Amino acid-amino acid interactions studied by charge-transfer chromatography

The interaction between amino acids was studied by charge-transfer reversed-phase thin-layer chromatography. The dependence of the lipophilicity of Trp on the concentration of other amino acids in the eluent was considered to be linearly related to the strength of interaction. Arg, Asn, Glu, Met, Phe and Thr interacted with Trp; Ala, Gly and Ser showed no interaction. Stepwise regression analysis indicated that the pK value of the amino acid side-chain and the lipophilicity of the amino acid had the greatest impact on the interaction, suggesting the simultaneous presence of weak hydrophilic and hydrophobic bonding forces between amino acids. Sodium acetate in the eluent increased the interactive strength between Phe and Trp; acetic acid and sodium chloride did not influence the interaction significantly. No significant difference was found between the effects of l- and d-Asn.     

Influence of salt and pH on the hydrophobicity parameters of antisense nucleosides studied by reversed-phase thin-layer chromatography and multivariate mathematical-statistical methods.

The lipophilicity and specific hydrophobic surface area of 12 8-substituted 2'-deoxyadenosine and 17 5-substituted-2'-deoxyuridine derivatives were determined by reversed-phase thin-layer chromatography in ion-free eluents and in eluents containing sodium chloride, sodium acetate and acetic acid. The strength and selectivity of the effect of eluent additives were separated by use of spectral mapping technique followed by two-dimensional nonlinear mapping. The relationship between the structural characteristics and hydrophobicity parameters was elucidated by stepwise regression analysis. Eluent additives exert a considerable influence on both hydrophobicity parameters. The effect of sodium chloride and acetic acid was higher than that of sodium acetate. The strength and selectivity of the sensitivity of nucleosides towards eluent additives significantly depended on the character of the ring structure and on the length of the apolar alkyl chain. The influence of the degree of unsaturation and the branching of the alkyl substituent was negligible.     

Effect of molecular parameters on the binding of phenoxyacetic acid derivatives to albumins

The interaction of 12 phenoxyacetic acid derivatives with human and serum albumin as well as with egg albumin was studied by charge-transfer reversed-phase (RP) thin-layer chromatography (TLC) and the relative strength of interaction was calculated. Each phenoxyacetic acid derivative interacted with human and bovine serum albumins whereas no interaction was observed with egg albumin. Stepwise regression analysis proved that the lipophilicity of the derivatives exert a significant impact on their capacity to bind to serum albumins. This result supports the hypothesis that the binding of phenoxyacetic acid derivatives to albumins may involve hydrophobic forces occurring between the corresponding apolar substructures of these derivatives and the amino acid side chains.     

Synthesis and neuropharmacological characterization of 2-O-substituted apomorphines

We have synthesized novel 2-O-substituted apomorphines with both different lengths of lipophilic alkyl chains and alkyl chains carrying free hydroxyl groups. Two bis-apomorphines formed as side products of the reactions with diols were isolated and characterized as well. The neuropharmacological profile of all these new compounds were investigated with respect to their binding affinities and activities to dopamine D(2) and D(1) receptors. The obtained data pointed to the fact that, in the examination of dopaminergic activities of 2-substituted apomorphines, the lipophilicity of the substituent is more important than its spatial parameters.     

The binding of amino acids to the herbicide 2,4-dichlorophenoxy acetic acid

Summary. The interaction of amino acids with the herbicide 2,4-dichlorophenoxyacetic acid (2,4-D) was studied by charge-transfer chromatography carried out on diatomaceous layers covered with different amount of 2,4-D and the effect of salts on the strength of interaction was elucidated. It was established that Arg, His, Lys, Orn, Phe and Trp binds to 2,4-D, the binding process is of saturation character. Principal component analysis proved that the concentration of 2,4-D exerts the highest impact on the interaction and the effect of salts is of secondary importance. The results suggest that these amino acid residues may account for the binding of 2,4-D to proteins and can play a considerable role in the detoxification processes by forming conjugates with 2,4-D. Received April 10, 1998, Accepted September 15, 1998     

Inhibition of nucleoside transport by new analogues of nitrobenzylthioinosine

Nitrobenzylthioinosine (NBTI, 1) was systematically modified by attachment of substituents at positions C6 and N9, and also by substitution of N1 with C. These modifications were chosen to reduce the polarity of the new compounds. Incorporation of the nitro functionality into a benzoxadiazole ring system was considered first. These new nucleosides showed high affinity (1.5-10nM) towards the nucleoside transport protein as present on human erythrocyte ghosts. Next, modification of this benzoxadiazole ring system with C, S and O in different positions produced a number of less polar nucleosides with affinity in the higher nanomolar range. Modification of N9 was achieved with different alkyl and alcohol substituents. An n-butyl substituent proved best, although all variations yielded substantial decreases in affinity. Replacement of N1 by a carbon atom in combination with a 2-Cl substituent also resulted in a relatively potent NBTI derivative (47 nM).     

Effect of Molecular Characteristics on Cellular Uptake,Subcellular Localization,and Phototoxicity of Zn(II) N-Alkylpyridylporphyrins

Tetra-cationic Zn(II) meso-tetrakis(N-alkylpyridinium-2 (or -3 or -4)-yl)porphyrins (ZnPs) with progressively increased lipophilicity were synthesized to investigate how the tri-dimensional shape and lipophilicity of the photosensitizer (PS) affect cellular uptake, subcellular distribution, and photodynamic efficacy. The effect of the tri-dimensional shape of the molecule was studied by shifting the N-alkyl substituent attached to the pyridyl nitrogen from ortho to meta and para positions. Progressive increase of lipophilicity from shorter hydrophilic (methyl) to longer amphiphilic (hexyl) alkyl chains increased the phototoxicity of the ZnP PSs. PS efficacy was also increased for all derivatives when the alkyl substituents were shifted from ortho to meta, and from meta to para positions. Both cellular uptake and subcellular distribution of the PSs were affected by the lipophilicity and the position of the alkyl chains on the periphery of the porphyrin ring. Whereas the hydrophilic ZnPs demonstrated mostly lysosomal distribution, the amphiphilic hexyl derivatives were associated with mitochondria, endoplasmic reticulum, and plasma membrane. A comparison of hexyl isomers revealed that cellular uptake and partition into membranes followed the order para > meta > ortho. Varying the position and length of the alkyl substituents affects (i) the exposure of cationic charges for electrostatic interactions with anionic biomolecules and (ii) the lipophilicity of the molecule. The charge, lipophilicity, and the tri-dimensional shape of the PS are the major factors that determine cellular uptake, subcellular distribution, and as a consequence, the phototoxicity of the PSs.     

Synthesis, study of 3D structures, and pharmacological activities of lipophilic nitroimidazolyl-1,4-dihydropyridines as calcium channel antagonist

QSAR studies indicated that the potency of nifedipine analogues was dependent upon lipophilicity, an electronic term and separated terms for each position on the DHP ring. Changes in the substitution pattern at the C3, C4, and C5 positions of DHPs alter potency, tissue selectivity, and the conformation of the 1,4-DHP ring. In this project a group of alkyl ester analogues of new derivatives of nifedipine, in which the ortho-nitrophenyl group at position 4 is replaced by a 1-methyl-5-nitro-2-imidazolyl substituent, and the methyl group at position 6 is replaced by a phenyl substituent, were synthesized and evaluated as calcium channel antagonist using the high K+ contraction of guinea-pig ileal longitudinal smooth muscle. The results for asymmetrical esters showed that lengthening of the substituent in C3 ester substituent increased activity. When increasing of the length is accompanied by increasing the hindrance, the activity decreased. The results demonstrate that all compounds were more active or similar in effect to that of the reference drug nifedipine.     

Structural optimization and evaluation of butenolides as potent antifouling agents: modification of the side chain affects the biological activities of compounds

A recent global ban on the use of organotin compounds as antifouling agents has increased the need for safe and effective antifouling compounds. In this study, a series of new butenolide derivatives with various amine side chains was synthesized and evaluated for their anti-larval settlement activities in the barnacle, Balanus amphitrite. Side chain modification of butenolide resulted in butenolides 3c-3d, which possessed desirable physico-chemical properties and demonstrated highly effective non-toxic anti-larval settlement efficacy. A structure-activity relationship analysis revealed that varying the alkyl side chain had a notable effect on anti-larval settlement activity and that seven to eight carbon alkyl side chains with a tert-butyloxycarbonyl (Boc) substituent on an amine terminal were optimal in terms of bioactivity. Analysis of the physico-chemical profile of butenolide analogues indicated that lipophilicity is a very important physico-chemical parameter contributing to bioactivity.     

Reversed-phase thin-layer chromatography used to study the simultaneous effect of pH and ion strength on the lipophilicity of chlorhexidine

The effect of ion strength and pH value of the eluent on the determination of the lipophilicity of chlorhexidine was studied by reversed-phase thin-layer chromatography. The method has been improved by using various buffers: aqueous solutions of formic, acetic and propionic acids and their sodium salts in different ratios and in various concentrations. Stepwise regression analysis separated the effect of pH value, ion strength and acid type on the lipophilicity of chlorhexidine and proved that the ion strength exerted a higher impact than the pH value did. The effect of alkyl chain length of the acids was of secondary importance.     

Structure-distribution studies on some 99mTc-o-hydroxybenzyliminodiacetic acid complexes

Biodistributions of a series of thirteen ^99mTc-o-hydroxy-benzyliminodiacetic acid complexes were carried out in rats and their hepatobiliary and urinary outputs correlated with lipophilicity, molecular weight, influence of substituent and plasma protein binding. Hepatobiliary output was moderate for those ligands with large alkyl substituents [t-butyl (36%), and iso-octyl (42%)] but compared to HIDA compounds was relatively low, indicating that they would not be suitable for clinical use. Halogen substituents had only a small effect on increasing hepatobiliary output but a large effect on reducing the urinary clearance.     

Efficient synthesis and in vitro cytostatic activity of 4-substituted triazolyl-nucleosides

We report herein an efficient synthesis of 4-substituted triazolyl-nucleosides and their in vitro cytostatic activity. The synthesis is based on a straightforward 1,3-dipolar cycloaddition between 1-azido-ribose 2 and terminal alkynes under a cooperative effect of microwave activation and copper (I) catalysis. All cycloadducts were obtained in nearly quantitative yield after a short reaction time (1 to 2min). After removal of acetyl protecting groups, the free nucleosides were evaluated against L1210, Molt4/C8, and CEM tumor cell lines. Structure-activity relationship study shows that the substituent on the triazole ring has a major effect since nucleosides 4c and 4g, containing, respectively, a long alkyl chain and an aryl donor group are the most active compounds in this series.     

Discovery of pyrrole-based hepatoselective ligands as potent inhibitors of HMG-CoA reductase

In an effort to identify hepatoselective inhibitors of HMG-CoA reductase, two series of pyrroles were synthesized and evaluated. Efforts were made to modify (3R,5R)-7-[3-(4-fluorophenyl)-1-isopropyl-4-phenyl-5-phenylcarbamoyl-1H-pyrrol-2-yl]-3,5-dihydroxy-heptanoic acid sodium salt 30 in order to reduce its lipophilicity and therefore increase hepatoselectivity. Two strategies that were explored were replacement of the lipophilic 3-phenyl substituent with either a polar function (pyridyl series) or with lower alkyl substituents (lower alkyl series) and attachment of additional polar moieties at the 2-position of the pyrrole ring. One compound was identified to be both highly hepatoselective and active in vivo. We report the discovery, synthesis, and optimization of substituted pyrrole-based hepatoselective ligands as potent inhibitors of HMG-CoA reductase for reducing low density lipoprotein cholesterol (LDL-c) in the treatment of hypercholesterolemia.     

Influence of substituent groups in regioselective acylation of nucleosides by Novozym 435 lipase

A comparative study on the substrate recognition was conducted successfully in Novozym 435-mediated acylation of various 2′- or 5-substituted nucleosides with acyl donors carrying different aliphatic chain lengths (C6, C10, and C14). The unexpected results revealed that the physicochemical property of the substituents (such as the size, hydrophobicity, and substitutional position) in nucleosides profoundly influenced the behavior of the enzyme. The different substrate-binding patterns derived from the existence of the substituents in 2′- or 5-position of the nucleosides could account for this. Moreover, another possible factor governing the regioselectivity might be ascribed to the interaction between the substituent and acyl donor in addition to the geometrical configuration of the lipase's active site.     

Histidine-aromatic interactions in barnase. Elevation of histidine pKa and contribution to protein stability.

Aromatic side-chains are found in the vicinity of histidine residues in many proteins and protein complexes. We have studied the interaction between a histidine residue (His18) and aromatic residues at position 94 in barnase. Three different techniques have been applied to show that Trp94 interacts more strongly with the protonated form of His18. The aromatic-histidine interaction stabilizes the protonated form of histidine by 0.8 to 1 kcal mol-1 relative to the unprotonated and, thereby, increases its pKa value. This was shown indirectly from the pH dependence of the stability of the wild-type protein and the mutant Trp94----Leu; and directly from the difference in pKa of His18 between wild-type barnase and the same mutant protein, and from double-mutant cycles that measure the total interaction energy of Trp94 with His18 at both low and high pH. When Trp94 is replaced by other aromatic amino acids, the strength of the interaction decreases in the series His-Trp greater than His-Tyr greater than His-Phe. The interaction is not masked by high salt concentrations. The raising of the pKa value of His18 by interaction with Trp94 is shown to be consistent with solution studies with model compounds. The histidine-aromatic interaction could have implications in binding and catalysis for modulation of the histidine pKa value.     

Synthesis, acute toxicities, and antitumor effects of novel 9-substituted beta-carboline derivatives

A series of novel 9-substituted beta-carboline derivatives was synthesized from harmine and l-tryptophan, respectively. Cytotoxic activities of these compounds in vitro were investigated. The results showed that most compounds of 9-substituted beta-carboline derivatives had more remarkable cytotoxic activities in vitro than their corresponding parent compounds. Acute toxicities and antitumor effects of the selected beta-carboline derivatives in mice were also examined. The results demonstrated that a short alkyl or benzyl substituent at position-9 increased the antitumor activities significantly and a ethoxycarbonyl or carboxyl substituent at position-3 reduced the acute toxicity and neurotoxicity of these beta-carboline derivatives dramatically. Moreover the compounds both with an alkoxycarbonyl or carboxyl substituent at position-3 and a short alkyl or benzyl substituent at positon-9 exhibited more significant antitumor activities and lower acute toxicities and neurotoxicities than the other compounds. The compound 8c, having an n-butyl and a carboxyl substituent at position-9 and 3, respectively, was found to have the highest antitumor effect and the lowest acute toxicity and neurotoxicity. These data suggested that (1) appropriate substituents at both position-9 and 3 of beta-carboline derivatives might play a crucial role in determining their enhanced antitumor activities and decreased acute toxicities and neurotoxic effects; (2) the beta-carboline derivatives have the potential to be used as antitumor drug leads.     

5-alkylvinyl-1,2,4-triazole nucleosides: Synthesis and biological evaluation

Abstract

Some 5-substituted ribavirin analogues have a high antiviral and anticancer activity, but their mechanisms of action are obviously not the same as their parent compound. The SAR studies performed on 3 (5)-substituted 1,2,4-triazole nucleosides have shown a high dependency between the structure of the 3 (5)-substituent and the level of antiviral/anticancer activity. The most active substances of the row contain coplanar with the 1,2,4-triazole ring aromatic substituent which is connected by a rigid ethynyl bond. However, the compounds with the trans-vinyl linker also had antiviral activity. We decided to study the antitumor activity of ribavirin analogues with alkyl/aryl vinyl substituents in the 5th position of the 1,2,4-triazole ring. Protected nucleoside analogues with various 5-alkylvinyl substituents were obtained by Horner-Wadsworth-Emmons reaction from the common precursor and converted to the nucleosides. Arylvinyl nucleosides were synthesised according the reported procedures. All compounds did not show significant antiproliferative activity on several tumour cell lines. Coplanar aromatic motif in the 5-substituent for the anticancer activity manifestation was confirmed.     

Design, efficient synthesis, and anti-HIV activity of 4'-C-cyano- and 4'-C-ethynyl-2'-deoxy purine nucleosides

Some 4'-C-ethynyl-2'-deoxy purine nucleosides showed the most potent anti-HIV activity among the series of 4'-C-substituted 2'-deoxynucleosides whose 4'-C-substituents were methyl, ethyl, ethynyl and so on. Our hypothesis is that the smaller the substituent at the C-4' position they have, the more acceptable biological activity they show. Thus, 4'-C-cyano-2'-deoxy purine nucleosides, whose substituent is smaller than the ethynyl group, will have more potent antiviral activity. To prove our hypothesis, we planned to develop an efficient synthesis of 4'-C-cyano-2'-deoxy purine nucleosides (4'-CNdNs) and 4'-C-ethynyl-2'-deoxy purine nucleosides (4'-EdNs). Consequently, we succeeded in developing an efficient synthesis of six 2'-deoxy purine nucleosides bearing either a cyano or an ethynyl group at the C-4' position of the sugar moiety from 2'-deoxyadenosine and 2,6-diaminopurine 2'-deoxyriboside. Unfortunately, 4'-C-cyano derivatives showed lower activity against HIV-1, and two 4'-C-ethynyl derivatives suggested high toxicity in vivo.     

Interaction of liposomes with homologous series of fluorescent berberine derivatives: New cationic probes for measuring membrane potential

The interaction of liposomes with a series of fluorescent berberine derivatives having different alkyl chain lengths has been investigated. The hydrophobicity of the binding site on the phospholipid membrane increases and mobility decreases with the length of the alkyl chain. If lauryl sulphate micelles are used to bind berberines, the hydrophobicity of the binding site is the same for all derivatives. The dye series represents a model with constant charge and growing lipophilicity. Both electrostatic forces and lipophilicity play an important role in binding. By virtue of the excellent sensitivity of the dyes to medium polarity, berberines prove to be suitable probes for measuring membrane potential, but only in cases when a negative charge is generated in the liposomal interior. The fluorescence response is a linear function of the potential magnitude.