Protonation equilibria of quinolone antibacterials

The acid-base properties of seven antibacterial 7-piperazinyl fluoroquinolone derivatives were studied by potentiometry and UV and NMR spectroscopy. These molecules contain two proton-binding sites of similar basicity, namely, the piperazine amino and the carboxylate groups, as proven by 1H NMR spectroscopy. The basicities are quantitated at the molecular level in terms of macroconstants, and also at the submolecular level in terms of microconstants. The microconstants are then used to calculate the concentration of the positive, zwitterionic, neutral, and negatively charged species (microspeciation). The zwitterionic forms always predominate over their neutral protonation isomers, but the zwitterionic:neutral concentration ratio is considerably different for the examined fluoroquinolone derivatives.     

Physico-chemical profiling of centrally acting molecules for prediction of pharmacokinetic properties

Physico-chemical profiling is a fundamental tool at the early stage of drug discovery in screening drug-like candidates. Complex physico-chemical profiling, including molecular properties such as solubility, ionization, lipophilicity and permeability, has been found to be of predictive power in ADME (absorption, distribution, metabolism, elimination). In the present thesis work, the physico-chemical properties of centrally acting compounds were investigated. We determined the protonation constants (K), the partition coeffitient in octanol/water (Poct) and cyclohexane/water (Pch) systems of antidepressive sertraline and 15 antipsychotic piperidine and piperazine derivatives and calculated the delta logP (logPoct-logPch) values of the molecules. Due to the poor water solubility of the compounds potentiometry using the "co-solvent" technique was applied for the determination of the protonation constants. The logP values were measured by the dual-phase potentiometric titration in octanol/water system and the traditional shake-flask method was used in cyclohexane/water system. Highly precise physico-chemical data were obtained by these validated methods. The relationship between the structure of the molecules and the physico-chemical data was investigated. The pharmacokinetic properties of the compounds were predicted by the physico-chemical parameters. Linear relationship has been found between the brain penetration characterized by the logBB values and the delta logP values. The validity of the equation was controlled by the delta logP and the logBB values of sertraline.     

Microequilibrium of drug molecules]

Protonation macro-, micro-, and submicro constants are physico-chemical parameters of crucial importance, concerning the fate of bio-, drug, and narcotic drug molecules in the body and in protic solvents. The most important characteristics, relationships, applications and biological significance of these parameters are reviewed, using acetylcysteine and cysteine, as examples. The mucolytic effect of acetylcysteine, an active principle in numerous drugs, is interpreted in terms of protonation state of the molecule and its thiolate site. Microscopic protonation constants of acetylcysteine, data that have not previously appeared, are also reported.     

Proton Speciation and Microspeciation of Vinpocetine and Related Compounds in Aqueous and Biomimetic Media

Purpose. The determination of protonation macroconslants of twelve compounds in the vincamine drug family and the determination of protonation microconstants of cis- and trans-apovincaminic acid in media of various solvent composition to characterise their site-specific basicity and to estimate the concentration of the membrane-penetrating and receptor-binding forms. Methods. UV-pH titrations have been used to determine the protonation macroconstants in 10–43 wt% methanol/water mixtures. Yasuda-Shedlovsky extrapolation was applied to obtain aqueous log values for compounds sparingly soluble in water. Protonation microconstants were also determined by deductive methods for compounds of free carboxylic group. Results. In the case of the two water-soluble compounds the extrapolated and the directly measured aqueous log K values were in good agreement, verifying all other extrapolated data. Compounds of cis-D/E ring anellation are 0.4-0.8 log K units more basic than their epimeric,trans counterparts. The pH-dependent distribution of apovincaminic acid microspecies in aqueous and membrane-like media is depicted in microspeciation diagrams. Conclusions. The N(4) nitrogen is more shielded by the adjacent ethyl group in trans-D/E ring anellation eburnanes than in cis ones, as reflected by the protonation constants. Solvent-dependent basicity data predict superiority of trans isomers in lipophilicity and membrane-penetrating ability.     

Integrated plasma and synovial fluid pharmacokinetics of tenoxicam in patients with rheumatoid arthritis and osteoarthritis: factors determining the synovial fluid/plasma distribution ratio

Single oral doses of 40 mg of the nonsteroidal antiinflammatory drug, tenoxicam, were given to four patients (three with rheumatoid arthritis, one with osteoarthritis). The concentrations of the drug in synovial fluid and plasma were measured by a specific high-performance liquid chromatography method. The unbound fractions of the drug in both fluids were determined at pH 7.4 and 37 degrees C by equilibrium dialysis. The possible influence of the pH on the protein binding was also assessed. The total concentration time curves in plasma and synovial fluid were fitted to linear oral 1 and 2 compartment body models with an additional synovial fluid compartment connected to the central compartment. The unbound fractions of drug in synovial fluid and plasma were on average 0.015 and 0.011, respectively: not significantly different from each other. The protein binding of tenoxicam was pH dependent with increased free fractions at pH values less than 7.4. The average peak concentrations of tenoxicam in plasma and synovial fluid were 4.3 and 1.4 micrograms/ml, respectively. The mean ratio of the areas under the total concentration time curves in synovial fluid and plasma was 0.42, which corresponded to the steady state of equilibrium ratio of the total drug concentrations in the two body fluids. Two hypotheses were tested: hypothesis I assuming that equilibration across the synovial tissue takes place between the unbound, unionized tenoxicam molecules; hypothesis II assuming that equilibration across the synovial tissue is established between the unbound (unionized + ionized) tenoxicam molecules. Based on the available evidence hypothesis II was rejected.     

Characterization of potential NMDA and cholecystokinin antagonists. II. Lipophilicity studies on 2-methyl-4-oxo-3H-quinazoline-3-alkyl-carboxylic acid derivatives

The lipophilicity of 17 newly synthesized potential NMDA and cholecystokinin antagonist 2-methyl-4-oxo-3H-quinazoline-3-alkyl-carboxylic acid derivatives has been investigated. The apparent partition coefficients of two amphoteric compounds of overlapping protonation (Q1 and Q2) were determined by shake-flask method and converted into true log P values using the protonation microconstants. The difference between their lipophilicity expressed with the true partition coefficients was less, than it could be expected from the 2D structures and was explained with conformational preferences and formation of intramolecular interactions. Out of the other 15 monoprotic quinazolone compounds the lipophilicity of ten molecules (Q8-Q17, experimental set) was determined by TLC method with the help of a calibration set consisting of 12 standard molecules, five quinazolones (Q3-Q7) and seven pyrido[1,2-a]pyrimidines (PP1-PP7). In order to justify the suitability of pyrido-pyrimidines as standards for the chromatographic log P determination of quinazolones, first Q3-Q7 were examined by TLC and HPLC using PP1-PP7 for calibration. Data showed good agreement of results obtained by shake-flask and two different chromatographic methods indicating the similar chromatographic behavior of the two bicyclic systems and the relevance of PP1-PP7 to extend the calibration set of quinazolones. The obtained log P values proved mostly the expected structure-activity relationships. Some findings, however, have revealed specific partition behavior of the compounds providing useful information in the estimation of their pharmacokinetics, and these are discussed in the paper.     

Relationship between lipophilicity and absorption from the liver surface of paraben derivatives and antipyrine in rats

Objectives The importance of drug lipophilicity on absorption from the liver surface was examined in rats using paraben derivatives, antipyrine, Sudan III, and Sudan blue. Methods The log partition coefficient (PC) of n‐octanol/water ranged from ?1.39 to 4.62. The compounds were applied to the rat liver surface using a cylindrical diffusion cell (i.d. 9 mm). Key findings The rate of absorption at 15 min was calculated to be 13.9% for paraben, much lower than that for its derivatives methylparaben, propylparaben and butylparaben (～80%). The obtained first‐order absorption rate constant (k_a) of paraben, methylparaben, propylparaben and antipyrine increased according to lipophilicity. Further lipophilicity resulted in a fall in k_a, implying the importance of affinity for lipids and water in absorption from the liver surface. Thus, a compound with a log PC of around 2.5 is considered to have maximum absorbability from the rat liver surface. A good relationship (r² = 0.97) was recognized between the log k_a and log reciprocal value with the square root of molecular weight of the compounds with a log PC below 2.5. Conclusions The rate of absorption of a drug from the liver surface could be estimated from physicochemical properties such as lipophilicity and molecular weight.     

Quantitative structure-antimicrobial activity relationship in 5 beta-cholanyl-24-benzylamine derivatives

Some representative physicochemical properties of benzylamido and amino derivatives of common bile acids have been determined and correlated with their antimicrobial activity against gram-positive bacterial strains. Steroid hydroxyls do not affect the basicity of amino derivatives; they promote solubility in a parallel way to unconjugated bile acids and mainly control hydrophobicity of this class of compounds as measured by log P values. Activity was correlated to hydrophobicity; that is, the nature of the side chain modulated activity, affected basicity, and facilitated changes in partition ability. Benzylamino derivatives proved to be even more active than the corresponding amides when ionization is taken into account. Trihydroxy derivatives possess the lowest log P values and were practically inactive. Decreased activity was also observed in those cases where, due to the orientation of the hydroxy group in the 6 or 7 position, the back beta face of the molecule had a reduced hydrophobic surface area. Antimicrobial activity, in terms of -log MIC (minimal inhibitory concentration), was found to correlate linearly with log P values of uncharged species. This linear relationship is discussed with respect to the structure of the steroid moiety and the ability of these molecules to cross cellular membranes.     

Chemical reactivity studies with naphthoquinones from Tabebuia with anti-trypanosomal efficacy

The biological activities of the naphthoquinones lapachol and its cyclization product beta-lapachone, extracted from trees of the genus Tabebuia, have been intensively studied. Given continuity to the studies about heterocyclic derivatives obtained from the reaction of these naphtoquinones with amino-containing reagents, 22 derivatives of beta-lapachone, nor-beta-lapachone and lapachol were synthesised and their activities against trypomastigote forms of T. cruzi were evaluated. The compounds were grouped as oxazolic, imidazolic, phenoxazinic, indolic, pyranic and cyclopentenic derivatives. The variability of the new structures is based on the great electrophilicity of 1,2-quinoidal carbonyls towards reagents containing nitrogen or carbon as nucleophilic centres. In relation to the trypanocidal activity of the synthesised compounds, in view of their structural diversity, tendencies only could be verified. Among the cyclofunctionalised products the oxazolic and imidazolic derivatives showed +/- 1.5 to 34.8 times higher activity than crystal violet, the standard drug for the sterilization of stored blood. These results corroborate the tendency of trypanocidal activity in imidazolic skeletons, and indicate that this moiety could be used as a guide for architectural delineation of molecules with potential value for the chemotherapy of Chagas disease.     

Electrodeless, accurate pH determination in highly basic media using a new set of (1)H NMR pH indicators

A set of indicator molecules was selected and applied to elaborate an NMR-based pH determination method, free of glass electrode errors in highly basic media. Accurate measurement of pH values and protonation constants was achieved by a successive build-up of overlapping, increasingly high pH solutions, using a collection of 8 compounds of appropriately incremented basicities. In order to verify the method, acid-base properties were quantified for two compounds with very high basicities in conflicting reports: two pharmaceutically important biguanidine drugs, metformin and phenformin.     

Synthesis and investigation of protonation properties and lipophilicity of some new quinazolone-3-carboxylic acid derivatives]

Several new quinazolone-carboxylic acid derivatives as potential NMDA and AMPA receptor antagonists have been synthesized, and the protonation properties and lipophilicity of some representative molecules have also been studied. The protonation macroconstants (logK) of 4(3H)-quinazolone (Q0) and two 2-methyl-4-oxo-3H-quinazoline-3-carboxylic acids (Q1, Q2) were determined by pH-potentiometry. The acid-base chemistry of Q1 and Q2, where protein-bindings take place in an overlapping fashion, was described in terms of protonation microconstants (logk) as well. Microspeciation was carried out by UV-pH titration and deductive method. Microspeciation revealed remarkable differences between the two homologue compounds (Q1 and Q2), namely insertion of a second methylene moiety into the aliphatic acid side-chain reversed the predominantly zwitterion-involved protonation pathway into neutral form-involved one. Lipophilicity of our molecules was described by the octanol-water partition coefficients. The apparent partition coefficients of Q1 and Q2 were determined by shake-flask method and converted into true logP values using the protonation microconstants. The unexpected differences between their true logP values were explained, similarly to the different protonation pathways with conformational preferences and formation of intramolecular interactions. Out of the other 15 monoprotic quinazolone compounds the lipophilicity of 10 molecules (Q8-Q17, experimental set) was determined by RP-TLC method with the help of a calibration set consisting of 12 standard molecules, five quinazolones (Q3-Q7, determined by shake-flask method) and seven pyrido[1,2a]pyrimidines (PP1-PP7). The obtained logP values proved mostly the expected structure-property relationships. These physico-chemical investigations are pieces of predictive information for the pharmacokinetics of our compounds. These are also discussed in the paper.     

Biochemical and pharmacological characterization of isatin and its derivatives: from structure to activity

Isatin, 1H-indole-2,3-dione, is a heterocyclic compound of significant importance in medicinal chemistry. It is a synthetically versatile molecule, a precursor for a large number of pharmacologically active compounds. Isatin and its derivatives have aroused great attention in recent years due to their wide variety of biological activities, relevant to application as insecticides and fungicides and in a broad range of drug therapies, including anticancer drugs, antibiotics and antidepressants. The purpose of this review is to provide an overview of the pharmacological activities of isatin and its synthetic and natural derivatives. Molecular modifications to tailor the properties of isatin and its derivatives are also discussed.     

Soft ligands--biodegradable complexants in plant cultivation and environmental protection

In this paper, the place and role of biodegradable complexants in light of modern plant cultivation criteria is discussed. From this point of view, micro- and macroelement (metal) chelators play a remarkable role. They are widely used in a variety of consumer products and processes, especially in agriculture. The great majority of the traditional chelators (e.g., EDTA) are, however, essentially non-biodegradable. We have systematically investigated some new biodegradable ligands with polyaminopolycarboxylic structure. We conclude that these compounds may be considered as a special group of the soft (safer) chemicals proposed first by Bodor based on retrometabolic concepts. We studied the structural conditions of biodegradability, especially regarding the new ligands, by the physicochemical characterization of their complexes formed with micro- and macroelements. The study of the complicated equilibrium conditions required the prior determination of the stability constants of the metal aqua/hydroxo complexes and the protonation constants of the ligands as well. Knowledge of the pH-dependence of the proper competitive equilibriums allowed the optimization of the investigated complexation reactions. The protonation and stability constants obtained were compared with the adequate values of the traditional, but weakly biodegradable EDTA. Excellent or very good linear correlations have been found between the logarithms of the different ligands' protonation constants, but the slopes of the straight lines were different and generally smaller than EDTA's corresponding values. Presumably, the log stability constants (logK(st)) of complexes formed by different ligands (X(m), X(n)) having analogue structures with a given metal-ion series (M(i), i = 1, 2, ...) are linearly correlated [i.e., log K(X(m)M(i)) = a log K(X(n)M(i)) + b], just as the stability constants of complexes formed by different ions (M(k), M(l)) with a given ligand series (X(j), j = 1, 2, ...). Based on the rich data basis of EDTA, these regression equations allow the calculation of some still unmeasured stability constants of the new biodegradable complexes. In the analysis of structural effects we would like to especially highlight the role of stereochemical relations in addition to the usual structural elements. From this point of view, the eventual presence of chiral nitrogen atoms is of particular importance. The stability of these compounds increases in their complexed form. As a special research field, we have examined the possibility of obtaining metal ion buffers. These systems may get an important role both from theoretical and practical points of view.     

Relationships between the lipophilicity of some 1,4-piperazine derivatives of aryloxyaminopropanols and their beta-andrenolytic activity

Nineteen 1,4-piperazine derivatives of aryloxyaminopropanol were evaluated with respect to beta-adrenolytic activity. The retention factors obtained from HPLC, RM values obtained from partition TLC and the lipophilic Hansch's (4) constants pi were determined and the compounds were studied with respect to their lipophilicity based on chromatographic properties. The study of the influence of different substituents introduced at the para position on the phenyl ring on the retention factor indicated the log k vs. the number of carbon atoms in R1 substituent to be a linear relationship. Attempts have been made to relate the beta-adrenolytic activity to the lipohydrophilic parameters by deriving a quantitative relationship between them. Significant parabolic correlation was observed between the beta-adrenolytic activity and the logarithm of the retention factor, log k. An analogous relationship was obtained between the beta-adrenolytic activity of the compounds and the RM values obtained from partition TLC as well as Hansch's lipophilic constants pi.     

Theoretical study of the protonation and tautomerization of adenosine, formycin, and their 2-NH2 and 2-F derivatives: functional implications in the mechanism of reaction of adenosine deaminase

A quantum chemical study of adenosine, formycin, and their 2-NH2 and 2-F derivatives is performed. The tautomerism of neutral and protonated species as well as the protonation of adenosine, formycin, and their derivatives are theoretically studied using semiempirical MNDO and AM1, as well as ab initio STO-3G methods. Calculations have been performed on a reduced model, in which the ribose moiety has been substituted by a hydroxy-methyl group. Results indicate that adenosine is mainly protonated at the N1 atom, whereas formycin can be protonated on N1 or N3, depending on the tautomeric form (N8-H or N7-H). The quantum chemical study of the N1-protonated molecules shows that a second protonation of adenosine is mainly on the N3 atom, whereas formycin can be protonated on N8 or N3, depending on the tautomeric form. On the other hand, results indicate that the protonation of formycin and its derivatives at the N1 atom leads to a change in their tautomeric preference from N7-H to N8-H. The importance of both tautomerism and protonation reactions in the mechanism of action of adenosine deaminase is studied by means of a quantitative structure activity relationships strategy. Significant correlations were found between several electronic parameters and the logarithm of the maximum rate of deamination (log Vm) of the studied compounds. For formycin and its derivatives, it was necessary to consider their N8-H tautomeric forms. The electronic parameters giving good correlations were as follows: energy of the minimum of the ab initio molecular electrostatic potential on N1, net charge over purine (pyrazolo-pyrimidine) and pyrimidine rings, and the N1 protonation energy. It must be noted that all these parameters are informative in relation to a proton attack. Adenosine and purine ribosides have been studied largely because of their high biological relevance. They are constituents of nucleic acids, intermediates in secondary metabolism, neuromodulators, and neurohormones. Their analogues have been extensively used because of their wide range of pharmacological effects (1). Formycin A (Fig. 1) is one of the most studied analogues of adenosine. It is a natural product extracted from Nocardia interforma (2) with proven antiviral (3-5), antibiotic (2), immunodepressant (6), antitumor (6), and antimetabolic (5) activities.     

The binding of oxicam derivatives to human serum albumin

The binding characteristics of several oxicam derivatives (tenoxicam, 4-hydroxy-2-methyl-N-phenyl-2H-1,2-benzothiazixine-3- carboxamide-1,1-dioxide (CP 14,304), 4-hydroxy-2-methyl-N-2-(3-methyl)-pyridyl-2H-1,2-benzothiazixine-3 -carboxamide- 1,1-dioxide (CP 16,460), piroxicam, meloxicam [corrected], isoxicam, 5-hydroxy-piroxicam) to 2% and 4% human serum albumin (HSA) were determined using a modified ultrafiltration process. The binding properties to HSA were characterized by determining the overall binding constant, the apparent binding constant, the slope, the free reaction energy, and the unbound portion of the drug. The following results were obtained: 1. These oxicam derivatives show a high affinity to HSA. The unbound fraction amounts to 1-3%. 2. The affinity of the compounds to HSA decreases in the order mentioned above. 3. Doubling of the HSA concentration reduces the unbound fraction, to the half, with piroxicam being the only exception.     

Synthesis, physicochemical and pharmacological properties of pentacyclic alkaloid-analogues

Quinazolinocarboline rutaecarpine and evodiamine (Evodia rutaecarpa) are main alkaloid components of traditional Chinese folk-remedies. Evodiamine exhibited selective antitumor and antimetastatic effects on several cancer cell lines and became lead structure of anticancer agents. During our synthetic research we achieved to gain alkaloid hybrid derivatives by combining the structural elements of quinazolinocarbolines with analogous alkaloids or drug molecules having similar effects by bioisosteric replacements. 8-norrutaecarpine, a hybrid molecule of rutaecarpine and luotonin A containing the indolo-pyrroloquinazolinone ring system has been synthesized. The hybrids of rutaecarpine and piroxicam bearing the indolo-pyridobenzothiazine and the 12-azaindolo-pyridobenzothiazine structures were prepared on two alternative routes. Two new heterocondensed pentacyclic compounds, 5-sulfarutaecarpine and 5-sulfa-8-norrutaecarpine were reached via bioisosteric replacement on the structure of rutaecarpine and 8-norrutaecarpine. Two new tricyclic ring systems, pyrido-benzothiadiazine and pyrrolo-benzothiadiazine were produced as intermediaries of these pentacyclic molecules. Series of substituted derivatives were prepared for pharmacological studies by modification of the structures with various substituents and solubilizing groups. During our work alternative way for synthesis of nauclefine (Nauclea latifolia) was laboured, and we published the synthesis of indolylquinazolinone derivative bouchardatine (Bouchardata neurococca) for the first time. Some of the physicochemical attributes of the synthesized intermediaries were defined, such as the pKa constants of 2,3-poly-methylene-benzothiadiazines. Proton/deuteron exchange kinetic constants of active methylene-groups of five tricyclic compounds were measured by 1H NMR technique. Solvent-dependent ratio of the Z/E isomers of phenyhydrazone-derivatives in polar and apolar solvents were determined. In the case of 18 produced compounds our work was completed by in vitro pharmacological studies performed within co-operation with the Institute of Pharmacology. The viability of HeLa cells was inhibited by five of our compounds to similar extent as the effect of evodiamine. Eight of our compounds induced apoptosis on HeLa cells to similar extent as evodiamine.     

A comprehensive method for determining hydrophobicity constants by reversed-phase high-performance liquid chromatography

The development of a method for determining hydrophobicity constants for small, organic molecules by reversed-phase liquid chromatography (RPLC) is presented. The method uses capacity ratios measured at a number of different compositions of methanol to obtain derived values, denoted log k'w, upon which a new scale of hydrophobicity constants can be developed. This scale eliminates potential problems such as peak inversion that hamper RPLC methods using isocratic data to estimate hydrophobicity. The differential hydrogen bond effect observed in most correlations of RPLC data with logarithms of octanol-water partition coefficients (log Po/w) for compounds of opposite net hydrogen bonding capabilities (noncongeners) was minimized by adding trace quantities of n-decylamine and 1-octanol to the eluent and using an octyl-modified silica gel stationary phase. Values of log k'w are shown to be largely column-independent as long as the hydrophobic properties of columns are similar. The correlation of log k'w values with the logarithms of bovine serum albumin binding constants (log 1/C) is shown to be statistically indistinguishable from the correlation of log 1/C with log Po/w, indicating that this data models log 1/C as well as log Po/w for these compounds. Additionally, the chromatographic system is automatable and thus capable of higher sample throughput than measurements of log Po/w by the shake-flask method.