) C(I 1) C(12) C(13)
x
y
z
Ueq
7722(2) 7416(4) 7552(3) 7386(3) 7565(3) 7409(4) 7262(4) 7258(4) 7354(5) 8893(7) 8171(10) 6115(10) 5530(6)
5809(2) 1629(2) 3640(2) 2718(2) 4169(3) 1847(3) 419(3) 895(3) 3166(3) 2231 (5) 2242(8) 2649(7) 2625(5)
338(1) 1246(1) 782(1) 182(1) 427( 1) 916(1) 649(1) 302(1) -208(1) -422(1 ) -807(1) -791( 1) -396(1)
53(1) 75(1) 47(1) 47(1) 42( 1) 51(1) 52(1) 51(1) 67(1) 117(2) 193(4) 195(4) 107(1)
made [13]. The crystal data, experimental details and refinement results are summarized in Table 1.
the corresponding refined atomic coordinates for the non-hydrogen atoms are given in Table 2.
2.4.2. Structure solution and refinement The structure was solved by direct methods [14], which revealed the position of all non-hydrogen atoms, and refined on F 2 by a full-matrix least-squares procedure using anisotropic displacement parameters for all non-hydrogen atoms [15]. Hydrogen atoms were located in difference maps and refined isotropically, except those linked to C10, C11, C12 and C13 the positions of which were generated from assumed geometries (C-H = 0.97 ,~), and then refined using a riding model (AFIX 23). The absolute structure was determined by racemic twinning [16]. Atomic scattering factors were taken from International Tables for X-Ray Crystallography [17]. Molecular graphics used ZORTEP [18]. After all shift/e.s.d, ratios were less than 0.001, the refinement converged to the agreement factors listed in Table 1;
2.4.3. AM1 theoretical study Optimal molecular geometries were calculated for compounds 2 - 7 by means of the semi-empirical quantum-chemical method AM1 [19] as implemented by the AMPAC program running on an SGI workstation [20]. Stable molecular geometries were searched for by varying the torsion angle x[C(5)N(4)-C(9)-C(13)] about the pseudo-glycosidic bond (N4-C9), from 0-360 ° in 10° increments. The preferred conformation of the cyclopentane ring of each of the resulting rotamers was also examined.
3. Results and discussion
Compounds 2 - 4 were prepared by condensation of the trimethylsilylated base (uracil, thymine or O
OTMS
~
Br
+
,~N~[
HN~I, R
CH2C12
R
oA'NJJ
(7
TMSO
2 R=H 3 R=CH 3 4
Scheme 1.
R=F
C. Terdn et al./Journal of Molecular Structure 448 (1998) 69-75
73
o x
0
5 6
o
X=CI X=Br 0
b
'
7 a) NXS (X = CI, Br), AcOH. b) 12, HNO3, dioxane Scheme 2.
5-fluorouracil) with bromocyclopentane (Scheme 1). The other 5-halogenated analogues were prepared from 2 as indicated in Scheme 2. Fig. 1 shows the molecular structure obtained for 2 by X-ray diffraction, together with the atomic numbering scheme used. A selection of bond lengths and angles in 2 is listed in Table 3. The bond lengths and angles in 2 are unremarkable, being similar to those found in other carbocyclic pyrimidine nucleosides [4,7]. The planar base (r.m.s. deviation from the fitted plane, 0.0053 ,~) makes an angle of 87.0(1) ° with the best plane through atoms C(10)-C(13) of the cyclopentane ring, which assumes
an envelope conformation with C(9) 0.538(5) below this plane. The appreciable difference between this dihedral angle in 2 and the value of 99.8 ° found for cis-l-[2-(hydroxymethyl)cyclopentyl]uridine [7] shows the extent to which the steric effect of the hydroxymethyl group can determine the orientation of the base. This was the only appreciable effect of this group, with the corresponding bond angles at the pseudo-glycosidic carbon [C(9)] in 2 and in cis- l-[ 2-(hydroxymethyl)cyclopentyl]uridine being practically the same. As would be expected, in the crystal lattice pairs of molecules are linked by hydrogen-bonding
O1 Fig. 1. ORTEP plot and numbering scheme of atoms. Thermal ellipsoids represent 50% probabilities.
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C. Terdn et aL/Journal of Molecular Structure 448 (1998) 69-75
Table 3 Selected bond lengths (,~) and angles (deg) for 1-cyclopentyluridine O(1)-C(5) O(2)-C(6) N(3)-C(5) N(3)-C(6) N(4)-C(8) N(4)-C(5) N(4)-C(9) C(6)-C(7) C(7)-C(8) C(9)-C(10) C(9)-C(13) C(IO)-C(11) C(11)-C(12) C(12)-C(13) C(5)-N(3)-C(6) C(8)-N(4)-C(5) C(8)-N(4)-C(9) C(5)-N(4)-C(9) O(1)-C(5)-N(3) O( I )-C(5)-N(4) N(3)-C(5)-N(4) O(2)-C(6)-N(3) O(2)-C(6)-C(7) N(3)-C(6)-C(7) C(8)-C(7)-C(6) C(7)-C(8)-N(4) N(4)-C(9)-C(10) N(4)-C(9)-C(13) C( 10)-C(9)-C(13) C(9)-C(10)-C(11) C(12)-C(11)-C(10) C(11)-C(12)-C(13) C(12)-C(13)-C(9)
1.221(2) 1.232(2) 1.369(2) 1.375(3) 1.377(3) 1.383(2) 1.478(2) 1.425(3) 1.329(3) 1.510(4) 1.525(4) 1.517(6) 1.496(9) 1.524(5) 127.08(17) 120.12(16) 121.16(17) 118.71(17) 121.58(17) 123.32(18) 115.10(17) 118.32(19) 126.8(2) 114.83(18) 119.3(2) 123.5(2) 114.0(2) 113.8(3) 105.6(3) 104.2(4) 107.3(4) 107.7(4) 101.7(4)
interactions. Specifically, the contact distance between 0(2) and NH(3) and the symmetry-related atoms defined by (x,y,z) ---. ( - x + 3/2, y + 1/2, 1 / 4 - z ) is only 2.756 A, the corresponding O(2)...H bond and HN(3) contact distances being 1.84(3) and 0.92 .~, and the N...H...O angle 177.2 °. The AM1 theoretical calculations indicated the maximum energy difference between the rotamers of each compound to be 7 kcal mol-l. Regardless of Table 4 Selected torsion angles (deg) for 1-cyclopentyluridine C(8)-N(4)-C(9)-C(10) C(5)-N(4)-C(9)-C(10) C(8)-N(4)-C(9)-C(13) C(5)-N(4)-C(9)-C(13)
-58.4(4) 121.0(3) 62.8(3) - 117.7(3)
the substituent at position five of the pyrimidine, a global energy minimum was observed at X between -115 and - 1 2 0 °. The value of X derived from the X-ray data for 2 (-117.7°; Table 4) also lies in this range, as does the corresponding torsional angle for tegafur (1) [21]. For these X the pyrimidine base lies a n t i to the cyclopentane ring, which is the orientation observed in natural nucleosides and in most nucleoside analogues. A local minimum about 2 kcal mol -l higher in energy was observed at X close to 60 °, at which the base lies s y n to the cyclopentyl ring. Once again, this minimum was unaffected by the nature of the substituent at position five of the pyrimidine. The cyclopentane ring showed great conformational flexibility. For X between - 9 0 and - 1 2 0 ° it adopted twist and envelope conformations in both the northern and southern hemispheres of the pseudo-rotational cycle. However, for X of around 60 °, i.e. with the pyrimidine in the s y n orientation, the cyclopentane preferred to adopt an envelope conformation with the pseudo-glycosidic carbon [C(9)] e x o or below the ring plane (equivalent to the I E conformation of furanoses). Interestingly, this is the conformation observed for the cyclopentane in the X-ray study, for the conformer with X = -117.7 °. By contrast, regardless of the orientation of the 5-fluorouracil, the furan ring of tegafur adopts an envelope conformation with the glycosidic carbon e n d o (IE).
Acknowledgements
Work at Vigo University was supported by Vicerrectorado de Investigaci6n of Vigo University. Work at the University of Santiago de Compostela was supported by the DGICYT (PM92-0090) and the Xunta de Galicia (XUGA20312B92 and 20306B95). References
[1] Y. Takemasa, M. Suehara, Y. Koyama, T. Kimura, Cancer Chemother. Jpn. 1 (1974) 259. [2] W. Saenger, Principles of Nucleic Acid Structure, Springer, New York, 1994, pp. 9-104. [3] P.C. Yates, Struct. Chem. 2 (1991) 611. [4] T. Kovacs, L. Parkanyi, I. Pelczer, F. Cervantes-Lee, K.H. Pannell, P.F. Torrence, J. Med. Chem. 34 (1991) 2595.
C. Ter6n et al./Journal of Molecular Structure 448 (1998) 69-75
[5] H. Maag, J.T. Nelson, J.L. Rios Steiner, E.J. Pfisbe, J. Med. Chem. 37 (1994) 431. and references cited therein. [6] L. Santana, M. Teijeira, E. Uriarte, J. Balzarini, E. De Clercq, Nucleosides and Nucleotides 14 (3-5) (1995) 521. [7] L. Santana, M. Teijeira, E. Uriarte, C. Ter~n, U. Casellato, R. Graziani, Nucleosides and Nucleotides 15 (6) (1996) 1179. [8] J.B. Rodrfguez, V. M~quez, M.C. Nicklans, H. Mitsuya, J.J. Barchi, J. Med. Chem. 37 (1994) 3389. [9] S. Neidle, L. Urpi, P. Serafinowsky, D. Whitby, Biochem. Biophys. Res. Commun. 161 (2) (1989) 910. [10] H.O. Kim, S.K. Ahn, A.J. Alves, J.W. Beach, L.S. Jeong, B.G. Choi, P. Van Roey, R.F. Schinazi, C.K. Chu, J. Med. Chem. 35 (1992) 1987. [11] H. Awano, S. Shuto, M. Baba, T. Kira, S. Shigeta, A. Matsuda, Bioorg. Med Chem. Lett. 4 (1994) 367. [12] CAD4-Express Software, Version 5.1, Enraf-Nonius, Delft, Netherlands, 1995.
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[ 13] A.L. Spek, HELENA. A Program for Data Reduction of CAD4 Data, University of Utrecht, Netherlands, 1997. [14] G.M. Sheldrick, Acta Crystallogr. Sect. A: 46 (1990) 467. [15] G.M. Sheldrick, SHELXL-97. Program for the Refinement of Crystal Structures, University of G6ttingen, Germany, 1997. [16] H.D. Flack, Acta Crystallogr. Sect. A: 39 (1985) 876. [17] International Tables for X-Ray Crystallography, Vol. C, Kluwer, Dordrecht, Netherlands, 1995. [18] L. Zsolnai, ZORTEP. A Program for the Presentation of Thermal Ellipsoids, University of Heidelberg, Germany, 1997. [19] M.J.S. Dewar, E.G. Zoebisch, E.F. Healy, J.J.P. Stewart, J. Am. Chem. Soc. 107 (1985) 3902. [20] BIOSYM Technologies, Inc., 10065 Barnes Canyon Road, San Diego, CA 92121. [21] Y. Nakai, K. Yamamoto, K. Terada, T. Uchida, N. Shimizu, S. Nishigaki, Chem. Pharm. Bull. 30 (1982) 2629.
MOLECULAR STRUCTURE ELSEVIER
Journal of Molecular Structure 448 (1998) 69-75
1-Cyclopentyluracils: synthesis and conformational analysis by X-ray crystallography and AM1 theoretical calculations C. Tenin a, M. Teijeira b, L. Santana b'*, E. Uriarte b, A. Castifieiras c aDepartment of Physico-Chemistry and Organic Chemistry, University of Vigo, 36200, Vigo, Spain bDepartment of Organic Chemistry, Faculty of Pharmacy, University of Santiago de Compostela, 15706, Santiago de Compostela, Spain CDepartmentof Inorganic Chemistry, Faculty of Pharmacy, University of Santiago de Compostela, 15706, Santiago de Compostela, Spain
Received 5 January 1998; revised 23 February 1998; accepted23 February 1998
Abstract A series of 1-cyclopentyl 5-substituted pyrimidines was synthesized and their energy-minimized structures were determined by AM1 and compared with the crystal structure of the parent compound, 1-cyclopentyluracil (2). As has been reported for other nucleoside analogues, the energy-minimized conformations were similar for all the compounds, but were not fully in agreement with the X-ray crystal structure obtained for 2. © 1998 Elsevier Science B.V. All rights reserved Keywords: Cyclopentyluracils; Synthesis; X-ray crystallography; AM1 theoretical calculations
I. Introduction Intense research in the nucleoside field has produced several 2',3'-dideoxy nucleosides and carbocyclic nucleosides with potent antiviral and/or antitumour activity, some of which are now in clinical use. However, relatively little attention has been paid to simpler analogues also lacking the 5'-hydroxymethyl group, the only exception being tegafur ( 1 ) - - a well known prodrug of the antitumour agent 5-fluorouracil [1].
On the basis of published work [2-5] and our own theoretical studies [6,7] of the conformations of analogues of pyrimidine nucleosides, we decided that these simple nucleoside analogues merited further study. Accordingly, in this work we synthesized a series of analogues with parent compound 1-cyclopentyluracil (2), determined their energy-minimized structures by AM1, and compared these structures with the crystal structure of 2. Special attention was focused on the relative orientation of the pyrimidine and cyclopentane rings, since this can be crucial to biological activity [2,8,9].
2. Experimental 2.1. Synthesis and characterisation
* Corresponding author
Melting points: Reichert Kofler or Biichi 510 apparatus, uncorrected. 1H and 13C spectra: Bruker
0022-2860/98/$19.00 © 1998 Elsevier Science B.V. All fights reserved PH S0022-2860(98)00367-6
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C. Terdn et al./Journal of Molecular Structure 448 (1998) 69-75
AMX (300 MHz), TMS as internal standard (6/ppm, J/Hz). IR spectra: Perkin-Elmer 1640FT spectrometer (KBr discs, v/cm-1). Mass spectra: Hewlett Packard 5988A spectrometer. Microanalyses: Perkin-Elmer 240B instrument (___0.4% of the calculated values). Flash chromatography (FC): Merck silica gel 60 (230-400 mesh). Analytical thin-layer chromatography (TLC): precoated silica gel plates Merck 60 F254 (0.25 mm).
2.2. Condensation of pyrimidine bases with cyclopentyl bromide. General procedure [10] A mixture of the pyrimidine (1.53 mmol) and a catalytic amount of ammonium sulphate in hexamethyldisilazane (12ml) was heated at 135°C for 12 h under an Ar atmosphere. The resulting clear solution was concentrated in vacuo under anhydrous conditions to yield the silylated pyrimidine as a colourless oil. This oil was immediately dissolved in dry 1,2-dichloroethane (4 ml), and a solution of cyclopentyl bromide (1.53 mmol) in the same solvent (4ml) was added. This mixture was heated to reflux under an inert atmosphere (for 48 h for compounds 2 and 3, and for 1 week for compound 4) and then cooled, treated with 6:1 methanol/water and filtered. The filtrate was extracted with CH2C12, and the solution obtained was dried over anhydrous Na2SO4. The drying agent was filtered out and the solvent was evaporated in vacuo; the residue was purified by FC using 3:1 hexane/ethyl acetate as eluent.
2.2.1. 1-Cyclopentyluracil 2 Prepared from uracil in 11% yield. M.p. 170172°C. Rf = 0.2 (3:1 ethyl acetate/hexane). 1H NMR (CDC13): 8.13 (s, 1H, NH), 7.22 (d, 1H, H-6, J = 8.10), 5.72 (dd, 1H, H-6, J = 8.10, 2.30), 4.90 (m, 1H, H-I'), 2.12 (m, 2H, 1H-2' + 1H-5'), 1.90-1.50 (m, 6H, 1H-2' + H-3' + H-4' + 1H-5'). 13C NMR (CDC13): 163.4, 151.3, 141.3, 102.8, 57.1, 31.8, 24.5. IR: 3000, 2875, 2826, 1678, 1613, 1470,1421, 1383, 1267. MS, m/z (%): 180 (M +, 19), 113 (JUra + 1] +, 100), 112 (Ura +, 16), 69 (11). Anal. (C9H12N202) C, H, N. 2.2.2. 1-Cyclopentyl-5-methyluracil 3 Prepared from 5-methyluracil in 10% yield. M.p. 1 7 4 - 1 7 6 ° C . R f = 0.11 (3:1 hexane/ethyl acetate).
1H NMR (CDC13): 9.10 (s, 1H, NH), 7.01 (q, 1H, H-6, J = 1.09), 4.91 (m, 1H, H-I'), 2.10 (m, 2H, 1H-2' + 1H-5'), 1.93 (d, 3H, CH3, J = 1.09), 1.831.55 (m, 6H, 1H-2' + H-3' + H-4' + 1H-5'). 13C NMR (CDC13): 164.1, 151.5, 137.1,111.3, 56.7, 31.6, 24.5. IR: 3174, 3038, 2956, 1690, 1659, 1474, 1269. MS, m~ z (%): 195 ([M + 1] +, 6), 127 ([Thy + 1] +, 63), 126 (Thy +, 100), 83 (16). Anal. (Ca0H14N202) C, H, N.
2.2.3. 1-Cyclopentyl-5-fluorouracil 4 Prepared from 5-fluorouracil in 13% yield. M.p. 1 7 8 - 1 8 0 ° C . R e = 0.68 (1:3 hexane/ethyl acetate). 1H NMR (CDC13): 9.45 (s, 1H, NH), 7.27 (d, 1H, H-6, J = 6.31), 4.93 (m, 1H, H-I'), 2.10 (m, 2H, 1H-2' + 1H-5'), 1.85-1.50 (m, 6H, 1H-2' + H-3' + H-4' + 1H-5'). 13C NMR (CDC13): 157.4 and 157.1 (d, Ca, J = 27), 150.3 (C2), 142.6 and 139.5 (d, C5, J = 237), 125.8 and 125.3 (d, C6, J = 32), 57.4, 31.7, 24.4. IR: 3169, 3048, 1710, 1670, 1659, 1261. MS, m/z (%): 199 ([M + 1] +, 4), 198 (M +, 25), 131 ([FUra + 1] +, 100), 130 (FUra +, 30). Anal. (C9H11FN202) C, H, N. 2.3. Halogenation of 2 using N-halosuccinimide. General procedure [11] To a solution of 2 (100 mg, 0.55 mmol) in acetic acid (3 ml) was added a solution of N-halosuccinimide (0.60 mmol) in acetic acid (6 ml), and the mixture was heated to reflux (for 48 h for compound 5, and for 6 h for compound 6). After the reaction mixture had cooled, the solvent was evaporated and the residue was purified by FC using 4:1 hexane/ethyl acetate as eluent.
2.3.1. 5-Chloro-l-cyclopentyluracil 5 Prepared from 2 and N-chlorosuccinimide in 68% yield. M.p. 196-198°C. R f = 0.60 (1:1 hexane/ethyl acetate). IH NMR (CDC13): 8.90 (s, 1H, NH), 7.45 (s, 1H, H-6), 4.90 (m, 1H, H-I'), 2.15 (m, 2H, 1H-2' + ill-5'), 1.95-1.55 (m, 6H, IH-2' +H-3' +H-4' + 1H5'). 13C NMR (CDC13): 159.4, 150.8, 138.5, 109.3, 57.8, 31.9, 24.4. IR: 3167, 3031, 2967, 1717, 1661, 1613, 1471, 1316. MS, m/z (%): 216 ([M + 2] +, 10), 214 (M +, 30), 149 ([C1Ura + 2] +, 35), 148 ([CIUra + 1] +, 30), 147 ([C1Ura] +, 100), 146 ([CIUra - 1] ÷, 72), 103 (25), 69([M - C1Ura]+,12), 68 ([M - C1Ura - 1] +, 10), 67 ( [ M - C1Ura- 2] +, 15). Anal. (C9H11C1N202) C, H, N.
C. TerSn et al./Journal of Molecular Structure 448 (1998) 69-75
2.3.2. 5-Bromo-l-cyclopentyluracil 6 Prepared from 2 and N-bromosuccinimide in 93% yield. M.p. 200-202°C. R f = 0.55 (1:1 hexane/ethyl acetate). 1H NMR (CDCI3): 8.82 (s, 1H, NH), 7.52 (s, 1H, H-6), 4.90 (m, 1H, H-I'), 2.14 (m, 2H, 1H-2' + IH-5'), 1.90-1.50 (m, 6H,1H-2' + H-3' + H-4' + 1H-5'). 13C NMR (CDC13): 159.6, 151.1, 141.1, 97.1, 57.9, 31.9, 24.4. IR: 3160, 3028, 2965, 2840, 1718, 1654, 1606, 1466, 1272. MS, m/z (%): 260 ([M + 2] +, 36), 258 (M +, 36), 193 ([BrUra + 3] +, 100), 192 ([BrUra + 2] +, 91), 191 ([BrUra + l] +, 98), 190 (BrUra +, 84), 149 (33), 147 (30), 69 ([M - BrUra] +, 13), 68 ([M - BrUra - 1]+, 11), 67 ([M - BrUra - 2] +, 16), 53 (13). Anal. ( C 9 H I I B r N 2 0 2 ) C, H , N. 2.3.3. 5-Iodo-l-cyclopentyluracil 7 A mixture of 2 (93mg, 0.52mmol), iodine (260mg, 1.04mmol) and 0.75M nitric acid (0.68 ml) in dioxane (8 ml) was stirred for 2 h at 100°C. After the reaction mixture had cooled, the solvent was evaporated in vacuo and the residue was purified by FC using 4:1 hexane/ethyl acetate as eluent which gave 7 (148mg, 94% yield). M.p. 220-222°C. Rf = 0.72 (1:3 hexane/ethyl acetate).
1H NMR (CDC13): 9.09 (s, 1H, NH); 7.61 (s, 1H, H-6), 4.88 (m, 1H, H-I'), 2.13 (m, 2H, 1H-2' + 1H5'), 1.90-1.53 (m, 6H, 1H-2' + H-3' + H-4' + 1H-5').
13C NMR (CDC13): 160.3, 151.1, 146.2, 68.3, 57.9, 31.9, 24.4. IR: 3148, 1955, 1839, 1699, 1664, 1599, 1427, 1271. MS, m/z (%): 306 (M +, 43), 239 ([IUra + 1]+, 48), 238 (IUra +, 100), 195 (34). Anal. ( C 9 H I I I N z O 2 ) C, H, N. 2.4. Structural study 2.4.1. X-ray collection data collection and data reduction A colourless prismatic crystal of 1-cyclopentyluridine (2) was mounted on a glass fibre in an Enraf-Nonius CAD4 automatic diffractometer [12] and used for data collection. Cell constants and an orientation matrix for data collection were obtained by least-squares refinement of the diffraction data for 25 reflections in the range of 23.04 < 0 < 42.83 ° . Data were collected at 293 K using Cu Ka radiation (X = 1.54184,~) with the co scan technique and were corrected for Lorentz and polarization effects [13]. A semi-empirical absorption correction was also
Table 1 Collection of crystal data and refinement of structure of 1-cyclopentyluridine Empirical formula Formula weight Temperature Crystal system, space group Unit cell dimensions
Volume z, Calculated density Absorption coefficient F(000) Crystal size 0 range for data collection Index ranges Observed unique reflections Completeness to 20 = 74.31 Max. and min. transmission Data/restraints/parameters Goodness-of-fit on F 2 Final R indices [I > 2a(/)] R indices (all data) Absolute structure parameter Extinction coefficient Max. peak-trough difference
71
CgH12N202 180.21 292(2) K Tetragonal, P412 t2 a = 7.1343(7) A b = 7.1343(5) ,~ c = 37.031(2) ,~ 1884.8(2) ~3 8, 1.270 Mg m 0.752 mm -1 768 0.35 × 0.25 × 0.10 mm 3 2.39-74.31 °
O<~h<~8,0<~k<~8,-46<~l<~O 2280/1917 (Ri,t = 0.0137) 100.0% 0,972 and 0,893 1917/0/138 1.032 RI = 0.0442, wR2 = 0,1151 Rt =0.0790, wR2 =0,1339 0.00(1) 0.0035(5) 0.161 and -0.142 e- ~ 3
a = 9o.oo(-)° /3 = 90.00(-) ° ,y = 90.00(-) °
72
C. Terdn et al./Journal of Molecular Structure 448 (1998) 69-75
Table 2 Atomic coordinates ( x 104)and equivalent isotropic displacement parameters (.~ 2x 103)for 1-cyclopentyluridine. Ueqis defined as one-third of the trace of the orthogonalized Uij tensor
O(1) 0(2) N(3) N(4) C(5) C(6) C(7) C(8) C(9) C(10) C(I 1) C(12) C(13)
x
y
z
Ueq
7722(2) 7416(4) 7552(3) 7386(3) 7565(3) 7409(4) 7262(4) 7258(4) 7354(5) 8893(7) 8171(10) 6115(10) 5530(6)
5809(2) 1629(2) 3640(2) 2718(2) 4169(3) 1847(3) 419(3) 895(3) 3166(3) 2231 (5) 2242(8) 2649(7) 2625(5)
338(1) 1246(1) 782(1) 182(1) 427( 1) 916(1) 649(1) 302(1) -208(1) -422(1 ) -807(1) -791( 1) -396(1)
53(1) 75(1) 47(1) 47(1) 42( 1) 51(1) 52(1) 51(1) 67(1) 117(2) 193(4) 195(4) 107(1)
made [13]. The crystal data, experimental details and refinement results are summarized in Table 1.
the corresponding refined atomic coordinates for the non-hydrogen atoms are given in Table 2.
2.4.2. Structure solution and refinement The structure was solved by direct methods [14], which revealed the position of all non-hydrogen atoms, and refined on F 2 by a full-matrix least-squares procedure using anisotropic displacement parameters for all non-hydrogen atoms [15]. Hydrogen atoms were located in difference maps and refined isotropically, except those linked to C10, C11, C12 and C13 the positions of which were generated from assumed geometries (C-H = 0.97 ,~), and then refined using a riding model (AFIX 23). The absolute structure was determined by racemic twinning [16]. Atomic scattering factors were taken from International Tables for X-Ray Crystallography [17]. Molecular graphics used ZORTEP [18]. After all shift/e.s.d, ratios were less than 0.001, the refinement converged to the agreement factors listed in Table 1;
2.4.3. AM1 theoretical study Optimal molecular geometries were calculated for compounds 2 - 7 by means of the semi-empirical quantum-chemical method AM1 [19] as implemented by the AMPAC program running on an SGI workstation [20]. Stable molecular geometries were searched for by varying the torsion angle x[C(5)N(4)-C(9)-C(13)] about the pseudo-glycosidic bond (N4-C9), from 0-360 ° in 10° increments. The preferred conformation of the cyclopentane ring of each of the resulting rotamers was also examined.
3. Results and discussion
Compounds 2 - 4 were prepared by condensation of the trimethylsilylated base (uracil, thymine or O
OTMS
~
Br
+
,~N~[
HN~I, R
CH2C12
R
oA'NJJ
(7
TMSO
2 R=H 3 R=CH 3 4
Scheme 1.
R=F
C. Terdn et al./Journal of Molecular Structure 448 (1998) 69-75
73
o x
0
5 6
o
X=CI X=Br 0
b
'
7 a) NXS (X = CI, Br), AcOH. b) 12, HNO3, dioxane Scheme 2.
5-fluorouracil) with bromocyclopentane (Scheme 1). The other 5-halogenated analogues were prepared from 2 as indicated in Scheme 2. Fig. 1 shows the molecular structure obtained for 2 by X-ray diffraction, together with the atomic numbering scheme used. A selection of bond lengths and angles in 2 is listed in Table 3. The bond lengths and angles in 2 are unremarkable, being similar to those found in other carbocyclic pyrimidine nucleosides [4,7]. The planar base (r.m.s. deviation from the fitted plane, 0.0053 ,~) makes an angle of 87.0(1) ° with the best plane through atoms C(10)-C(13) of the cyclopentane ring, which assumes
an envelope conformation with C(9) 0.538(5) below this plane. The appreciable difference between this dihedral angle in 2 and the value of 99.8 ° found for cis-l-[2-(hydroxymethyl)cyclopentyl]uridine [7] shows the extent to which the steric effect of the hydroxymethyl group can determine the orientation of the base. This was the only appreciable effect of this group, with the corresponding bond angles at the pseudo-glycosidic carbon [C(9)] in 2 and in cis- l-[ 2-(hydroxymethyl)cyclopentyl]uridine being practically the same. As would be expected, in the crystal lattice pairs of molecules are linked by hydrogen-bonding
O1 Fig. 1. ORTEP plot and numbering scheme of atoms. Thermal ellipsoids represent 50% probabilities.
74
C. Terdn et aL/Journal of Molecular Structure 448 (1998) 69-75
Table 3 Selected bond lengths (,~) and angles (deg) for 1-cyclopentyluridine O(1)-C(5) O(2)-C(6) N(3)-C(5) N(3)-C(6) N(4)-C(8) N(4)-C(5) N(4)-C(9) C(6)-C(7) C(7)-C(8) C(9)-C(10) C(9)-C(13) C(IO)-C(11) C(11)-C(12) C(12)-C(13) C(5)-N(3)-C(6) C(8)-N(4)-C(5) C(8)-N(4)-C(9) C(5)-N(4)-C(9) O(1)-C(5)-N(3) O( I )-C(5)-N(4) N(3)-C(5)-N(4) O(2)-C(6)-N(3) O(2)-C(6)-C(7) N(3)-C(6)-C(7) C(8)-C(7)-C(6) C(7)-C(8)-N(4) N(4)-C(9)-C(10) N(4)-C(9)-C(13) C( 10)-C(9)-C(13) C(9)-C(10)-C(11) C(12)-C(11)-C(10) C(11)-C(12)-C(13) C(12)-C(13)-C(9)
1.221(2) 1.232(2) 1.369(2) 1.375(3) 1.377(3) 1.383(2) 1.478(2) 1.425(3) 1.329(3) 1.510(4) 1.525(4) 1.517(6) 1.496(9) 1.524(5) 127.08(17) 120.12(16) 121.16(17) 118.71(17) 121.58(17) 123.32(18) 115.10(17) 118.32(19) 126.8(2) 114.83(18) 119.3(2) 123.5(2) 114.0(2) 113.8(3) 105.6(3) 104.2(4) 107.3(4) 107.7(4) 101.7(4)
interactions. Specifically, the contact distance between 0(2) and NH(3) and the symmetry-related atoms defined by (x,y,z) ---. ( - x + 3/2, y + 1/2, 1 / 4 - z ) is only 2.756 A, the corresponding O(2)...H bond and HN(3) contact distances being 1.84(3) and 0.92 .~, and the N...H...O angle 177.2 °. The AM1 theoretical calculations indicated the maximum energy difference between the rotamers of each compound to be 7 kcal mol-l. Regardless of Table 4 Selected torsion angles (deg) for 1-cyclopentyluridine C(8)-N(4)-C(9)-C(10) C(5)-N(4)-C(9)-C(10) C(8)-N(4)-C(9)-C(13) C(5)-N(4)-C(9)-C(13)
-58.4(4) 121.0(3) 62.8(3) - 117.7(3)
the substituent at position five of the pyrimidine, a global energy minimum was observed at X between -115 and - 1 2 0 °. The value of X derived from the X-ray data for 2 (-117.7°; Table 4) also lies in this range, as does the corresponding torsional angle for tegafur (1) [21]. For these X the pyrimidine base lies a n t i to the cyclopentane ring, which is the orientation observed in natural nucleosides and in most nucleoside analogues. A local minimum about 2 kcal mol -l higher in energy was observed at X close to 60 °, at which the base lies s y n to the cyclopentyl ring. Once again, this minimum was unaffected by the nature of the substituent at position five of the pyrimidine. The cyclopentane ring showed great conformational flexibility. For X between - 9 0 and - 1 2 0 ° it adopted twist and envelope conformations in both the northern and southern hemispheres of the pseudo-rotational cycle. However, for X of around 60 °, i.e. with the pyrimidine in the s y n orientation, the cyclopentane preferred to adopt an envelope conformation with the pseudo-glycosidic carbon [C(9)] e x o or below the ring plane (equivalent to the I E conformation of furanoses). Interestingly, this is the conformation observed for the cyclopentane in the X-ray study, for the conformer with X = -117.7 °. By contrast, regardless of the orientation of the 5-fluorouracil, the furan ring of tegafur adopts an envelope conformation with the glycosidic carbon e n d o (IE).
Acknowledgements
Work at Vigo University was supported by Vicerrectorado de Investigaci6n of Vigo University. Work at the University of Santiago de Compostela was supported by the DGICYT (PM92-0090) and the Xunta de Galicia (XUGA20312B92 and 20306B95). References
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C. Ter6n et al./Journal of Molecular Structure 448 (1998) 69-75
[5] H. Maag, J.T. Nelson, J.L. Rios Steiner, E.J. Pfisbe, J. Med. Chem. 37 (1994) 431. and references cited therein. [6] L. Santana, M. Teijeira, E. Uriarte, J. Balzarini, E. De Clercq, Nucleosides and Nucleotides 14 (3-5) (1995) 521. [7] L. Santana, M. Teijeira, E. Uriarte, C. Ter~n, U. Casellato, R. Graziani, Nucleosides and Nucleotides 15 (6) (1996) 1179. [8] J.B. Rodrfguez, V. M~quez, M.C. Nicklans, H. Mitsuya, J.J. Barchi, J. Med. Chem. 37 (1994) 3389. [9] S. Neidle, L. Urpi, P. Serafinowsky, D. Whitby, Biochem. Biophys. Res. Commun. 161 (2) (1989) 910. [10] H.O. Kim, S.K. Ahn, A.J. Alves, J.W. Beach, L.S. Jeong, B.G. Choi, P. Van Roey, R.F. Schinazi, C.K. Chu, J. Med. Chem. 35 (1992) 1987. [11] H. Awano, S. Shuto, M. Baba, T. Kira, S. Shigeta, A. Matsuda, Bioorg. Med Chem. Lett. 4 (1994) 367. [12] CAD4-Express Software, Version 5.1, Enraf-Nonius, Delft, Netherlands, 1995.
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[ 13] A.L. Spek, HELENA. A Program for Data Reduction of CAD4 Data, University of Utrecht, Netherlands, 1997. [14] G.M. Sheldrick, Acta Crystallogr. Sect. A: 46 (1990) 467. [15] G.M. Sheldrick, SHELXL-97. Program for the Refinement of Crystal Structures, University of G6ttingen, Germany, 1997. [16] H.D. Flack, Acta Crystallogr. Sect. A: 39 (1985) 876. [17] International Tables for X-Ray Crystallography, Vol. C, Kluwer, Dordrecht, Netherlands, 1995. [18] L. Zsolnai, ZORTEP. A Program for the Presentation of Thermal Ellipsoids, University of Heidelberg, Germany, 1997. [19] M.J.S. Dewar, E.G. Zoebisch, E.F. Healy, J.J.P. Stewart, J. Am. Chem. Soc. 107 (1985) 3902. [20] BIOSYM Technologies, Inc., 10065 Barnes Canyon Road, San Diego, CA 92121. [21] Y. Nakai, K. Yamamoto, K. Terada, T. Uchida, N. Shimizu, S. Nishigaki, Chem. Pharm. Bull. 30 (1982) 2629.