Analogues of vaccinia virus DNA topoisomerase I modified at the active site tyrosine

The mechanism of type IB topoisomerase-mediated DNA relaxation was studied by modification of vaccinia topoisomerase I at the active site tyrosine (position 274) with several tyrosine analogues. These analogues had varied steric, electronic, and stereochemical features to permit assessment of those structural elements required to support topoisomerase function. Eleven tyrosine analogues were successfully incorporated into the active site of vaccinia topoisomerase I. It was found that only tyrosine analogues having the phenolic -OH group in the normal position relative to the protein backbone were active. Modifications that replaced the nucleophilic tyrosine OH (pKa approximately 10.0) group with NH2 (pKa 4.6), SH (pKa approximately 7.0), or I groups or that changed the orientation of the nucleophilic OH group essentially eliminated topoisomerase I function. For the active analogues, the electronic effects and H-bonding characteristics of substituents in the meta-position of the aromatic ring may be important in modulating topoisomerase I function. The pH profile for the functional analogues revealed a small shift toward lower pH when compared with wild-type topoisomerase I.     

Prediction of pKa values of nido-carboranes by density functional theory methods

A great parallel exists between metal complexes of cyclopentadienyl and arene ligands on one side and metal complexes of the nido derivatives of the icosahedral o-carborane clusters. With few exceptions, the metal complexation in the cluster can be viewed as the substitution of one or more bridging hydrogen atoms by the metal. Therefore, a necessary requirement for the complexation is the deprotonation of the nido cluster to generate a coordination site for that metal. The reaction to remove these protons, which most probably is one of the most commonly done processes in boron and metallaborane chemistry, is barely known, and no quantitative data are available on the magnitude of their pKa values. With the purpose of determining the acidity of nido-carboranes, a procedure to calculate the pKa values of nido boron clusters is presented in this paper for the first time. To this objective, some nido clusters have been selected and their geometry and NMR-spectroscopic properties have been studied, giving a good correlation between the theoretical and experimental data in both geometry distances and 11B NMR spectroscopy. Of notice is the result that proves that the singular carbon atom in the thermodynamic isomer of [C2B10H13]- is definitely part of the cluster and that its connection with the C2B3 face would be better defined by adding additional interactions with the two boron atoms nearest to the second cluster carbon. The pKa values of the nido species have been calculated by correlating experimental pK(a) values and calculated reaction Gibbs energies DeltaG(s). Some pKa values of importance are -4.6 and +13.5 for 7,8-[C2B9H13] (1) and 7,8-[C2B9H12]- (2), respectively.     

Varying acidity of aqua ligands in dependence on the microenvironment in mononucleobase (nb) complexes of type cis- and trans-[Pt(NH3)2(nb)(H2O)]n+

Aqua ligands in mixed aqua/nucleobase metal complexes are potential sites of acid-base catalysis and/or, when present as hydroxo ligands, can directly be involved in hydrolysis reactions. pKa values of close to 7 are consequently of particular interest and potential significance. Here we report on the differential acidity of aqua complexes in model nucleobase (nb) complexes of cis- and trans-[Pt(NH3)2 (nb)(H2O)]n+ and discuss reasons as to why the nb in cis complexes influences the pKa (pKa 4.8-7.0), whereas in trans complexes the pKa values are rather constant (pKa approximately 5.2-5.3). The results of DFT calculations of a series of mono(nucleobase) complexes derived from cis-Pt(NH3)2 are critically examined with regard to the role of exocyclic groups of nucleobases in stabilizing aqua/hydroxo ligands through intracomplex hydrogen bond formation. This applies in particular to the exocyclic amino groups of nucleobases, for which gas-phase calculations suggest that they may act as H bond acceptors in certain cases, yet in the condensed phase this appears not to be the case.     

Delamination and aromatic amine intercalation of layered aluminophosphate with [Al3P4O16]3- stoichiometry

The delamination and intercalation of a layered microporous aluminophosphate, [Al3P4O16](3-).3[CH3(CH2)NH3]+ (AlP), with aromatic amine have been carried out and were followed by XRD and SEM measurements. The basicity of the amine plays an important role in this process, as do the dielectric constant of the solution and the amount of amine added. A saturated benzylamine (pKa of 9.34) intercalate of the aluminophosphate are obtained in solutions with dielectric constant of 50-70 and an amine concentration of 10 mmol/g AlP, while no aniline (pKa of 4.60) intercalates are formed under similar conditions. The remarkable effect of basicity of the amine can be explained by a metathetical balanced reaction model, which was proved by the results of the intercalation processes of 4-methylpiridine (pKa = 6.00) and 4-methylimidazole (pKa = 7.55).     

The chemical nature of the 2'-substituent in the pentose-sugar dictates the pseudoaromatic character of the nucleobase (pKa) in DNA/RNA

We here show that the pKa (error limit: 0.01 to 0.03 pKa unit) of a nucleobase in a nucleotide can be modulated by the chemical nature of the 2'-substituent at the sugar moiety. This has been evidenced by the measurement of nucleobase pKa in 47 different model nucleoside 3',5'-bis- and 3'-mono-ethylphosphates. The fact that the electronic character of each of the 2'-substituents (Fig. 1) alters the chemical shift of the H2' sugar proton, and also alters the pKa of the nucleobase in the nucleotides has been evidenced by a correlation plot of pKa of N3 of pyrimidine (T/C/U) or pKa of N7 of 9-guaninyl with the corresponding deltaH2' chemical shifts at the neutral pH, which shows linear correlation with high Pearson's correlation coefficients (R = 0.85-0.97). That this modulation of the pKa of the nucleobase by a 2'-substituent is a through-bond as well as through-space effect has been proven by ab initio determined pKa estimation. Interestingly, experimental pKas of nucleobases from NMR titration and the calculated pKas (by ab initio calculations utilizing closed shell HF 6-31G** basis set) are linearly correlated with R = 0.98. It has also been observed that the difference of ground and protonated/de-protonated HOMO orbital energies (DeltaHOMO, a.u.) for the nucleobases (A/G/C/T/U) are well correlated with their pK(a)s in different 2'-substituted 3',5'-bis-ethylphosphate analogs suggesting that only the orbital energy of HOMO can be successfully used to predict the modulation of the chemical reactivity of the nucleobase by the 2'-substituent. It has also been demonstrated that pKa values of nucleobases in 3',5'-bis-ethylphosphates (Table 1) are well correlated with the change in dipole moment for the respective nucleobases after protonation or de-protonation. This work thus unambiguously shows that alteration of the thermodynamic stability (Tm) of the donor-acceptor complexes [ref. 20], as found with various 2'-modified duplexes in the antisense, siRNA or in triplexes by many workers in the field, is a result of alteration of the pseudoaromatic character of the nucleobases engineered by alteration of the chemical nature of the 2'-substitution.     

Acid-base properties of the nucleic-acid model 2'-deoxyguanylyl(5'-->3')-2'-deoxy-5'-guanylate, d(pGpG)3-, and of related guanine derivatives

The dinucleotide d(pGpG) is an often employed DNA model to study various kinds of interactions between DNA and metal ions, but its acid-base properties were not yet described in detail. In this study the six deprotonation reactions of H4[d(pGpG)]+ are quantified. The acidity constants for the release of the first proton from the terminal P(O)(OH)2 group (pKa = 0.65) and for one of the (N7)H+ sites (pKa = 2.4) are estimated. The acidity constants of the remaining four deprotonation reactions were measured by potentiometric pH titrations in aqueous solution (25 degrees C; I = 0.1 M, NaNO3): The pKa values for the deprotonations of the second (N7)H+, the P(O)2(OH)-, and the two (N1)H sites are 2.98, 6.56, 9.54 and 10.11, respectively. Based on these results we show how to estimate acidity constants for related systems that have not been studied, e.g. pGpG, which is involved in the initiation step of a rotavirus RNA polymerase. The relevance of our results for nucleic acids in general is briefly indicated.     

Ruthenium(II) complex of Hbopip: synthesis, characterization, pH-induced luminescence "off-on-off" switch, and avid binding to DNA

A novel ruthenium(II) complex of [Ru(bpy)2(Hbopip)](ClO4)2 (in which bpy=2,2'-bipyridine, Hbopip=2-(4-benzoxazolyl)phenylimidazo[4,5-f][1,10]phenanthroline) was synthesized and characterized. The spectrophotometric pH titrations of the complex showed that it acted as a pH-induced luminescence "off-on-off" switch: a luminescence off-on switch with a luminescence enhancement factor of IpH=3.0/IpH=1.0=20 occurring over a narrow pH range of 1.00-3.00 plus a luminescence on-off switch with a luminescence enhancement factor of 3 over a pH range of 3.20-9.40. The excited-state ionization constant of the complex derived, pKa1*=3.06, is 1.36 pKa units greater than the ground-state pKa1=1.70, and pKa2*=5.01 and pKa3*=8.22 are comparable to the ground-state pKa2=5.23 and pKa3=8.22, respectively. The complex avidly bound to calf thymus DNA with a large binding constant of (1.2+/-0.3)x10(7) M-1 in buffered 50 mM NaCl, as evidenced by UV-vis and luminescence titrations, steady-state emission quenching by [Fe(CN)6]4-, DNA competitive binding with ethidium bromide, viscosity measurements, and DNA melting experiments.     

An equilibrium analysis of the aqueous H+-MoO4(2-)-(HP)O(3)2- and H+-MoO4(2-)-(HP)O(3)2--HPO4(2-) systems

The speciation in the phosphitomolybdate system, H+-MoO4(2-)-(HP)O(3)2-, has been determined from combined potentiometric and 31P NMR measurements in 0.600 M Na(Cl) medium at 298(1) K. Potentiometric titration data were collected in the ranges 2.5<-log[H+]<6.2, 40.0相似文献   

Synthesis, structure, and acid-base and redox properties of a family of new Ru(II) isomeric complexes containing the trpy and the dinucleating Hbpp ligands

Three pairs of mononuclear geometrical isomers containing the ligand 3,5-bis(2-pyridyl)pyrazole (Hbpp) of general formula in- and out-[RuII(Hbpp)(trpy)X](n+) (trpy=2,2':6',2' '-terpyridine; X=Cl, n=1, 2a,b; X=H2O, n=2, 3a,b; X=py (pyridine), n=2, 4a,b) have been prepared through two different synthetic routes, isolated, and structurally characterized. The solid state structural characterization was performed by X-ray diffraction analysis of four complexes: 2a-4a and 4b. The structural characterization in solution was performed by means of 1D and 2D NMR spectroscopy for complexes 2a,b and 4a,b and coincides with the structures found in the solid state. All complexes were also spectroscopically characterized by UV-vis which also allowed us to carry out spectrophotometric acid-base titrations. Thus, a number of species were spectroscopically characterized with the same oxidation state but with a different degree of protonation. As an example, for 3a three pKa values were obtained: pKa1(RuII)=2.13, pKa2(RuII)=6.88, and pKa3(RuII)=11.09. The redox properties were also studied, giving in all cases a number of electron transfers coupled to proton transfers. The pH dependency of the redox potentials allowed us to calculate the pKa of the complexes in the Ru(III) oxidation state. For complex 3a, these were found to be pKa1(RuIII)=0.01, pKa2(RuIII)=2.78, and pKa3(RuIII)=5.43. The oxidation state Ru(IV) was only reached from the Ru-OH2 type of complexes 3a or 3b. It has also been shown that the RuIV=O species derived from 3a is capable of electrocatalytically oxidizing benzyl alcohol with a second-order rate constant of kcat=17.1 M(-1) s(-1).     

Evaluation of 2-methyl-3-hydroxy-4-pyridinecarboxylic acid as a possible chelating agent for iron and aluminium

In view of a possible application to Fe and Al chelation therapy, 2-methyl-3-hydroxy-4-pyridinecarboxylic acid (DT2) was synthesised, and its complex formation, electrochemical and cytotoxic properties were studied. The complexing properties of DT2 towards Fe(III) and Al(III) were investigated in aqueous 0.6 m (Na)Cl at 25 degrees C by means of potentiometric titrations, UV-vis spectrophotometry, and 1H NMR spectroscopy. DT2 is a triprotic acid (H3L+) having pKa1 = 0.47, pKa2 = 5.64 and pKa3 = 11.18. The metal-ligand complexes observed in solution and their corresponding stability constants (log beta values) are the following: FeLH (19.38), FeL (16.01), FeLH(-1) (12.28), FeL2H2 (37.29), FeL3H3 (53.41), FeL3H2 (47.99), FeL3H (41.21) and FeL3 (34.1); AlLH (17.43), AlL2H2 (33.74), AlL2H (27.6), AlL3H3 (48.72), AlL3H2 (42.67), AlL3H (35.8) and AlL3 (27.92). The complex formation between DT2 and Fe(II) was studied by UV-vis: the weak complex FeLH (log beta = 15.8) was detected. DT2 shows a lower complexation efficiency with Fe(III) and Al(III) than that of other available chelators, but higher than that of its non-methylated analogue 3-hydroxy-4-pyridinecarboxylic acid (DT0). The electrochemical behaviour of DT2 was investigated by means of cyclic voltammetry, indicating that the oxidation of the ligand proceeds through a two electron process with a CECE mechanism. Voltammetric curves suggest that the oxidation or the reduction of DT2 in vivo is unlikely. According to the thermodynamic data, also the Fe(III)-DT2 complexes do not undergo redox cycling at physiological pH. Amperometric titrations of solutions containing Fe(III) and DT2 at pH = 5 indicated the same Fe(III) : ligand stoichiometric ratio as calculated from potentiometric data. The toxicity of DT2 and of other simple hydroxypyridinecarboxylic acids was investigated in vitro and no cytotoxic activity was observed (IC50 > 0.1 mM) on cancer cell lines and also on primary human cells, following a three day exposure.     

Equilibrium and kinetic studies of the aquation of the dinuclear platinum complex [[trans-PtCl(NH3)2]2(mu-NH2(CH2)6NH2)]2+: pKa determinations of aqua ligands via [1H,15N] NMR spectroscopy

By the use of [1H,15N] heteronuclear single quantum coherence (HSQC) 2D NMR spectroscopy and electrochemical methods we have determined the hydrolysis profile of the bifunctional dinuclear platinum complex [[trans-PtCl(15NH3)2]2(mu-15NH2(CH2)(6)15NH2)]2+ (1,1/t,t (n = 6), 15N-1), the prototype of a novel class of potential antitumor complexes. Reported are estimates for the rate and equilibrium constants for the first and second aquation steps, together with the acid dissociation constant (pKa1 approximately pKa2 approximately pKa3). The equilibrium constants determined by NMR at 25 and 37 degrees C (I = 0.1 M) were similar, pK1 approximately pK2 = 3.9 +/- 0.2, and from a chloride release experiment at 37 degrees C the values were found to be pK1 = 4.11 +/- 0.05 and pK2 = 4.2 +/- 0.5. The forward and reverse rate constants for aquation determined from this chloride release experiment were k1 = (8.5 +/- 0.3) x 10(-5) s-1 and k-1 = 0.91 +/- 0.06 M-1 s-1, where the model assumed that all the liberated chloride came from 1. When the second aquation step was also taken into account, the rate constants were k1 = (7.9 +/- 0.2) x 10(-5) s-1, k-1 = 1.18 +/- 0.06 M-1 s-1, k2 = (10.6 +/- 3.0) x 10(-4) s-1, k-2 = 1.5 +/- 0.6 M-1 s-1. The rate constants compare favorably with other complexes with the [PtCl(am(m)ine)3]+ moiety and indicate that the equilibrium of all these species favors the chloro form. A pKa value of 5.62 was determined for the diaquated species [[trans-Pt(15NH3)2(H2O)]2(mu-15NH2(CH2)(6)15NH2)]4+ (3) using [1H,15N] HSQC NMR spectroscopy. The speciation profile of 1 and its hydrolysis products under physiological conditions is explored.     

Formation of ammonia in the reactions of a tungsten dinitrogen with ruthenium dihydrogen complexes under mild reaction conditions

Treatment of cis-[W(N2)2(PMe2Ph)4] (5) with an equilibrium mixture of trans-[RuCl(eta 2-H2)(dppp)2]X (3) with pKa = 4.4 and [RuCl(dppp)2]X (4) [X = PF6, BF4, or OTf; dppp = 1,3-bis(diphenylphosphino)propane] containing 10 equiv of the Ru atom based on tungsten in benzene-dichloroethane at 55 degrees C for 24 h under 1 atm of H2 gave NH3 in 45-55% total yields based on tungsten, together with the formation of trans-[RuHCl(dppp)2] (6). Free NH3 in 9-16% yields was observed in the reaction mixture, and further NH3 in 36-45% yields was released after base distillation. Detailed studies on the reaction of 5 with numerous Ru(eta 2-H2) complexes showed that the yield of NH3 produced critically depended upon the pKa value of the employed Ru(eta 2-H2) complexes. When 5 was treated with 10 equiv of trans-[RuCl(eta 2-H2)(dppe)2]X (8) with pKa = 6.0 [X = PF6, BF4, or OTf; dppe = 1,2-bis(diphenylphosphino)ethane] under 1 atm of H2, NH3 was formed in higher yields (up to 79% total yield) compared with the reaction with an equilibrium mixture of 3 and 4. If the pKa value of a Ru(eta 2-H2) complex was increased up to about 10, the yield of NH3 was remarkably decreased. In these reactions, heterolytic cleavage of H2 seems to occur at the Ru center via nucleophilic attack of the coordinated N2 on the coordinated H2 where a proton (H+) is used for the protonation of the coordinated N2 and a hydride (H-) remains at the Ru atom. Treatment of 5, trans-[W(N2)2(PMePh2)4] (14), or trans-[M(N2)2(dppe)2] [M = Mo (1), W (2)] with Ru(eta 2-H2) complexes at room temperature led to isolation of intermediate hydrazido(2-) complexes such as trans-[W(OTf)(NNH2)(PMe2Ph)4]OTf (19), trans-[W(OTf)(NNH2)(PMePh2)4]OTf (20), and trans-[WX(NNH2)(dppe)2]+ [X = OTf (15), F (16)]. The molecular structure of 19 was determined by X-ray analysis. Further ruthenium-assisted protonation of hydrazido(2-) intermediates such as 19 with H2 at 55 degrees C was considered to result in the formation of NH3, concurrent with the generation of W(VI) species. All of the electrons required for the reduction of N2 are provided by the zerovalent tungsten.     

Superoxide radical anion adduct of 5,5-dimethyl-1-pyrroline N-oxide (DMPO). 3. Effect of mildly acidic pH on the thermodynamics and kinetics of adduct formation

The nitrone, 5,5-dimethylpyrroline N-oxide (DMPO), is a commonly used spin trap for the detection of superoxide radical anion (O2*-) using electron paramagnetic resonance spectroscopy. This work investigates the reactivity of DMPO to O2*- in mildly acidic pH (5.0-7.0). Mild acidity is characteristic of acidosis and has been observed in hypoxic systems, e.g., ischemic organs and cancer cells. Although the established pKa for O2*- is 4.8, the pKa for DMPO is unknown. The pKa of the conjugate acid of DMPO was determined to be 6.0 using potentiometric, spectrophotometric, 1H and 13C NMR, and computational methods. 1H and 13C NMR were employed to investigate the site of protonation. An alternative mechanism for the spin trapping of O2*- in mildly acidic pH was proposed, which involves protonation of the oxygen to form the N-hydroxy imino cation and subsequent addition of O2*-. The exoergicity of O2*- addition to protonated DMPO was rationalized using density functional theory (DFT) at the PCM/B3LYP/6-31+G**//B3LYP/6-31G* level of theory.     

Mass spectrometry analysis of in vitro nitration of a recombinant human IgG1 monoclonal antibody

Nitration of a recombinant human monoclonal antibody was carried out in vitro by incubating the antibody with the nitrating reagent tetranitromethane (TNM). The susceptible sites of nitration were identified using high-performance liquid chromatography/mass spectrometry (HPLC/MS). In general, tyrosine residues in the variable domains of the antibody are more susceptible to nitration, while tyrosine residues in the constant domains are relatively resistant to nitration. However, one tyrosine residue in the CH1 domain and one tyrosine residue in the CH2 domain are highly susceptible to nitration. Interestingly, the susceptible tyrosine residue in the CH2 domain is followed by the conserved asparagine residue that is glycosylated.     

Photodynamic Treatment with Benzoporphyrin Derivative Monoacid Ring A Produces Protein Tyrosine Phosphorylation Events and DNA Fragmentation in Murine P815 Cells

Treatment with benzoporphyrin derivative monoacid ring A (BPD-MA, verteporfin) and broad-spectrum fluorescent light rapidly produced apoptosis in murine P815 mastocytoma cells. Fragmentation of DNA, a fundamental characteristic of cells undergoing apoptosis, was evident within 3 h following the photodynamic treatment. Western immunoblot analysis using the specific antiphosphotyrosine monoclonal antibody 4G10 indicated that molecular species of >200 kDa were phosphorylated on tyrosine residues during or immediately following the irradiation of cells loaded with BPD-MA. Increased tyrosine phosphorylation of a 15 kDa protein was evident by 15 min postirradiation. In the absence of light, BPD-MA did not affect the status of cellular protein tyrosine phosphorylation or cause DNA fragmentation. The protein kinase inhibitor staurosporine prevented tyrosine phosphorylation of the >200 kDa species but did not affect tyrosine phosphorylation of the 15 kDa protein or the level of DNA fragmentation produced by the photo-dynamic treatment. The protein tyrosine phosphorylation events observed for P815 cells treated with cytotoxic levels of BPD-MA and light may not be directly related to the induction of the apoptotic cell death pathway.     

UV/vis, 1H, and 13C NMR spectroscopic studies to determine mangiferin pKa values

The acid constants of mangiferin (a natural xanthonoid) in aqueous solution were determined through an UV/vis spectroscopic study employing the SQUAD program as a computational tool. A NMR study complements the pK(a) values assignment and evidences a H-bridge presence on 1-C. The chemical model used was consistent with the experimental data obtained. The pK(a) values determined with this procedure were as follows: H(4)(MGF)=H(3)(MGF)(-)+H(+), pKa1 (6-H)=6.52+/-0.06; H(3)(MGF)(-)=H(2)(MGF)(2-)+H(+), pKa2 (3-H)=7.97+/-0.06; H(2)(MGF)(2-)=H(MGF)(3-)+H(+), pKa3 (7-H)=9.44+/-0.04; H(MGF)(3-)=(MGF)(4-)+H(+), pKa4 (1-H)=12.10+/-0.01; where it has been considered mangiferin C(19)H(18)O(11) as H(4)(MGF). Mangiferin UV/vis spectral behavior, stability study in aqueous solution as well as NMR spectroscopy studies: one-dimensional (1)H,(13)C, 2D correlated (1)H/(13)C performed by (g)-HSQC and (g)-HMBC methods; are also presented. pK(a) values determination of H(4)(MGF) in aqueous solution is a necessary contribution to subsequent pharmacokinetic study, and a step towards the understanding of its biological effects.     

Basicity of some P1 phosphazenes in water and in aqueous surfactant solution

The pKa values in water and in dilute surfactant solution for 15 ring-substituted phenyl P1 pyrrolidino phosphazenes PhN=P(NC4H8)3 and the phenyl P1 dimethylamino phosphazene PhN=P(NMe2)3 previously studied in acetonitrile (AN) and tetrahydrofuran (THF) are reported. The nonionic surfactant Tween 20 was used for the basicity measurements of some compounds to overcome the solubility problems. Measurements with a control group of phosphazenes in both media were used to validate the use of the obtained pKa values as estimates of aqueous values. The pK(a) values of the studied phosphazenes in aqueous medium vary from 6.82 (2,6-dinitro-) to 12.00 (4-dimethylamino-). The basicity span is 5.18 pKa units. The aqueous pKa values of the P1 phosphazenes were correlated with the respective basicity data in AN and THF and from these correlations the pK(a) values in water for the parent compounds HN=P(NC4H8)3 and HN=P(NMe2)3 were estimated as 13.9 and 13.3. Also a comparison of the basicity of phosphazenes and some guanidines, amines and pyridines was made. In water the parent phosphazenes and guanidines are the strongest of all the groups of bases studied. In AN and THF the parent phosphazenes are clearly the strongest bases followed by guanidines, amines and pyridines which are bracketed between the basicities of phenyl phosphazenes. In the gas phase the phosphazenes for which data are available are clearly more basic than the other compounds referred to here. Comparison of the basicity data of P1 phosphazenes and some guanidines confirms earlier conclusions about the partly ylidic character of the N=P double bond.     

Tetrakis- and tris(1-Methyluracil) complexes of Pt(II): formation and properties of a carbon-bonded nucleobase species as well as of heternonuclear derivatives

The reaction of K2PtCl4 with an excess of 1-methyluracilate (1-MeU) in water at 60 degrees C leads to the formation of two major products, K2[Pt(1-MeU-N3)4].10H2O (1) and trans-K[Pt(1-MeU-N3)2(1-MeU-C5)(H2O)].3H2O (2). Addition of CuCl2 to an aqueous solution of 2 yields the mixed-metal complex trans-[PtCl(1-MeU-N3,O4)2(1-MeU-C5,O4)Cu(H2O)].H2O (4). Single-crystal X-ray analysis was carried out for 1 and 4. In both compounds, the heterometals (K+ in 1 and Cu2+ in 4) are bonded to exocyclic oxygens atoms of the 1-MeU ligands, giving rise to intermetallic distances of 3.386(2) and 3.528(2) A in 1 and 2.458(1) A in 4. The shortness of the Pt-Cu separation in 4 is consistent with a dative bond between PtII and CuII. The aqua ligand in 2 is readily substituted by a series of other ligands (e.g., 1-MeC, 9-MeGH, and CN-), as demonstrated by 1H NMR spectroscopy, with 3J(195Pt-1H(6)) coupling constants being sensitive indicators. Acid-base equilibria of 1 and 2 have been studied in detail and reveal some unexpected features: 1 has a relatively high basicity, with protonation starting below pH 5, and first and second pKa values being ca. 3.4 and 0.4, respectively. These pKa values are markedly higher than those of related neutral 2:1 or cationic 1:1 complexes and are attributed to both charge effects (-2 charge of 1) and a favorable stabilization of oxygen-protonated species by the arrangement of four exocyclic oxygen groups of 1-MeU ligands at either sides of the platinum coordination planes. Whereas in 2, H+ affinities of the three uracil ligands are in the normal range, there is a surprisingly low acidity of N(3)H of the C5-bonded uracil with a pKa of approximately 12.2, which compares with 9.75 for free 1-methyluracil. This implies that the C5-bonded PtII does not induce the typical acidifying effect of a PtII metal entity when bonded to a ring nitrogen atom of a neutral nucleobase. Rather, the effect is qualitatively similar to that of a metal ion bonded to N3 of an anionic 1-MeU ligand, which likewise increases its overall basicity as compared to neutral 1-MeUH.     

Synthesis and acid-base properties of (1H-benzimidazol-2-yl-methyl)phosphonate (Bimp2-). Evidence for intramolecular hydrogen-bond formation in aqueous solution between (N-1)H and the phosphonate group

The synthesis of (1H-benzimidazol-2-yl-methyl)phosphonic acid, H2(Bimp)+/-, is described: 2-chloromethylbenzimidazole was reacted with ethylchloroformate to give 1-carboethoxy-2-chloromethylbenzimidazole which was treated with trimethyl phosphite and after hydrolysis with aqueous HBr H2(Bimp)+/- was obtained. In H2(Bimp)+/- one proton is at the N-3 site and the other at the phosphonate group; both acidity constants were determined in aqueous solution by potentiometric pH titrations (25 degrees C; I = 0.1 M, NaNO3) and this furnished the pKa values of 5.37 +/- 0.02 and 7.41 +/- 0.02, respectively. The acidity constant for the release of the primary proton from the P(O)(OH)2 group of H3(Bimp)+ was estimated: pKa = 1.5 +/- 0.2. Moreover, Bimp2- can be further deprotonated at its neutral (N-1/N-3)H site to give the benzimidazolate residue, but this reaction occurs only in strongly alkaline solution (KOH); application of the H_ scale developed by G. Yagil (J. Phys. Chem., 1967, 71, 1034) together with UV spectrophotometric measurements gave pKa = 14.65 +/- 0.12. Comparisons with acidity constants taken from the literature show that this latter pKa value is far too large and this allows the conclusion that an intramolecular hydrogen bond is formed between the (N-1/N-3)H site and the phosphonate group of Bimp2-; the formation degree of this hydrogen-bonded isomer is estimated to be 98 +/- 2%. The general relevance of this and the other results are shortly discussed and the species distribution for the Bimp system in dependence on pH is provided.     

Rationalisation of unusual changes in efficiency and retention with temperature shown for bases in reversed-phase high-performance liquid chromatography at intermediate pH

Despite the possibility of poorer peak shapes, analysis of pharmaceuticals and other bases using reversed-phase high-performance liquid chromatography (RP-HPLC) at intermediate pH gives useful increases in retention, selectivity and column loading capacity compared with low pH. Retention times of some bases showed anomalous increases with column temperature. Peak shapes for bases improved significantly at elevated temperature (up to 70 degrees C), with the weakest bases studied (pKa approximately 8) giving greater improvement than the strongest bases (pKa approximately 10). In contrast, quaternary ammonium compounds showed reduced retention with increasing column temperature (normal behaviour) and only modest improvements in peak shape. Considering these results, and pKa measurements of the bases made using capillary electrophoresis, it appears that increases in retention and improvement in efficiency may be influenced significantly by reduction in the pKa of bases with temperature, leading to reduction in protonation. It is less likely that efficiency improvements are due to the speeding up of the kinetics of silanol ion exchange, at least in the temperature range studied here.