Transition and Group IIB Metal Complexes With “Active Aldehyde” Derivatives of Thiamine

The Zn2+, Cd2+, Hg2+, Co2+ and Ni2+ ions produce zwitterionic type complexes with the ligands (L), 2-(α-hydroxy-benzyl)thiamine=HBT and 2-(α-hydroxy-cyclohexyl-methyl)thiamine = HCMT, of the type MLCl3. The ligands are in the S conformation, the metals are bound to N1, of the pyrimidine moiety of thiamine and the complexes have a trigonally distorted tetrahedral structure, as the crystal structure of the complex Zn(HCMT)Cl3 (orthorombic, a=14.4 b=14.1 c=17.4 β=105.6O V=3392A3 R=13.8%), the one and two dimensional 1H nmr spectra of the Zn2+, Cd2+ and Hg2+ complexes and the electronic spectra of the Co2+ and Ni2+ complexes show. A brief review of the previous techniques (structure of the Hg(HBT)Cl3 complex, IR-Raman spectra, 13C nmr in solution and solid state etc) used to characterize these complexes, is also given here and the proper conclusions drawn.

metal-ligand bonding, due to the net positive charge on thiamine and to the easy protonation of the N 1, site of pyrimidine (pKa-5) [3]. Despite this difficulty, a few metal complexes of thiamine with Cd2+, Zn2+, Pt2+, Mn 2+ etc containing mainly a M-N 1, direct bonding were prepared in recent years and their structures were solved with X-rays [4].
The use however of the "active aldehyde" intermediates of thiamine as ligands for the formation of complexes with bivalent metal ions, presented the advantage of the delocalization of the net positive charge on N 3 of thethiazolium moiety to the sulfur atom and resulted to the easier formation of compounds with M-N 1, bonds [3][4][5][6][7].
We summarize here all the techniques used thus far for the characterization of these complexes, adding a few more, as well as the conclusions drawn concerning the enzymatic action of thiamine. M. Louloudi, NickHadjiliadis, J.-P. Laussac andR. Bau Metal-BasedDngs Materials and Methods: The preparation of the ligands (L) 2-(ahydroxybenzyl)thiamine HBT and 2-(a-hydroxycyclohexyl-methyl) thiamine HCMT and the complexes MLC13 (M=Zn2+, Cd2+, Hg2+, Co2+, Ni2+) was described previously [3,6]. The structure of the complex Zn(2-(ahydroxycyclohexylmethyl) thiamine)C13 was solved with the same method and instrument used for the Hg(HBT)C13 complex [3]. Crystals of the complex were grown by slowly diffusing acetone and ether in a solution of the complex in methanol. They were unstable in the absence of the solvents and were mounted in a sealed capillary and the data collection followed.
The 1D-1H nmr spectra were recorded on a Brucker AC 200 Spectrometer with TMS as an internal standard.
The 2D-1H nmr spectra were also obtained on a Brucker AC 200 spectrometer at 200 MHz. The pulse sequence 90-tl-90-rm-90 was followed with mixing time of 500 ms.
The DRS and the solution UV-Vis spectra were recorded as described [6].
Results and Discussion: Metal complexes of the li B group metal ions as well as Co2+, Ni 2+ and Cu 2+ were used for the preparation of the 1:1 complexes with the "active aldehyde" derivatives of the thiamin HBT and HCMT that corresponded to the general formulae MLC13, with L the above thiamine derivatives and M all the metal ions except Cu 2+ [3][4][5][6][7]. The latter was oxydizing both ligands to thiochrome [6,8] (i) X-ray crystal structures The first crystal structure reported was the one of Hg 2+ with 2-(ahydroxybenzyl)thiamine, Hg(HBT)C13 [3]. Here the ligand had the less common S  [9] defined the S conformation of thiamine with bT=+100o, bp=+lS0o, the V conformation with T=+90 , p=+90 and the F conformation with T=0 , bp=+/-90). These was a direct Hg-N 1, bonding of 2.23k [3].
A second crystal structure of xhe compound Zn(2-(cx-hydroxyclohexylmethyl)thiamine)C13, Zn(HCMT)C13 was subsequently solved but only to an R value of about 13.8%. Despite the low accurancy in the details of the structure, it is clear that the bonding of the metal takes place again with the N 1, of the pyrimidine moiety (The bond distance Zn-N 1, is about 2.1 A). Volume 1, No,: 2-3, 1994 Transition and Group lib Metal Complexes with "Active Aldehyde"Derivatives of Thiamine The HCMT ligand is also in the S conformation with .p~172.8 and T~-92.7 and the configuration around Zn is pseudotetrahedral [3]. (ii) Vibrational IR-Raman Spectra The comparison of the IR and Raman spectra of the various complexes with the ones of known structure, showed that they were all similar band by band, except the metal-ligand (M-N, M-C1) streching vibrations [5]. Another difference of the various complexes was the difference at the position of the first vC=N vibration of the pyrimidine moiety, varying with the bulkiness of the N 1, coordinated metal ion and the protonation. This frequency was increasing in the order H + Co 2+ Ni 2+ Zn 2+ Cd 2+ Hg 2+ [3,5].

CI
(iii)__NMR Spectra Comparison of the 13C nmr of the various metal complexes in solution and the solid state showed that they were all isostructural in both phases, to the ones with known structures and that both the M-N 1' bonding and the S conformation of the ligands were retained in D20 and DMSO-d 6 solutions [7]. This was also confirmed with the 199Hg nmr spectra [7] of the mercury complexes. ()_IH nrnr spectra (1D): The 1H nmr chemical shifts of the various ligands and their II B metal complexes in DMSO-d 6 solutions, are included in Table I. Protonation of both ligands at N 1, of pyrimidine causes downfield shifts to its adjacent protons, C6,-H and C2,-CH 3. This however, does not cause significant changes to the neighboring coupling constants (Table I), indicating the absence of conformational changes of the molecule upon protonation. The broad bands of HBT.HC1 and HCMT.HC1 located at -8.9 and 9.254 ppm respectively, are assigned to the N(4'a)H 2 protons in exchange with their N(I')-H protons. This is evidenced from the fact that in the non protonated ligands, they are located at 7.3-7.5 ppm as sharp bands as expected for aminopyrimidines [10]. Unexpectedly, the neighboring protons to the N 1, site of pyrimidine are not shifted significantly upon metal coordination.
In a first approach this can be explained as the result of two opposite effects"  [11,12]. Similar were the results of the 13C nmr in D20 and DMSO-d 6 solutions for the carbon atoms adjacent to N 1, that can not be due to the breaking of the M-N 1, bonding in solution, since the behavior was also similar in the solid state 13C nmr spectra [7].
All the protons of pyrimidine in HBT, HBT.HC1 and their complexes are more deshielded compared to the ones of HCMT, HCMT.HC1 and their complexes and therefore more upfield shifted than the latter (Table I). This is due to the stacking effect of the benzyl ring, being almost parallel to the pyrimidine in the HBT derivatives (dihedral angle 9 and shorter distance 3.4A) [3,13,14]. The same was observed in the solution and solid state 13C nmr spectra of all the HBT and HCMT complexes for the adjacent to N 1, carbon atoms [7]. Hg(HBT)C13 [3]. The same is true in the structure of Zn(HCMT)C13 that does not change in solution as well [7]. It should be noted here that the C(3,5')H 2 methylenic protons behave as an AB system in the protonated and non protonated ligands due to the chiral center at C(10) [15]. In their metal complexes however, they are seen as isochronous.  Fig.2. 1H nmr NOESY spectrum (contour plot) of the complex Zn(HBT)C13, in DMSO-d6. Volume 1, No,: 2-3, 1994 Transition and Group lib Metal Complexes with "Active Aldehyde" Derivatives of Thiamine interaction between the benzyl and the pyrimidine rings found in the solid state [3] persists also in solution, we have recorded the 2D-NOE spectrum of the complex Zn(HBT)C13 in DMSO-d 6 (Fig.2). For comparison the 2D-NOE spectrum of the complex Zn(HCMT)C13 was recorded as well. From Fig.2, it is easily seen that enhanced connectivities exist between the protons C(6')-H/C(10)-H, C(6')-H/C(2",6")-H and a weak cross peak between C(6')-H and O(ll)-H. Therefore the pyrimidine C(6')-H proton approaches the C(10)-H and O(ll)-H and the C(2",6") protons of the benzene ring. This indicates that the benzene and pyrimidine rings are almost parallel, like in the solid phase.
Comparison of the 2D-NOE spectra of the complexes Zn(HBT)C13 and Zn(HCMT)C13 show that in the second case there were not observed any cross peak connectivities between pyrimidine and cyclohexane, thus excluding any interaction (stacking, hydrophobic) between these two rings, as this is also true in the solid phase.
(iv) Electronic Spectra The Co 2+ and Ni 2+ complexes of the general formulae MLC13 should have similar structures (M-N 1, bonding and $ conformation of the ligands, pseudotetrahedral structures around the metals) with the ones of known structures Hg(HBT)C13 and Zn(HCMT)C13 as exposed above (similar IR-Raman spectra band by band).Their pseudotetrahedral structures are further substantiated by their electronic DRS (diffuse reflectance spectra) and DMF solution spectra (Table II).
The same geometries are retained in DMF solutions, while in D20 they are transformed to octahedral ones (Table II)    In the case of the Ni(HBT)C13 the multiple bands near 8700 cm -1 and 15900 Volume 1, Nos. [2][3]1994 Transition arid Group liB Metal Complexes with "4ctive Aldehyde" Derivatives of Thiamine cm -1 in the DRS are due to the 3TI(F) ---3A2(F) and 3TI(F) ---, 3TI(P) transitions in a tetrahedral environment [16]. Their multiplicity is again due to a trigonal distortion (C3v symmetry) (See Table II) [18]. In DMF solutions the geometry is retained but it becomes again octahedral in aqueous solutions (See Table II). The band at 4390 cm -1 is assigned to the (Vl) 3T2(F)---,3A2(F) transition. Therefore 10Dq=vl=4390 cm-1.
Finally, the ligand HBT shows a maximum at 274 nrn, the HBT.HC1 at 276 nm and the complexes Co(HBT)C13 and Ni(HBT)C13 at 275.5 and 275 nm respectively, in DMF solutions. The HCMT, HCMT.HC1 and Co(HCMT)C13 show the same band at 272, 273 and 272.5 nm correspondingly. This band is assigned to a rt---rt* transition of the pyrimidine ring of thiamine. Its bathochromic shift in the protonated and metallated ligands indicate the involvement of the pyrimidine (N1,) moiety to protonation and metallation [20].
Concluding Remarks: The easy formation of metal complexes with the "active aldehyde" derivatives of thiamine with a M-N 1, bonding, may indicate that the intervention of the metal ions follows the formation of these intermediates during the enzymatic process. The S conformation of the ligand seems to be important during the enzymatic action, since it persists in all the C(2a) substituted thiamine derivatives. (The V conformation of thiamine and the pyrophosphate metal binding are also important, after liberation of the C(2a) substituent, since it was recently found in the structure of transketolase, containing Ca 2+ ions [21].