- Email: [email protected]
Coordination behavior of sulfathiazole. Crystal structure of dichloro-disulfathiazole ethanol Cu(II) complex. Superoxide dismutase activity
Coordination Behavior of Sulfathiazole. Crystal Structure of DichloroDisulfathiazole Ethanol C&I) Complex. Superoxide Dismutase Activity J. Casanova, G. Alzuet, J. Borrh, J. Timoneda, S. Garcfa-Granda, and I. Chdano-Gonzilez JC, GA, JB. Departamento de Quimica Ino&nica, Facultad de Farmacia, Universidad de Valencia, Valencia, Spain.-JT. Departamento de Bioquimica, Facultad de Farmacia, Uniuersidad de Valencia, Valencia, Spain.-SG-G, IC-G. Departamento de Qulmica Fisica y Anal&a, Facultad de Q&mica, Universidad de Oviedo, Oviedo, Spain
ABSTRACT The crystaj structure of [Cu(sulfathiazole),(EtOH)C12] (triclinic; a = 7.82(1),-b = 11.441(3), c = 16.606(6) A, (Y= 109.68(4), p = 102.24(4), y = 91.27(4Y’, Z = 2, space group Pl) has been determined from the intensities of 2605 independent reflections (graphite monochromator MoKa; 4-circle diffractometer; R = 4.5%). The coordination polyhedron around Cu(I1) ion is intermediate between square pyramid and trigonal bipyramid. The spectroscopic data and the rhombic powder EPR spectrum are in agreement with the distorted geometry of the C&I) ion. The superoxide dismutase activity of the title compound has been measured and compared with those of the free drug and other sulfathiazole complexes.
INTRODUCTION 4-Amino-N-2-thiazolylbencenosulfonamide ically one of the most used sulfanilamide of bacterial infections in humans [l].
H2N-@Ihl<’
FIGURE 1.
(sulfathiazole = Hstz) (Fig. 1) is clinderivatives in the prevention and cure
SuIfathiaxole.
Address reprint requests and correspondence to: Dr. J. Borras Departamento de Quimica Inorganica, Facultad de Farmacia, Universidad de VaIencia, 46100-Bujassot, Valencia, Spain. Journal of Inorganic Biochemistry, 56,65-76 (1994) 0 1994 Ekvier Science Inc., 655 Avenue of the Americas, NY, NY 10010
65
0162-0134/94/$7.00
66
.I Casanova et al.
Copper complexes have found possible medical uses in the treatment of many diseases including cancer [2], [3]. It has been suggested that the anticancer activity of some copper complexes may be based on their ability to inhibit DNA synthesis or on the possible scavenging of superoxide anions [2]. Although the synthesis and characterization of sulfanilamide metal complexes have already been reported, it is often incomplete and conflicting [4]. In fact, the only crystal structures of the sulfathiazole complexes have been recently reported by us. The crystal structure of the Zn(stz),H,O complex shows the drug acting as a bridging ligand that interacts through the Nanilinoand the Nthiazole atoms, while in the [Cu(Hstz),(MeOH)Cl,] the drug behaves as a monodentate ligand binding to the metal ion via the Nanilinoatom [5-71. These results seem to suggest that the different behavior of such ligands depends on its deprotonation and/or the metal ion; thus it is difficult to give a generally valid account of the structures of other metal-sulfonamides. In this paper we report the preparation, properties, and crystal structure of the [Cu(Hstz),(EtOH)Cl,] complex and examine its superoxide radical scavenging activity and those of the previously reported Zn(stz),H,O and [Cu(Hstz),(MeOH)Cl,l complexes. The activities of these compounds are also compared with that of the parent drug.
EXPERIMENTAL Synthesis 0.76 Mmol of CuC1,.2H,O were added, with continuous stirring, to a hot mixture (60°C) of 15 ml of methanol and 75 ml of ethanol containing 1.57 mmol of Hstz; then, the solution was stirred for half an hour and left to stand at room temperature. Within one day, prismatic green crystals of [Cu(Hstz),(EtOH)Cl,] were obtained. Anal. Calc. for [Cu(Hstz),(EtOH)Cl,]: C, 34.7; N, 12.2; H, 3.5, Cu 9.2%. Found: C, 34.3; N, 12.0; H, 3.2, Cu 8.5%. Crystallographic
Data Collection and Refinement of the Structure
Crystal data and crystallographic data collection are shown in Table 1. Green crystal, 0.2 X 0.13 X 0.05 mm size. Throughout the experiment Moka radiation was used with a graphite crystal monochoromator on an Enraf-Nonius CAD4 single crystal diffractometer (h = 0.73071 A). The unit cell dimensions were determined from an angular setting of 25 reflections with 19 between 10” and 15”. The space group was determined from the structure determination. The intensity of 5139 reflections, in the hkl range (0, - 13, - 19) to (9, 13, 19) and 8 limits (0” < 8 < 25”) were measured using the (~28 scan technique and a variable scan rate with a primary scan time of 60 set per reflection. The intensity of the primary beam was checked throughout the data collection by monitoring three standard reflections every 60 min. The final drift correction factors were between 1.00 and 1.04. Profile analysis was performed on all reflections [8, 91. Semiempirical absorption correction was applied, using rcI_scans[lo], ~(MoKcu) = 13.41 cm-’ (corrections factors were in the range 0.92 to 1.00). Some double measured reflections were averaged, Rint = Z (I-(I))/& I = 0.056, resulting in 4753, of which 2605 were observed with I > 3~0). Lorentz and polarization corrections were applied and the data were reduced to IFol values. The structure
COORDINATION
BEHAVIOR
OF SULFATHIAZOLE
67
TABLE 1. Crystal Data and CrystallographicData Collection for the Cu(Hstz),(EtOH)Cl, Complex Formula Formula weight Crystal system Space group
a(A)
b (A) c (A) ; Y v (2) Crystal dimensions(mm) Z Dx(Mg me3) P No. observed,reflections No. variables F WOO) R wR
CuC,H,N&O,Cl, 691.14 Triclinic
pi
7.820) 11&l(3) 16.606(6) 109.68(4) 102.24(4) 91.27(4)” 1360.0(9) 0.2 x 0.13 x 0.05 2 1.69 13.41 cm-’ 2605 352 706 0.045 0.045
was solved by Direct Methods using the program SHELXS86 Ill]. Isotropic least squares refinements, using a local version of the program SHELX76 [121, converged to 0.98. At this stage additional empirical absorption correction was performed using DIFABS [13]. The maximum and minimum absorption correction factors were 0.80 and 1.21, respectively. All hydrogen atoms were geometrically placed and refined riding on their parent atoms. All nonhydrogen atoms were anisotropically refined, except the two carbon atoms of the ethanol ligand which were found disordered and refined as two rigid groups, 56/44%, with a common isotropic thermal parameter. The final conventional agreement factors were R = 0.045 and WR = 0.045 for the 2605 observed reflections and 352 variables. The function minimized was Zw(Fo - Fcj2, w = l/[ a2(Fo) + 0.00045 Fo2] with a(Fo) from counting statistics. The maximum shift to esd ratio in the last full-matrix least-squares cycle was less than 0.054 for parameters of the disordered molecule, and 0.004 for the rest. The final difference Fourier map showed no peaks higher than 0.44 eAm3 or deeper than -0.52 eAe3. Atomic scattering factors were taken from International Tables for X-Ray Crystallography [141. Geometrical calculations were made with PARST [15]. The molecular plot was made using the EUCLID package [16]. All calculations were made on a MicroVAX-3400 at the Scientific Computer Center of the University of Gviedo. Techniques Infrared spectrum was recorded on a Perkin-Elmer 843 instrument. The sample was prepared by using the KBr technique. Solid UV-VIS spectra was recorded with a Per-kin-Elmer Lambda 15 spectrophotometer.
68 J. Casanova et al.
Polycrystalline powder EPR spectrum of the [Cu(Hstz),(EtOH)Cl, I complex was registered at room temperature on a Bruker ER 200D spectrometer at the X-band frequency. Superoxide Dismutase
Assay
Superoxide dismutase activity of the metal complexes was determined according to Oberley and Spitz [17] with minor modifications. Xanthine (1.5 X lop4 M) and xanthine in 50 mM potassium phosphate buffer, pH 7.8, were used to generate a reproducible and constant flux of superoxide anions. An amount of xanthine oxidase giving in the control assays an A,,, rate of O.OS/min was used. Reduction of NBT (5.6 X 10e5 M) was used as an indicator of superoxide anion production and followed spectrophotometrically at 560 nm. The metal complexes were dissolved in 50 mM Tris-HCl buffer, pH 7.8, and added to the assay mixture in a volume representing one-tenth of the total. The percentage inhibition of NBT reduction was used as a measure of SOD activity of the complexes. Xanthine, xanthine oxidase, NBT, and superoxide dismutase (Bovine erytrocyte) were from Sigma Chemical Co. The inhibition of xanthine oxidase by the complexes was calculated measuring uric acid formation from xanthine at 310 nm in the presence of the complex concentrations used. The inhibition percentage of enzyme activity was subtracted from that of NBT reduction. RESULTS AND DISCUSSION
The final positional and equivalent isotropic thermal parameters are given in Table 2. Bond lengths and angles are listed in Table 3. The structure of the compound is shown in Figure 2 together with the atomic numbering. The stereochemistry of Cu(I1) is five coordinated with a CuN,Cl,O chromophore. The Cu(I1) ion is surroOunded by two Nanilino atoms frcm two Hstz ligands at a distance of 2.069(5) A, two Cl- anio9s at 2.261(2) A, and the 0 atom of the ethanol at the distance of 2.282(5) A. All bond lengths can be regarded as normal. The coordination polyhedron is intermediate between square pyramid and trigonal bipyramid [18]. The geometry of the molecule can be better described as a distorted tetragonal pyramid where the basal plane consists of tram nitrogen atoms from the sulfathiazole molecules and tram chloride ions. The. apical site is occupied by the ethanolic oxygen atom. The ligand atoms which form the basal pl$ne are not strictly coplanar. The Cl(l) and Cl(2) atoms lie 0.2700 and 0.3086 A below the baDsalplane of the pyramid and the N(1) and N(lA) atom: are 0.3842 and 0.3741 A above this plane. The copper atom is displaced 0.2204 A out of this same plane in the direction of the apical ethanol ligand. Due to the out-of-plane of the ethanol ligand, the Cu-O(3) line is not at 90” but rather at an angle of 8.2(2Y’ to the perpendicular to this plane. The value of T5 (T = mean in-plane Cu-L bond distance/mean out-of-plane Cu-L bond distance) is 0.949. The complex has a T value of 0.28 [T = ( p - a)/60], where p is the largest and (Y is the next largest metal-ligand bond angle, which in the complex correspond to N(l)-Cu-N(2), 171.0” and Cl-Cu-Cl, 153.9”, respectively]. The structural parameter r is applicable to five coordinate structures as an index of the degree of trigonality; for perfect tetragonal-pyramidal and
COORDINATION
BEHAVIOR OF SULFATHIAZOLE
TABLE 2. Fractional Positional and Thermal Parameters (1/3)ZiZjUijaiaja~a~ Atom Cdl) Cl(l) s(1) Wa) Cl(2) s(2) s(2a) N(l) o(3)
NOa) N(3a) N(3) OCW OOa) o(1) N(2) c(1) N(2a) o(2) Ula) C(4) cm C(4a) c(5) C(7a) c(6) C(7) C(3a) C(6a) c(5a) c(3) c(2) c(8) c(8a) c(9) CX9a) c(10) Cm c(lO’)
(Xl’) #Occupation
Z
0.3136 (9) 0.4067 (9) 0.4065 (9) 0.6647 (9) 0.2923 (9) 0.9313 (8) 0.1595 (9) 0.4641(9) 0.5840 (9) 0.3932 (9) 0.5692 (9) 0.389 (1) 0.256 (1) OF357 (1) 0.970 (1) 0.369 (1) 0.944 (1) -0.348 (4) -0.419 (4)
0.74126(7) 0.5464 (2) 0.7180 (2) 0.7823 (2) 0.9479 (2) 0.8464 (2) 0.6528 (2) 0.7694 (5) 0.6749 (5) 0.7135 (5) 0.8811(5) 0.6189 (5) 0.6610 (4) 0.8651(5) 0.6379 (5) 0.6368 (5) 0.7598 (6) 0.8629 (5) 0.8413 (5) 0.7288 (6) 0.7368 (6) 0.6256 (6) 0.7594 (6) 0.6335 (6) 0.8101(6) 0.6452 (6) 0.6898 (6) 0.6419 (6) 0.8476 (6) 0.8634 (6) 0.8514 (6) 0.8635 (6) 0.6850 (8) 0.8146 (8) 0.8071(9) 0.6924 (8) 0.733 (3) 0.797 (2)
0.50738(5) 0.4418 (1) 0.1977 (1) 0.8082 (1) 0.5773 (1) 0.0482 (2) 0.9602 (1) 0.3891(3) 0.4872 (4) 0.6266 (3) 1.0567 (4) - 0.0495 (4) 0.7933 (3) 0.7728 (3) 0.2322 (3) 0.0954 (3) 0.3370 (4) 0.9116 (3) 0.2103 (3) 0.6762 (4) 0.2461(4) 0.6690 (4) 0.7615 (4) 0.2310 (4) 0.9719 (4) 0.2758 (4) 0.0356 (5) 0.7122 (4) 0.7279 (4) 0.7704 (4) 0.3049 (4) 0.3504 (5) - 0.1060 (5) 1.1146 (5) - 0.0642 (6) 1.0732 (6) 0.469 (2) 0.544 (2)
-0.318 -0.369
0.757 (3) 0.797 (3)
0.544 (2) 0.480 (2)
0.0944 (1) 0.1701(3) 0.5897 (2) 0.8926 (2) 0.1506 (3) 0.4526 (3) 0.9110 (3) 0.0144 (7) - 0.1940 (7) 0.1335 (7) 0.9612 (7) 0.4068 (7) 0.9552 (6) 0.9687 (6) 0.7005 (7) 0.5167 (7) 0.1442 (8) 0.9206 (7) 0.6638 (7)
(5) (4)
factor, 0.56(2); “Occupation
trigonal-bipyramidal
WA.
Y
X
(with es&).
69
Ueq =
Ueq ( * 100) 2.80 (3) 4.25 (7) 4.02 (7) 3.67 (7) 5.47 (8) 5.8 (1) 5.53 (9) 3.0 (2) 5.5 (2) 3.0 (2) 3.8 (2) 3.8 (2) 5.1 (2) 4.9 (2) 5.8 (2) 3.7 (2) 2.9 (2) 3.6 (2) 6.1 (2) 3.1 3.0 3.5 3.1 3.4 3.3 3.3 3.3 3.4 3.2 3.2 4.0 4.1 5.2 5.0 6.0 5.7 11.3 11.3
(3) (3) (3) (3) (3) (3) (3) (3) (3) (3) (3) (3) (3) (4) (3) (4) (4) (5)X (5)”
11.3 (5)“X 11.3 (5P
factor, O&l(2).
geometries the value of T is zero and unity, respectively
Comparison with the T value of 0.36 in the structurally related methanol complex [61 suggests that the ethanol compound lies closer to the ideal&d square pyramidal geometry. Possible electronic spectroscopic implications of the more square pyramidal character are considered below. In general, the geometry of both methanol and ethanol complexes is comparable. The major difference in the stereochemistry of the CuN2Cl,0 chromophore
70 J. Casanova et al.
TABLE 3. Bond Lengths (A) (with esd’s) CumCl(l) C&)-O(3) W-W) S(la)-0(2a) S(laHX4a) WaXt7a) O(3)-Cc101 N(la)_C(la) N(3)-C(7) C(lM6) C(laX(2a) C(4M3) C(4a)_Wa) C(3M2)
2.281(2) 2.282(S) 1.606(5) l&1(5) 1.762(7) 1.744(7) 1.41(3) l&7(8) 1.342(8) 1.389(8) 1.3&l(8) 1.376(9) 1.401(8) 1.394(9)
cumCl(2) Cu(l)-N(la) W-O(2) S(la)-O(la) S(2)-c(7) S(2aM9a) O(3)-C(10’) N(3aM7a) N(3)-c(8) CWc(2) CXlaX(6a) C(2a)-C(3a) CWCt6) C(Wx9)
2.241(2) 2.071(5) 1.444(5) 1.448(5) 1.740(7) 1.734(8) 1.60 (3) 1.334(8) 1.393(8) 1.383(g) 1.386(g) 1.397(9) 1.384(9) 1.33 (1)
Cu(l)-N(1) W-O(l) S(lxx4) S(la)-N(2a) S(2Mx9) N(l)-C(1) O(3)-C(11’) N(3aM8a) N(2)-C(7) N(2a)-C(7a) C(4Mx5) C(4aM3a) C(6aKX5a) C(8aM9a)
Bond Angles (“) (with esd’s) c1(2)-Cu(l)-cl(1) N(l)-Cu(l)-Cl(2) O(3)-Cu( l)-Cl(2) N(la)-CuWClW N(la)_Cu(l)-N(1) N(2)-S(l)-00) 0(2M(l)-N(2) CW-S(l)-N(2) O(la)-S(labO(2a) N(2a)-S(la)-O(la) CX4a)-S(la)-O(la) cx9)-s(2Mx7) c(l)-N(l)-Cu(1) cx10’)-0(3)-cu(1) c(ll’)-o(32cu(l) C(11 ‘)-o(3)-C(10’) C@a)-N(3a)-C(7a) c(7)-N(2)-SW C(2)-C(l)-NW Cf7a)-N(2a)-S(la) Cf6a)-C(la)-N(la) c(5)-c(4)_S(l) cx3Mx4M5) CX3a)-C(4aHla) C(5a)-C(4a)-C(3a) N(3a)_C(7a)-S(2a) N(2a)-C(7a)-N(3a) NW-C(7H2) N(2)-C(7)-N(3) C(Sa)-CX6aMla) Ct2)-Cx3m4) C(9)-C(8)-N(3) c(8)-C(9)-S(2) CW-cxlO)-O(3)
153.9(l) 89.9(2) 111.8(2) 90.7(2) 171.0(2) 104.3(3) 111.7(3) 106.8(3) 117.8(3) 104.3(3) 106.8(3) 90.7(4) 116.7(4) 122.0) 117.0) 40X2) 114.7(6) 121.9(5) 120.1(6) 122.3(5) 119.4(5) 119.5(5) 120.7(6) 120.3(5) 120.4(6) 110.0(5) 119.9(6) 109.8(5) 120.1(6) 119.5(6) 119.8(6) 113.0(7) 1X6(6) 115.(2)
N(l)-Cu(l)-Cl(l) o(3)_cu(l)-cl(1) 0(3)-0.1(1)-N(l) N(la)-Cu(GCl(2) N(la)-Cu(l)-O(3) ocWWo(1) C(4)-SW-O(l) C(4)-W-O(2) N(2a)-S( la)-O(2a) C(4a)-S(la)-O(2a) C(4a)-S(la)-N(2a) C(9a)-S(2a)-C(7a) c(10)-0(3)-cu(1) c(10’)-0(3)-q10) c(11’)-0(3)-q10) C(la)-N(la)-Cu(1) C(8)-N(3)-C(7) C(6)-C(l)-N(1) C(2)-C(l)-C(6) C(2a)-C(la)-N(la) C(6akC(labC(2a) C(3)-C(4)-S(1) C(3a)-C(2a)-C(la) C(Sa)-C(4aMla) C(6)-C(5)-C(4) N(2a)-C(7a)-S(2a) C(5M6HXl) N(2)-C(7)-S(2) C(4a)-C(3aX(2a) C(6a)-C(5a)-C(4a) c(3)-c(2)-cx1) C(9a)_C@a)-N(3a) C@a)-C(9a)-S(2a) c(10%00)-0(3)
92.9(l) 94.3(2) 85.4(2) 90.5(2) 86.2(2) 118.7(3) 107.7(3) 107.1(3) 112.5(3) 107.6(3) 107.4(3) 90.6(4) 130. (1) 45.(l) 15.(2) 116.6(4) 114.8(6) 119.6(6) 120.3(6) 119X(6) 120.8(6) 119xX5) 119.5(6) 119.2(5) 119.8(6) 130.1(5) 119.8(6) 130.1(5) 120.0(6) 119.8(6) 119.7(6) 112.6(7) 112.0(6) 76.0(3)
2.066(5) 1.448(5) 1.768(6) 1.615(5) 1.737(8) 1.450(g) 1.99 (3) l&5(8) 1.328(8) 1.323(8) 1.385(9) 1.370(9) 1.385(9) 1.32 (1)
COORDINATION
FIGURE 2.
BEHAVIOR
OF SULFATHIAZOLE
71
ORTEP drawing of the [Cu(Hst.&(EtOH)C1J.
is the O(3)-Cu-Cl(l) bond angle which may be related to a different hydrogen bond and to the larger size of the ethanol molecule. Both complexes represent a series of distortion isomers whose geometries reflect the flexible stereochemistry of the C&I) ion or “plasticity effect,” the apical group being the main cause for the structural differences. I&and Conformation
and Molecular Packing
There are no unusual bond lengths or angles in the coordinated Hstz ligand, %xcept for the NW-C(l) bond di$ance that it is significantly increased [1.450@) A in the complex and 1.401(4) A in the sulfathiazole polymorph II [2OU.This fact, also found in the methanol complex is due to the interaction of the N(1) atom with the metal ion. The study of the W-N(2), N(2)_CX7), and c(7)_N(3), distances suggests that, as it occurs in the [Cu(Hstz&&kOH)c1,1 compound, the sulfathiazole presents the imido form. Neighboring molecules are interconnected by a hydrogen bonding system involving the nitrogen atoms, the chloride anions, and the sulfonamide and
72 J. Casanova et al.
ethanolic oxygen atoms. Relevant hydrogen bonding distances are given in Table 4. Figure 3 shows a perspective view of the detail of the molecular packing showing the N-H.. . H interbonding. Spectroscopic Data IR and Electronic Spectra. The IR spectrum shows the characteristic bands of the Hstz ligand. The bands of 3300 and 3240 cm-‘, assigned to the v,(N-H) and I.&N-H) vibrations of the NH, group, are shifted respect to those of the ligand (3320 and 3280 cm-‘). As expected, the bands due to Y(SO&~ ,(1320 cm-‘), v(SO,),(1140 cm-‘), scissors and wagging SO2 (570 and 550 cm-‘) vibrations remain unchanged. There is no change of the bands at 1540, 920, and 680-640 cm-‘, which have been assigned to the characteristic thiaxole ring and v(S-N) and v(C-S) vibrations, respectively. The presence of coordinated alcohol is indicated by a strong broad band at 3550-3400 cm-’ [211. The new band at 1060 cm-’ is attributed to bending M-OH vibrations [22]. The d-d electronic spectrum of the complex consists of an asymmetric broad band with a maximum approximately at 14140 cm-‘. In order to reveal the correlation of the d orbitals, the solid state visible spectrum was deconvoluted into Gaussian component bands by curve-fitting iteration processes. Three Gaussian component bands (15730, 14470, and 12050 cm-‘) with that of the highest frequency having the greatest intensity were resolved. On the basis of
TABLE 4. Distances and Angles for H-Bonds (with esd’s) Donor-H N(l)_Hll 1.08oc3) NWH12 l.ow7) 0(3)_H31 0.97(l) N(la&Hlla l.ow7) N(la)_Hlla 1.08oc7) N(la)_HUa 1.08oc7~ N(la)-H12a 1.080(7) NW-H12 l.o800 N(la>Hlla 1.W7)
a3Mr31 0.97(l) No-H21 l.Oso@)
Donor..Acceptor
H..Acceptor
N(l)..CI (2) 3.045w N(l)..0 (3) 2.95(l) 0(3)..N (la) 2.977(7) N(la)..Q (1) 3.099(6) N(laI.0 (3) 2.977(7) N(laLCl(2) 3.065(7) N(lak.0 (la) (1) 3.025(7) NW...0 (1) (1) 3.093(7) N(la)..Cl (1) (2) 3.46X6)
Hll...Cl(2) 2.687(5) H12...0 (3) 2.58(l) H3l...N (la) 2.648(7) Hlla..Cl(l) 2.8320 Hlla...O (3) 2.590@) H12a...Cl(2) 2.786(7) HEa... (la) (1) 2.086(S) H12...0 (1) (1) 2.151(7) Hlla...Cl(l) (2) 2.405(6) H31...Cl(l) (2) 2.1940) H21...N (3) (3) 1.848@)
o(3x.a (1) (2) 3.124X8) N(2h.N (3) (3) 2.875(8)
symmetry codes: (1) -X - 1, +Y, +z; (2) -x,
-y
+ I, --z + 1; (3)
Donor..Acceptor N(l)_Hll...Cl(2) 98.8(5) N(l)_H12...0 (3) 99.5(5) y(&fI$...N (la) N(l&Hlla...Cl(l) 93.90 N(la)_Hlla...O (3) 100.2(5) N(la)_HKIa...Cl(2) 94.4(5) N(la>HUa...O (la) (1) 143.7(6) N(lkH12...0 (1) (1) 144.X6) N(la)_Hlla...Cl (1) (2) 16&X.3(6) cm&y
...Cl (1) (2)
N(2iH21...N 157.3(7) -X
+
I, -y
+
(3) (3)
I, -x.
COORDINATION
BEHAVIOR
OF SULFATHIAZOLE
73
FIGURE 3. Perspective view of a detail of the molecular packing, showing the N-H.. . N interactions.
the d orbital splitting, the orbital sequence of the title complex is intermediate between those of trigonal bipyramidal and square pyramidal symmetry [23, 241 A comparison of this spectrum with that of the methanol complex does not give evidence to establish if the basic metal geometry in the title compound leads to a more square pyramid, such as has been indicated by the structural parameter 7. EPR Spectrum of the [Cu(Hst.d2(EtOH)Cl,] Complex The polycrystalline EPR spectrum is rhombic (Fig. 4). The calculated EPR parameters are g, = 2.05, g, = 2.09, and g, = 2.28. Fo r systems with g, >g, >g, the ratio R=(g,gr)& - g2) indicates the ground state of the complex [251. If R > 1 the ground state is predominantly d,z while the ground state is predominantly d,z_,,z when the value of R is less than one. The complex presents an R value of 0.17 indicating d,z_y2 as the ground state. Thus the complex may be five coordinated square pyramid, which has been confirmed by its crystal structure. The calculated R value of 0.5 for the [Cu(Hstz),(MeOH)Cl,] complex is higher than that for the ethanol compound, suggesting a more trigonal geometry for the methanol complex as it also indicated by its T value. Superoxide Dismutation Activity The superoxide dismutase activity of the drug and the [Cu(Hstz),(EtOH)cl,], [Cu(Hstz),(MeOH)Cl,], and Zn(stz),.H,O complexes were assayed by their
74
J. Casanova et al.
FIGURE 4.
Polycrystalline powder EPR spectrum of the Cu(Hstzl,(EtOH)Cl~ cmpkx.
ability to inhibit the reduction of nitroblue tetrazolium [26]. The superoxide scavenging data (Fig. 5) indicates that the Zn(I1) complex is only marginally more active than the free drug. The copper complexes exhibit a greatly increased superoxide dismutase activity compared with the parent drug molecule. Table 5 shows the chelate or superoxide dismutase concentrations required to produce 50% inhibition of NE3T reduction (IC,,). On a molar basis the methanol
0
5
IO
15
20
Concentration (PM) 5. Inhibition of. NBT reduction in the presence of metal complexes 0- 0 Cu(Hstz),(MeOHlCl,, 0-0 Cu(Hstz),(EtOH)Cl,, q - 0 Zn(Stz),H,O, n - n Hstz. Each point repreknts the mean f standard deviation of triplicate determinations.
FIGURE
COORDINATION
BEHAVIOR
OF SULFATHIAZOLE
75
TABLE 5. Superoxide Dismutase-Mimetic Activity IC,,a ( PM) Cu(Hstz)z(MeOH)cl z Cu(Hstz),(EtOH)Cl, Suoeroxidedismutase
2.510 5.170 0.006
- Log Ic,o 5.60 5.28 8.21
‘IC,, is defined as the concentration of complex or enzyme which produces 50% inhibition of NBT reduction. A molecular weight of 31,200 was considered for calculating the enzyme concentration.
coordinated complex is more efficient as superoxide radical scavenger than the ethanol complex being the former, 400, and the latter, 800 times less effective than superoxide dismutase. For the zinc complex a proper IC,, could not be established. The values presented here are similar to the ones published by Sorenson et al. [27] and Bijloo et al. [28] for the enzyme and other copper complexes. The mechanism believed to be operating in the naturally occurring superoxide dismutases involves one-electron reduction of the metal ion of the active center by superoxide followed by reoxidation of the reduced metal ion by a second superoxide anion. Metal complexes that can undergo such redox cycling are likely to function as superoxide scavengers. It is assumed that electron transfer between the metal center and superoxide anion radicals occurs by direct binding [29]. A fast exchange of coordinated EtOH or MeOH and limited steric hindrance to the approach of the superoxide anion are considered essential requirements for the successful binding of the 0; radical [30]. The high superoxide dismutase activity could also be explained considering a favorable response of r-electrons of the aromatic side chain in stabilizing the 01-o; interactions. Moreover, as suggested by Srivastava et al. [31], the distorted geometry of these complexes may favor the geometrical change, which is essential for the catalysis as the geometry of copper in SOD enzyme changes from distorted squared pyramidal (for C&I)) to distorted tetrahedral (for C&I)) during the catalysis. We consider that these requirements are satisfied in the distorted complexes [Cu(Hstz),(EtOH)Cl,] and [Cu(Hstz),(MeOH)Cl,] and that the doubled superoxide dismutase-mimetic activity of the latter can be related with its higher geometrical distortion. Supplementary
Material
Anisotropic temperature factors, a list of observed and calculated structure factors, fractional positional parameters of the hydrogen atoms, torsion angles and angles between least-square planes and lines are available from the authors on request. JC, GA, and JB appreciate financial suppoti from FAR91 -197(CICYT). JC acknowledges the Generalitat Valenciana for a grant. JT appreciates FIS grant No. 92 /26X
REFERENCES 1.
A. Bult, in Metal Ions in Biological Systems, H. Sigel, Ed., Marcel Dekker, New York 1983.
16
J. Casanova
et al.
2. J. R. J. Sorenson, Chem. in Brit. 16, 1110 (1984). 3. R. K. Gouch, T. W. Kensler, L. W. Oberley, and J. R. J. Sorenson, in Biochemical and Inorganic Copper Chemistry, K. D. Karlin and J. Zubieta, Eds., Adenine, New York, 1986, Vol. 7. 4. A. Buh, Pharm. Weekblad Sci. Ed. 3,213 (1981). 5. J. Casanova, G. AIzuet, S. Ferrer, J. Borras, S. Garcia-Granda, and E. Perez-Carrefio, J. Inotg. B&hem., 51, 689 (1993). 6. J. Casanova, G. AIzuet, J. Borrh, J. M. Amigo, and T. Debaerdemaeker, Zeit. jiir fit., 209, 271 (1994). 7. J. Casanova, G. Aizuet, J. BorrL, L. David, and D. Gatteschi, Inorg. Chim. Acta, 211, 183 (1993). 8. M. S. Lehman and F. K. Larsen, Acta Cryst. A30, 580 (1974). 9. D. F. Grant and E. J. Gabe, J. Appl. Cryst. 11, 114 (1978). 10. A. C. T. North, D. C. Phillips, and F. S. Mathews, Acta Cryst. A24, 351 (1968). 11. G. M. Sheldrick, in Crystallographic Computing 3, edited by G. M. Sheldrick, C. Kruger, and R. Goddard, Eds., &ford, Ciarendon Press, 1985, p. 175. 12. G. M. Sheldrick, SHELX76. Program for crystal structure determination, Univ. of Cambridge, England, 1976. 13. N. Walker and D. Stuart, Acta Cryst. A39,158 (1983). 14. International Tables for X-Ray Crystallography, Byrmingham, Kynoch Press (Present distributor KIuwer Acad. Publ., Dordrecht), 1974, Vol. IV. 15. M. Nardelh, Comp. Chem. 7,95 (1983). 16. A. L. Spek, in Computational Crystallography, edited by SAYRE, Gxford, Clarendon Press, 1982, p. 528. 17. L. W. Oberley and D. R.‘Spitz, in Handbook of Methods for Gqgen Radicals Research, R. A. Greenwald, Ed., CRC Press, Boca Raton, 1986, pp. 217-220. 18. B. J. Hathaway, in Comprehensive Coordination Chemistry, Eds. G. Wilkinson, R. D. Gillard, and J. A. Cleverty, Eds., Pergamon Press, New York, 1987, Vol. 5. 19. W. Addison, T. N. Rao, J. Reedijk, J. van Rijn, and G. C. Verschoor, J. Chem. Sot., Dalton Trans., 1349 (1984). 20. G. J. Kruger and G. Gafner, Acta Ctyst., B27, 326 (1971). 21. S. S. Tandon, L. K. Thompson, and R. C. Hynes, Znotg. Chem. 31, 2210 (1992). 22. K. Nakamoto, Infired and Raman Spectra of Inorganic and Coordination Compounds, John Wiley & Sons, New York, 1986,4th edn. 23. B. J. Hathaway and D. E. Billing, Coord. Chem. Rev. 5, 143 (1970). 24. (a) A. Bencini and D. Gatteschi, Trans. Met. Chem. 8, 2 (1980); (b) A. Bencini, I. Bertini, D. Gatteschi, and A. Scozzafava, Inorg. Chem. 17, 3194 (1978). 25. M. F. El-ShazIy, A. El-Dissowky, T. Salem, and M. Osman, Znorg. Chim. Acta 40, 1 (1980). 26. A. Bury, A. E. Underhill, D. R. Kemp, N. J. O’Shea, J. P. Smith, and P. S. Gomm, Znorg. Chim. Acta 13885 (1987). 27. H. H. A. DoIIwet, J. B. McNicholas, A. Pezeshk, and J. R. J. Sorenson, Trace Elements in Medicine 4, 13 (1987). 28. G. J. Bijloo, H. van der Goot, A. Bast, and H. Timmerman, J. Znotg. Biochem. 40, 237 (1990). 29. J. A. Fee, in Metal Ions in Biological Systems, H. Sigel, Ed., Marcel Dekker, New York, 1981, Vol. 13. 30. M. Boden, M. C. Holmes, and P. F. Knowles, Biochem. Biophys. Res. Commun. 57, 845 (1974). 31. R. G. Bhirud and T. S. Srivastava, Znorg. Chim. Acta 179, 125 (1991). Received July 13, 1993; accepted September 15, 1993
ABSTRACT The crystaj structure of [Cu(sulfathiazole),(EtOH)C12] (triclinic; a = 7.82(1),-b = 11.441(3), c = 16.606(6) A, (Y= 109.68(4), p = 102.24(4), y = 91.27(4Y’, Z = 2, space group Pl) has been determined from the intensities of 2605 independent reflections (graphite monochromator MoKa; 4-circle diffractometer; R = 4.5%). The coordination polyhedron around Cu(I1) ion is intermediate between square pyramid and trigonal bipyramid. The spectroscopic data and the rhombic powder EPR spectrum are in agreement with the distorted geometry of the C&I) ion. The superoxide dismutase activity of the title compound has been measured and compared with those of the free drug and other sulfathiazole complexes.
INTRODUCTION 4-Amino-N-2-thiazolylbencenosulfonamide ically one of the most used sulfanilamide of bacterial infections in humans [l].
H2N-@Ihl<’
FIGURE 1.
(sulfathiazole = Hstz) (Fig. 1) is clinderivatives in the prevention and cure
SuIfathiaxole.
Address reprint requests and correspondence to: Dr. J. Borras Departamento de Quimica Inorganica, Facultad de Farmacia, Universidad de VaIencia, 46100-Bujassot, Valencia, Spain. Journal of Inorganic Biochemistry, 56,65-76 (1994) 0 1994 Ekvier Science Inc., 655 Avenue of the Americas, NY, NY 10010
65
0162-0134/94/$7.00
66
.I Casanova et al.
Copper complexes have found possible medical uses in the treatment of many diseases including cancer [2], [3]. It has been suggested that the anticancer activity of some copper complexes may be based on their ability to inhibit DNA synthesis or on the possible scavenging of superoxide anions [2]. Although the synthesis and characterization of sulfanilamide metal complexes have already been reported, it is often incomplete and conflicting [4]. In fact, the only crystal structures of the sulfathiazole complexes have been recently reported by us. The crystal structure of the Zn(stz),H,O complex shows the drug acting as a bridging ligand that interacts through the Nanilinoand the Nthiazole atoms, while in the [Cu(Hstz),(MeOH)Cl,] the drug behaves as a monodentate ligand binding to the metal ion via the Nanilinoatom [5-71. These results seem to suggest that the different behavior of such ligands depends on its deprotonation and/or the metal ion; thus it is difficult to give a generally valid account of the structures of other metal-sulfonamides. In this paper we report the preparation, properties, and crystal structure of the [Cu(Hstz),(EtOH)Cl,] complex and examine its superoxide radical scavenging activity and those of the previously reported Zn(stz),H,O and [Cu(Hstz),(MeOH)Cl,l complexes. The activities of these compounds are also compared with that of the parent drug.
EXPERIMENTAL Synthesis 0.76 Mmol of CuC1,.2H,O were added, with continuous stirring, to a hot mixture (60°C) of 15 ml of methanol and 75 ml of ethanol containing 1.57 mmol of Hstz; then, the solution was stirred for half an hour and left to stand at room temperature. Within one day, prismatic green crystals of [Cu(Hstz),(EtOH)Cl,] were obtained. Anal. Calc. for [Cu(Hstz),(EtOH)Cl,]: C, 34.7; N, 12.2; H, 3.5, Cu 9.2%. Found: C, 34.3; N, 12.0; H, 3.2, Cu 8.5%. Crystallographic
Data Collection and Refinement of the Structure
Crystal data and crystallographic data collection are shown in Table 1. Green crystal, 0.2 X 0.13 X 0.05 mm size. Throughout the experiment Moka radiation was used with a graphite crystal monochoromator on an Enraf-Nonius CAD4 single crystal diffractometer (h = 0.73071 A). The unit cell dimensions were determined from an angular setting of 25 reflections with 19 between 10” and 15”. The space group was determined from the structure determination. The intensity of 5139 reflections, in the hkl range (0, - 13, - 19) to (9, 13, 19) and 8 limits (0” < 8 < 25”) were measured using the (~28 scan technique and a variable scan rate with a primary scan time of 60 set per reflection. The intensity of the primary beam was checked throughout the data collection by monitoring three standard reflections every 60 min. The final drift correction factors were between 1.00 and 1.04. Profile analysis was performed on all reflections [8, 91. Semiempirical absorption correction was applied, using rcI_scans[lo], ~(MoKcu) = 13.41 cm-’ (corrections factors were in the range 0.92 to 1.00). Some double measured reflections were averaged, Rint = Z (I-(I))/& I = 0.056, resulting in 4753, of which 2605 were observed with I > 3~0). Lorentz and polarization corrections were applied and the data were reduced to IFol values. The structure
COORDINATION
BEHAVIOR
OF SULFATHIAZOLE
67
TABLE 1. Crystal Data and CrystallographicData Collection for the Cu(Hstz),(EtOH)Cl, Complex Formula Formula weight Crystal system Space group
a(A)
b (A) c (A) ; Y v (2) Crystal dimensions(mm) Z Dx(Mg me3) P No. observed,reflections No. variables F WOO) R wR
CuC,H,N&O,Cl, 691.14 Triclinic
pi
7.820) 11&l(3) 16.606(6) 109.68(4) 102.24(4) 91.27(4)” 1360.0(9) 0.2 x 0.13 x 0.05 2 1.69 13.41 cm-’ 2605 352 706 0.045 0.045
was solved by Direct Methods using the program SHELXS86 Ill]. Isotropic least squares refinements, using a local version of the program SHELX76 [121, converged to 0.98. At this stage additional empirical absorption correction was performed using DIFABS [13]. The maximum and minimum absorption correction factors were 0.80 and 1.21, respectively. All hydrogen atoms were geometrically placed and refined riding on their parent atoms. All nonhydrogen atoms were anisotropically refined, except the two carbon atoms of the ethanol ligand which were found disordered and refined as two rigid groups, 56/44%, with a common isotropic thermal parameter. The final conventional agreement factors were R = 0.045 and WR = 0.045 for the 2605 observed reflections and 352 variables. The function minimized was Zw(Fo - Fcj2, w = l/[ a2(Fo) + 0.00045 Fo2] with a(Fo) from counting statistics. The maximum shift to esd ratio in the last full-matrix least-squares cycle was less than 0.054 for parameters of the disordered molecule, and 0.004 for the rest. The final difference Fourier map showed no peaks higher than 0.44 eAm3 or deeper than -0.52 eAe3. Atomic scattering factors were taken from International Tables for X-Ray Crystallography [141. Geometrical calculations were made with PARST [15]. The molecular plot was made using the EUCLID package [16]. All calculations were made on a MicroVAX-3400 at the Scientific Computer Center of the University of Gviedo. Techniques Infrared spectrum was recorded on a Perkin-Elmer 843 instrument. The sample was prepared by using the KBr technique. Solid UV-VIS spectra was recorded with a Per-kin-Elmer Lambda 15 spectrophotometer.
68 J. Casanova et al.
Polycrystalline powder EPR spectrum of the [Cu(Hstz),(EtOH)Cl, I complex was registered at room temperature on a Bruker ER 200D spectrometer at the X-band frequency. Superoxide Dismutase
Assay
Superoxide dismutase activity of the metal complexes was determined according to Oberley and Spitz [17] with minor modifications. Xanthine (1.5 X lop4 M) and xanthine in 50 mM potassium phosphate buffer, pH 7.8, were used to generate a reproducible and constant flux of superoxide anions. An amount of xanthine oxidase giving in the control assays an A,,, rate of O.OS/min was used. Reduction of NBT (5.6 X 10e5 M) was used as an indicator of superoxide anion production and followed spectrophotometrically at 560 nm. The metal complexes were dissolved in 50 mM Tris-HCl buffer, pH 7.8, and added to the assay mixture in a volume representing one-tenth of the total. The percentage inhibition of NBT reduction was used as a measure of SOD activity of the complexes. Xanthine, xanthine oxidase, NBT, and superoxide dismutase (Bovine erytrocyte) were from Sigma Chemical Co. The inhibition of xanthine oxidase by the complexes was calculated measuring uric acid formation from xanthine at 310 nm in the presence of the complex concentrations used. The inhibition percentage of enzyme activity was subtracted from that of NBT reduction. RESULTS AND DISCUSSION
The final positional and equivalent isotropic thermal parameters are given in Table 2. Bond lengths and angles are listed in Table 3. The structure of the compound is shown in Figure 2 together with the atomic numbering. The stereochemistry of Cu(I1) is five coordinated with a CuN,Cl,O chromophore. The Cu(I1) ion is surroOunded by two Nanilino atoms frcm two Hstz ligands at a distance of 2.069(5) A, two Cl- anio9s at 2.261(2) A, and the 0 atom of the ethanol at the distance of 2.282(5) A. All bond lengths can be regarded as normal. The coordination polyhedron is intermediate between square pyramid and trigonal bipyramid [18]. The geometry of the molecule can be better described as a distorted tetragonal pyramid where the basal plane consists of tram nitrogen atoms from the sulfathiazole molecules and tram chloride ions. The. apical site is occupied by the ethanolic oxygen atom. The ligand atoms which form the basal pl$ne are not strictly coplanar. The Cl(l) and Cl(2) atoms lie 0.2700 and 0.3086 A below the baDsalplane of the pyramid and the N(1) and N(lA) atom: are 0.3842 and 0.3741 A above this plane. The copper atom is displaced 0.2204 A out of this same plane in the direction of the apical ethanol ligand. Due to the out-of-plane of the ethanol ligand, the Cu-O(3) line is not at 90” but rather at an angle of 8.2(2Y’ to the perpendicular to this plane. The value of T5 (T = mean in-plane Cu-L bond distance/mean out-of-plane Cu-L bond distance) is 0.949. The complex has a T value of 0.28 [T = ( p - a)/60], where p is the largest and (Y is the next largest metal-ligand bond angle, which in the complex correspond to N(l)-Cu-N(2), 171.0” and Cl-Cu-Cl, 153.9”, respectively]. The structural parameter r is applicable to five coordinate structures as an index of the degree of trigonality; for perfect tetragonal-pyramidal and
COORDINATION
BEHAVIOR OF SULFATHIAZOLE
TABLE 2. Fractional Positional and Thermal Parameters (1/3)ZiZjUijaiaja~a~ Atom Cdl) Cl(l) s(1) Wa) Cl(2) s(2) s(2a) N(l) o(3)
NOa) N(3a) N(3) OCW OOa) o(1) N(2) c(1) N(2a) o(2) Ula) C(4) cm C(4a) c(5) C(7a) c(6) C(7) C(3a) C(6a) c(5a) c(3) c(2) c(8) c(8a) c(9) CX9a) c(10) Cm c(lO’)
(Xl’) #Occupation
Z
0.3136 (9) 0.4067 (9) 0.4065 (9) 0.6647 (9) 0.2923 (9) 0.9313 (8) 0.1595 (9) 0.4641(9) 0.5840 (9) 0.3932 (9) 0.5692 (9) 0.389 (1) 0.256 (1) OF357 (1) 0.970 (1) 0.369 (1) 0.944 (1) -0.348 (4) -0.419 (4)
0.74126(7) 0.5464 (2) 0.7180 (2) 0.7823 (2) 0.9479 (2) 0.8464 (2) 0.6528 (2) 0.7694 (5) 0.6749 (5) 0.7135 (5) 0.8811(5) 0.6189 (5) 0.6610 (4) 0.8651(5) 0.6379 (5) 0.6368 (5) 0.7598 (6) 0.8629 (5) 0.8413 (5) 0.7288 (6) 0.7368 (6) 0.6256 (6) 0.7594 (6) 0.6335 (6) 0.8101(6) 0.6452 (6) 0.6898 (6) 0.6419 (6) 0.8476 (6) 0.8634 (6) 0.8514 (6) 0.8635 (6) 0.6850 (8) 0.8146 (8) 0.8071(9) 0.6924 (8) 0.733 (3) 0.797 (2)
0.50738(5) 0.4418 (1) 0.1977 (1) 0.8082 (1) 0.5773 (1) 0.0482 (2) 0.9602 (1) 0.3891(3) 0.4872 (4) 0.6266 (3) 1.0567 (4) - 0.0495 (4) 0.7933 (3) 0.7728 (3) 0.2322 (3) 0.0954 (3) 0.3370 (4) 0.9116 (3) 0.2103 (3) 0.6762 (4) 0.2461(4) 0.6690 (4) 0.7615 (4) 0.2310 (4) 0.9719 (4) 0.2758 (4) 0.0356 (5) 0.7122 (4) 0.7279 (4) 0.7704 (4) 0.3049 (4) 0.3504 (5) - 0.1060 (5) 1.1146 (5) - 0.0642 (6) 1.0732 (6) 0.469 (2) 0.544 (2)
-0.318 -0.369
0.757 (3) 0.797 (3)
0.544 (2) 0.480 (2)
0.0944 (1) 0.1701(3) 0.5897 (2) 0.8926 (2) 0.1506 (3) 0.4526 (3) 0.9110 (3) 0.0144 (7) - 0.1940 (7) 0.1335 (7) 0.9612 (7) 0.4068 (7) 0.9552 (6) 0.9687 (6) 0.7005 (7) 0.5167 (7) 0.1442 (8) 0.9206 (7) 0.6638 (7)
(5) (4)
factor, 0.56(2); “Occupation
trigonal-bipyramidal
WA.
Y
X
(with es&).
69
Ueq =
Ueq ( * 100) 2.80 (3) 4.25 (7) 4.02 (7) 3.67 (7) 5.47 (8) 5.8 (1) 5.53 (9) 3.0 (2) 5.5 (2) 3.0 (2) 3.8 (2) 3.8 (2) 5.1 (2) 4.9 (2) 5.8 (2) 3.7 (2) 2.9 (2) 3.6 (2) 6.1 (2) 3.1 3.0 3.5 3.1 3.4 3.3 3.3 3.3 3.4 3.2 3.2 4.0 4.1 5.2 5.0 6.0 5.7 11.3 11.3
(3) (3) (3) (3) (3) (3) (3) (3) (3) (3) (3) (3) (3) (4) (3) (4) (4) (5)X (5)”
11.3 (5)“X 11.3 (5P
factor, O&l(2).
geometries the value of T is zero and unity, respectively
Comparison with the T value of 0.36 in the structurally related methanol complex [61 suggests that the ethanol compound lies closer to the ideal&d square pyramidal geometry. Possible electronic spectroscopic implications of the more square pyramidal character are considered below. In general, the geometry of both methanol and ethanol complexes is comparable. The major difference in the stereochemistry of the CuN2Cl,0 chromophore
70 J. Casanova et al.
TABLE 3. Bond Lengths (A) (with esd’s) CumCl(l) C&)-O(3) W-W) S(la)-0(2a) S(laHX4a) WaXt7a) O(3)-Cc101 N(la)_C(la) N(3)-C(7) C(lM6) C(laX(2a) C(4M3) C(4a)_Wa) C(3M2)
2.281(2) 2.282(S) 1.606(5) l&1(5) 1.762(7) 1.744(7) 1.41(3) l&7(8) 1.342(8) 1.389(8) 1.3&l(8) 1.376(9) 1.401(8) 1.394(9)
cumCl(2) Cu(l)-N(la) W-O(2) S(la)-O(la) S(2)-c(7) S(2aM9a) O(3)-C(10’) N(3aM7a) N(3)-c(8) CWc(2) CXlaX(6a) C(2a)-C(3a) CWCt6) C(Wx9)
2.241(2) 2.071(5) 1.444(5) 1.448(5) 1.740(7) 1.734(8) 1.60 (3) 1.334(8) 1.393(8) 1.383(g) 1.386(g) 1.397(9) 1.384(9) 1.33 (1)
Cu(l)-N(1) W-O(l) S(lxx4) S(la)-N(2a) S(2Mx9) N(l)-C(1) O(3)-C(11’) N(3aM8a) N(2)-C(7) N(2a)-C(7a) C(4Mx5) C(4aM3a) C(6aKX5a) C(8aM9a)
Bond Angles (“) (with esd’s) c1(2)-Cu(l)-cl(1) N(l)-Cu(l)-Cl(2) O(3)-Cu( l)-Cl(2) N(la)-CuWClW N(la)_Cu(l)-N(1) N(2)-S(l)-00) 0(2M(l)-N(2) CW-S(l)-N(2) O(la)-S(labO(2a) N(2a)-S(la)-O(la) CX4a)-S(la)-O(la) cx9)-s(2Mx7) c(l)-N(l)-Cu(1) cx10’)-0(3)-cu(1) c(ll’)-o(32cu(l) C(11 ‘)-o(3)-C(10’) C@a)-N(3a)-C(7a) c(7)-N(2)-SW C(2)-C(l)-NW Cf7a)-N(2a)-S(la) Cf6a)-C(la)-N(la) c(5)-c(4)_S(l) cx3Mx4M5) CX3a)-C(4aHla) C(5a)-C(4a)-C(3a) N(3a)_C(7a)-S(2a) N(2a)-C(7a)-N(3a) NW-C(7H2) N(2)-C(7)-N(3) C(Sa)-CX6aMla) Ct2)-Cx3m4) C(9)-C(8)-N(3) c(8)-C(9)-S(2) CW-cxlO)-O(3)
153.9(l) 89.9(2) 111.8(2) 90.7(2) 171.0(2) 104.3(3) 111.7(3) 106.8(3) 117.8(3) 104.3(3) 106.8(3) 90.7(4) 116.7(4) 122.0) 117.0) 40X2) 114.7(6) 121.9(5) 120.1(6) 122.3(5) 119.4(5) 119.5(5) 120.7(6) 120.3(5) 120.4(6) 110.0(5) 119.9(6) 109.8(5) 120.1(6) 119.5(6) 119.8(6) 113.0(7) 1X6(6) 115.(2)
N(l)-Cu(l)-Cl(l) o(3)_cu(l)-cl(1) 0(3)-0.1(1)-N(l) N(la)-Cu(GCl(2) N(la)-Cu(l)-O(3) ocWWo(1) C(4)-SW-O(l) C(4)-W-O(2) N(2a)-S( la)-O(2a) C(4a)-S(la)-O(2a) C(4a)-S(la)-N(2a) C(9a)-S(2a)-C(7a) c(10)-0(3)-cu(1) c(10’)-0(3)-q10) c(11’)-0(3)-q10) C(la)-N(la)-Cu(1) C(8)-N(3)-C(7) C(6)-C(l)-N(1) C(2)-C(l)-C(6) C(2a)-C(la)-N(la) C(6akC(labC(2a) C(3)-C(4)-S(1) C(3a)-C(2a)-C(la) C(Sa)-C(4aMla) C(6)-C(5)-C(4) N(2a)-C(7a)-S(2a) C(5M6HXl) N(2)-C(7)-S(2) C(4a)-C(3aX(2a) C(6a)-C(5a)-C(4a) c(3)-c(2)-cx1) C(9a)_C@a)-N(3a) C@a)-C(9a)-S(2a) c(10%00)-0(3)
92.9(l) 94.3(2) 85.4(2) 90.5(2) 86.2(2) 118.7(3) 107.7(3) 107.1(3) 112.5(3) 107.6(3) 107.4(3) 90.6(4) 130. (1) 45.(l) 15.(2) 116.6(4) 114.8(6) 119.6(6) 120.3(6) 119X(6) 120.8(6) 119xX5) 119.5(6) 119.2(5) 119.8(6) 130.1(5) 119.8(6) 130.1(5) 120.0(6) 119.8(6) 119.7(6) 112.6(7) 112.0(6) 76.0(3)
2.066(5) 1.448(5) 1.768(6) 1.615(5) 1.737(8) 1.450(g) 1.99 (3) l&5(8) 1.328(8) 1.323(8) 1.385(9) 1.370(9) 1.385(9) 1.32 (1)
COORDINATION
FIGURE 2.
BEHAVIOR
OF SULFATHIAZOLE
71
ORTEP drawing of the [Cu(Hst.&(EtOH)C1J.
is the O(3)-Cu-Cl(l) bond angle which may be related to a different hydrogen bond and to the larger size of the ethanol molecule. Both complexes represent a series of distortion isomers whose geometries reflect the flexible stereochemistry of the C&I) ion or “plasticity effect,” the apical group being the main cause for the structural differences. I&and Conformation
and Molecular Packing
There are no unusual bond lengths or angles in the coordinated Hstz ligand, %xcept for the NW-C(l) bond di$ance that it is significantly increased [1.450@) A in the complex and 1.401(4) A in the sulfathiazole polymorph II [2OU.This fact, also found in the methanol complex is due to the interaction of the N(1) atom with the metal ion. The study of the W-N(2), N(2)_CX7), and c(7)_N(3), distances suggests that, as it occurs in the [Cu(Hstz&&kOH)c1,1 compound, the sulfathiazole presents the imido form. Neighboring molecules are interconnected by a hydrogen bonding system involving the nitrogen atoms, the chloride anions, and the sulfonamide and
72 J. Casanova et al.
ethanolic oxygen atoms. Relevant hydrogen bonding distances are given in Table 4. Figure 3 shows a perspective view of the detail of the molecular packing showing the N-H.. . H interbonding. Spectroscopic Data IR and Electronic Spectra. The IR spectrum shows the characteristic bands of the Hstz ligand. The bands of 3300 and 3240 cm-‘, assigned to the v,(N-H) and I.&N-H) vibrations of the NH, group, are shifted respect to those of the ligand (3320 and 3280 cm-‘). As expected, the bands due to Y(SO&~ ,(1320 cm-‘), v(SO,),(1140 cm-‘), scissors and wagging SO2 (570 and 550 cm-‘) vibrations remain unchanged. There is no change of the bands at 1540, 920, and 680-640 cm-‘, which have been assigned to the characteristic thiaxole ring and v(S-N) and v(C-S) vibrations, respectively. The presence of coordinated alcohol is indicated by a strong broad band at 3550-3400 cm-’ [211. The new band at 1060 cm-’ is attributed to bending M-OH vibrations [22]. The d-d electronic spectrum of the complex consists of an asymmetric broad band with a maximum approximately at 14140 cm-‘. In order to reveal the correlation of the d orbitals, the solid state visible spectrum was deconvoluted into Gaussian component bands by curve-fitting iteration processes. Three Gaussian component bands (15730, 14470, and 12050 cm-‘) with that of the highest frequency having the greatest intensity were resolved. On the basis of
TABLE 4. Distances and Angles for H-Bonds (with esd’s) Donor-H N(l)_Hll 1.08oc3) NWH12 l.ow7) 0(3)_H31 0.97(l) N(la&Hlla l.ow7) N(la)_Hlla 1.08oc7) N(la)_HUa 1.08oc7~ N(la)-H12a 1.080(7) NW-H12 l.o800 N(la>Hlla 1.W7)
a3Mr31 0.97(l) No-H21 l.Oso@)
Donor..Acceptor
H..Acceptor
N(l)..CI (2) 3.045w N(l)..0 (3) 2.95(l) 0(3)..N (la) 2.977(7) N(la)..Q (1) 3.099(6) N(laI.0 (3) 2.977(7) N(laLCl(2) 3.065(7) N(lak.0 (la) (1) 3.025(7) NW...0 (1) (1) 3.093(7) N(la)..Cl (1) (2) 3.46X6)
Hll...Cl(2) 2.687(5) H12...0 (3) 2.58(l) H3l...N (la) 2.648(7) Hlla..Cl(l) 2.8320 Hlla...O (3) 2.590@) H12a...Cl(2) 2.786(7) HEa... (la) (1) 2.086(S) H12...0 (1) (1) 2.151(7) Hlla...Cl(l) (2) 2.405(6) H31...Cl(l) (2) 2.1940) H21...N (3) (3) 1.848@)
o(3x.a (1) (2) 3.124X8) N(2h.N (3) (3) 2.875(8)
symmetry codes: (1) -X - 1, +Y, +z; (2) -x,
-y
+ I, --z + 1; (3)
Donor..Acceptor N(l)_Hll...Cl(2) 98.8(5) N(l)_H12...0 (3) 99.5(5) y(&fI$...N (la) N(l&Hlla...Cl(l) 93.90 N(la)_Hlla...O (3) 100.2(5) N(la)_HKIa...Cl(2) 94.4(5) N(la>HUa...O (la) (1) 143.7(6) N(lkH12...0 (1) (1) 144.X6) N(la)_Hlla...Cl (1) (2) 16&X.3(6) cm&y
...Cl (1) (2)
N(2iH21...N 157.3(7) -X
+
I, -y
+
(3) (3)
I, -x.
COORDINATION
BEHAVIOR
OF SULFATHIAZOLE
73
FIGURE 3. Perspective view of a detail of the molecular packing, showing the N-H.. . N interactions.
the d orbital splitting, the orbital sequence of the title complex is intermediate between those of trigonal bipyramidal and square pyramidal symmetry [23, 241 A comparison of this spectrum with that of the methanol complex does not give evidence to establish if the basic metal geometry in the title compound leads to a more square pyramid, such as has been indicated by the structural parameter 7. EPR Spectrum of the [Cu(Hst.d2(EtOH)Cl,] Complex The polycrystalline EPR spectrum is rhombic (Fig. 4). The calculated EPR parameters are g, = 2.05, g, = 2.09, and g, = 2.28. Fo r systems with g, >g, >g, the ratio R=(g,gr)& - g2) indicates the ground state of the complex [251. If R > 1 the ground state is predominantly d,z while the ground state is predominantly d,z_,,z when the value of R is less than one. The complex presents an R value of 0.17 indicating d,z_y2 as the ground state. Thus the complex may be five coordinated square pyramid, which has been confirmed by its crystal structure. The calculated R value of 0.5 for the [Cu(Hstz),(MeOH)Cl,] complex is higher than that for the ethanol compound, suggesting a more trigonal geometry for the methanol complex as it also indicated by its T value. Superoxide Dismutation Activity The superoxide dismutase activity of the drug and the [Cu(Hstz),(EtOH)cl,], [Cu(Hstz),(MeOH)Cl,], and Zn(stz),.H,O complexes were assayed by their
74
J. Casanova et al.
FIGURE 4.
Polycrystalline powder EPR spectrum of the Cu(Hstzl,(EtOH)Cl~ cmpkx.
ability to inhibit the reduction of nitroblue tetrazolium [26]. The superoxide scavenging data (Fig. 5) indicates that the Zn(I1) complex is only marginally more active than the free drug. The copper complexes exhibit a greatly increased superoxide dismutase activity compared with the parent drug molecule. Table 5 shows the chelate or superoxide dismutase concentrations required to produce 50% inhibition of NE3T reduction (IC,,). On a molar basis the methanol
0
5
IO
15
20
Concentration (PM) 5. Inhibition of. NBT reduction in the presence of metal complexes 0- 0 Cu(Hstz),(MeOHlCl,, 0-0 Cu(Hstz),(EtOH)Cl,, q - 0 Zn(Stz),H,O, n - n Hstz. Each point repreknts the mean f standard deviation of triplicate determinations.
FIGURE
COORDINATION
BEHAVIOR
OF SULFATHIAZOLE
75
TABLE 5. Superoxide Dismutase-Mimetic Activity IC,,a ( PM) Cu(Hstz)z(MeOH)cl z Cu(Hstz),(EtOH)Cl, Suoeroxidedismutase
2.510 5.170 0.006
- Log Ic,o 5.60 5.28 8.21
‘IC,, is defined as the concentration of complex or enzyme which produces 50% inhibition of NBT reduction. A molecular weight of 31,200 was considered for calculating the enzyme concentration.
coordinated complex is more efficient as superoxide radical scavenger than the ethanol complex being the former, 400, and the latter, 800 times less effective than superoxide dismutase. For the zinc complex a proper IC,, could not be established. The values presented here are similar to the ones published by Sorenson et al. [27] and Bijloo et al. [28] for the enzyme and other copper complexes. The mechanism believed to be operating in the naturally occurring superoxide dismutases involves one-electron reduction of the metal ion of the active center by superoxide followed by reoxidation of the reduced metal ion by a second superoxide anion. Metal complexes that can undergo such redox cycling are likely to function as superoxide scavengers. It is assumed that electron transfer between the metal center and superoxide anion radicals occurs by direct binding [29]. A fast exchange of coordinated EtOH or MeOH and limited steric hindrance to the approach of the superoxide anion are considered essential requirements for the successful binding of the 0; radical [30]. The high superoxide dismutase activity could also be explained considering a favorable response of r-electrons of the aromatic side chain in stabilizing the 01-o; interactions. Moreover, as suggested by Srivastava et al. [31], the distorted geometry of these complexes may favor the geometrical change, which is essential for the catalysis as the geometry of copper in SOD enzyme changes from distorted squared pyramidal (for C&I)) to distorted tetrahedral (for C&I)) during the catalysis. We consider that these requirements are satisfied in the distorted complexes [Cu(Hstz),(EtOH)Cl,] and [Cu(Hstz),(MeOH)Cl,] and that the doubled superoxide dismutase-mimetic activity of the latter can be related with its higher geometrical distortion. Supplementary
Material
Anisotropic temperature factors, a list of observed and calculated structure factors, fractional positional parameters of the hydrogen atoms, torsion angles and angles between least-square planes and lines are available from the authors on request. JC, GA, and JB appreciate financial suppoti from FAR91 -197(CICYT). JC acknowledges the Generalitat Valenciana for a grant. JT appreciates FIS grant No. 92 /26X
REFERENCES 1.
A. Bult, in Metal Ions in Biological Systems, H. Sigel, Ed., Marcel Dekker, New York 1983.
16
J. Casanova
et al.
2. J. R. J. Sorenson, Chem. in Brit. 16, 1110 (1984). 3. R. K. Gouch, T. W. Kensler, L. W. Oberley, and J. R. J. Sorenson, in Biochemical and Inorganic Copper Chemistry, K. D. Karlin and J. Zubieta, Eds., Adenine, New York, 1986, Vol. 7. 4. A. Buh, Pharm. Weekblad Sci. Ed. 3,213 (1981). 5. J. Casanova, G. AIzuet, S. Ferrer, J. Borras, S. Garcia-Granda, and E. Perez-Carrefio, J. Inotg. B&hem., 51, 689 (1993). 6. J. Casanova, G. AIzuet, J. Borrh, J. M. Amigo, and T. Debaerdemaeker, Zeit. jiir fit., 209, 271 (1994). 7. J. Casanova, G. Aizuet, J. BorrL, L. David, and D. Gatteschi, Inorg. Chim. Acta, 211, 183 (1993). 8. M. S. Lehman and F. K. Larsen, Acta Cryst. A30, 580 (1974). 9. D. F. Grant and E. J. Gabe, J. Appl. Cryst. 11, 114 (1978). 10. A. C. T. North, D. C. Phillips, and F. S. Mathews, Acta Cryst. A24, 351 (1968). 11. G. M. Sheldrick, in Crystallographic Computing 3, edited by G. M. Sheldrick, C. Kruger, and R. Goddard, Eds., &ford, Ciarendon Press, 1985, p. 175. 12. G. M. Sheldrick, SHELX76. Program for crystal structure determination, Univ. of Cambridge, England, 1976. 13. N. Walker and D. Stuart, Acta Cryst. A39,158 (1983). 14. International Tables for X-Ray Crystallography, Byrmingham, Kynoch Press (Present distributor KIuwer Acad. Publ., Dordrecht), 1974, Vol. IV. 15. M. Nardelh, Comp. Chem. 7,95 (1983). 16. A. L. Spek, in Computational Crystallography, edited by SAYRE, Gxford, Clarendon Press, 1982, p. 528. 17. L. W. Oberley and D. R.‘Spitz, in Handbook of Methods for Gqgen Radicals Research, R. A. Greenwald, Ed., CRC Press, Boca Raton, 1986, pp. 217-220. 18. B. J. Hathaway, in Comprehensive Coordination Chemistry, Eds. G. Wilkinson, R. D. Gillard, and J. A. Cleverty, Eds., Pergamon Press, New York, 1987, Vol. 5. 19. W. Addison, T. N. Rao, J. Reedijk, J. van Rijn, and G. C. Verschoor, J. Chem. Sot., Dalton Trans., 1349 (1984). 20. G. J. Kruger and G. Gafner, Acta Ctyst., B27, 326 (1971). 21. S. S. Tandon, L. K. Thompson, and R. C. Hynes, Znotg. Chem. 31, 2210 (1992). 22. K. Nakamoto, Infired and Raman Spectra of Inorganic and Coordination Compounds, John Wiley & Sons, New York, 1986,4th edn. 23. B. J. Hathaway and D. E. Billing, Coord. Chem. Rev. 5, 143 (1970). 24. (a) A. Bencini and D. Gatteschi, Trans. Met. Chem. 8, 2 (1980); (b) A. Bencini, I. Bertini, D. Gatteschi, and A. Scozzafava, Inorg. Chem. 17, 3194 (1978). 25. M. F. El-ShazIy, A. El-Dissowky, T. Salem, and M. Osman, Znorg. Chim. Acta 40, 1 (1980). 26. A. Bury, A. E. Underhill, D. R. Kemp, N. J. O’Shea, J. P. Smith, and P. S. Gomm, Znorg. Chim. Acta 13885 (1987). 27. H. H. A. DoIIwet, J. B. McNicholas, A. Pezeshk, and J. R. J. Sorenson, Trace Elements in Medicine 4, 13 (1987). 28. G. J. Bijloo, H. van der Goot, A. Bast, and H. Timmerman, J. Znotg. Biochem. 40, 237 (1990). 29. J. A. Fee, in Metal Ions in Biological Systems, H. Sigel, Ed., Marcel Dekker, New York, 1981, Vol. 13. 30. M. Boden, M. C. Holmes, and P. F. Knowles, Biochem. Biophys. Res. Commun. 57, 845 (1974). 31. R. G. Bhirud and T. S. Srivastava, Znorg. Chim. Acta 179, 125 (1991). Received July 13, 1993; accepted September 15, 1993