Interactions of Sea Anemone Toxins with Insect Sodium Channel—Insights from Electrophysiology and Molecular Docking Studies

Animal venoms are considered as a promising source of new drugs. Sea anemones release polypeptides that affect electrical activity of neurons of their prey. Voltage dependent sodium (Nav) channels are the common targets of Av1, Av2, and Av3 toxins from Anemonia viridis and CgNa from Condylactis gigantea. The toxins bind to the extracellular side of a channel and slow its fast inactivation, but molecular details of the binding modes are not known. Electrophysiological measurements on Periplaneta americana neuronal preparation revealed differences in potency of these toxins to increase nerve activity. Av1 and CgNa exhibit the strongest effects, while Av2 the weakest effect. Extensive molecular docking using a modern SMINA computer method revealed only partial overlap among the sets of toxins’ and channel’s amino acid residues responsible for the selectivity and binding modes. Docking positions support earlier supposition that the higher neuronal activity observed in electrophysiology should be attributed to hampering the fast inactivation gate by interactions of an anemone toxin with the voltage driven S4 helix from domain IV of cockroach Nav channel (NavPaS). Our modelling provides new data linking activity of toxins with their mode of binding in site 3 of NavPaS channel.     

Structure and function of Buthus eupeus scorpion neurotoxins

At present the investigation of the complex mechanism of the signal conductance between nerve cells and their target organ is closely connected with application of neurotoxins as a valuable tool for study of important sites involved in nerve transmission. Particular interest is attributed to neurotoxins from scorpion venoms which slow down the inactivation rate of fast sodium channels of the excitable membrane. Fifteen different toxins have been isolated from Buthus eupeus scorpion venom by means of the combination of gel and ion exchange chromatography. The homologous insectotoxins I₁, I₃, I₄, and I₅ belong to the new structural type of scorpion toxins. The essential features of these toxins are low molecular weight (ca. 4000) and the presence of two or three methionine residues in their amino acid compositions. The complete amino acid sequences of two insectotoxins and one mammalian toxin have been estimated. All mammalian neurotoxins in concentration from 10^?9M to 10^?7M slow down the inactivation rate of fast sodium channels. The direct covalent binding of photosensitive radioactive neurotoxin derivatives presents one of the most effective approaches to the localization of the functionally important components of the fast sodium channels. Thus far 2,4-dinitro-5-fluorophenylazide has been used for chemical modification of the mammalian neurotoxin M₁₀. The prepared monosubstituted photosensitive neurotoxin derivative possessed a high biological activity according to the voltage clamp data. The neurotoxin derivative has been iodinated by ¹²⁵I, the resulted product being both radioactively and photoaffinity labeled. Both native neurotoxin M₁₀ and sea anemone toxin competitively inhibit the reception of the scorpion toxin derivative by rat brain synaptosomes. It was shown that covalent toxin-receptor complexes are composed of two protein components with molecular weights 76,000 and 51,000, being near equal for synaptosomes, neuroblastoma cells, and crab nerve plasma membranes.     

Synthesis and calcium channel antagonist activities of new derivatives of dialkyl 1,4-dihydro-2,6-dimethyl-4-(5-phenylisoxazol-3-yl)pyridine-3,5-dicarboxylates

The new analogues of nifedipine, in which 2-nitrophenyl group at position 4 is replaced by phenylisoxazolyl substituent, were synthesized. The symmetrical dialkyl 1,4-dihydro-2,6-dimethyl-4-(5-phenylisoxazol-3-yl)pyridine-3,5-dicarboxylates were prepared by classical Hantzsch condensation, and the asymmetrical analogues were synthesized using a procedure reported by Dagnino that involved the condensation of alkyl acetoacetate with alkyl 3-aminocrotonate and 5-phenylisoxazole-3-carboxaldehyde. The structure of all compounds was confirmed by IR, ¹H NMR and Mass spectra. In vitro calcium channel antagonist activities were evaluated as calcium channel antagonists using the high K⁺ concentration of guinea-pig ileum longitudinal smooth muscle (GPILSM) assay. These compounds exhibited moderate calcium antagonist activity (IC₅₀ = 10^?7 to 10^{? 5} M range) relative to the reference drug nifedipine (IC₅₀ = 1.10 ± 0.40 × 10^?8 M).     

Tx1, from Phoneutria nigriventer spidervenom, interacts with dihydropyridine sensitive-calcium channels in GH3 cells

Summary The aim of this work was to use the binding assay of tritiated-dihydropyridine and radioiodinated Tx1, isolated from the Phoneutria nigriventer venom, in order to show the presence of Ca_v1 calcium channels on pituitary tumour cell (GH3). We showed that GH3 cells have specific sites for ¹²⁵I-Tx1, which are sensitive to nifedipine (~20%). Reverse competition assay with ³H-PN200-110 (40% inhibition) and electrophysiological data (50% inhibition) suggest that Ca_v1 calcium channels are target sites for this toxin. To summarize, Tx1 binds to specific sites on GH3 cells and this interaction results in Ca_v1 calcium channel blockade. ³H-PN200-110 and ¹²⁵I-Tx1 binding assays proved to be useful tools to show the presence of calcium channels on GH3 cells.     

Feasibility of gamma irradiation as a stabilisation technique in the preparation of tissue reference materials for a range of shellfish toxins

The effect of γ-irradiation on concentrations of hydrophilic and lipophilic phycotoxins has been investigated by use of HPLC–UV and LC–MS. Pure toxins in organic solvents and toxins in mussel (Mytilus edulis) tissues were irradiated at three different doses. In solution all toxin concentrations were reduced to some extent. Most severe decreases were observed for domoic acid and yessotoxin, for which the smallest dose of irradiation led to almost complete destruction. For pectenotoxin-2 the decrease in concentration was less severe but still continuous with increasing dose. Azaspiracid-1 and okadaic acid were the least affected in solution. In shellfish tissue the decrease in toxin concentrations was much reduced compared with the effect in solution. After irradiation at the highest dose reductions in concentrations were between ca. 5 and 20% for the lipophilic toxins and there was no statistical difference between control and irradiated samples for azaspiracids in tissue. Irradiation of shellfish tissues contaminated with domoic acid led to a more continuous decrease in the amount of the toxin with increasing dose. The effect of irradiation on the viability of microbial activity in shellfish tissues was assessed by using total viable counting techniques. Microbial activity depended on the type of shellfish and on the pretreatment of the shellfish tissues (with or without heat treatment). As far as we are aware this is the first investigation of the effectiveness of irradiation as a technique for stabilising tissue reference materials for determination of phycotoxins. Our results suggest that this technique is not effective for materials containing domoic acid. It does, however, merit further investigation as a stabilisation procedure for preparation of shellfish tissue materials for some lipophilic toxins, in particular azaspiracids. Chemical structures of the toxins investigated in the study     

Feasibility study for the rapid screening of target molecules using translational diffusion coefficients: diffusion-ordered NMR spectroscopy of biological toxins

A panel of 15 biological toxins ranging between ~60–28,000 g/mol was used to evaluate the feasibility of screening aqueous samples for toxin analytes based on their translational diffusion coefficients, D _t. Toxin D _t values were measured by pulsed-field gradient ¹H NMR spectroscopy using a bipolar pulse pair, longitudinal eddy current delay pulse sequence incorporating water suppression to achieve the maximum dynamic range for toxin signals. To collect data for an effective screening protocol, reference D _t values were determined from five independent measurements at both 25 and 37 °C for all toxins in the panel. In the protocol, D _t values are measured at both temperatures for a suspected toxin target in a sample, and for assignment as a potential toxin analyte, the measurements are required to fall within ±0.25 × 10⁻⁶ cm²/s of both reference D _t values for at least one toxin in the panel. Only solution viscosity was found to influence sample D _t measurements appreciably; however, the measurements are easily corrected for viscosity effects by calculating the D _t value of the suspected toxin at infinite dilution. In conclusion, the protocol provides a rapid and effective means for screening aqueous samples for all toxins in the panel, narrowing toxin identification to ≤2 possibilities in virtually all cases.     

Multi-detection method for five common microalgal toxins based on the use of microspheres coupled to a flow-cytometry system

Freshwater and brackish microalgal toxins, such as microcystins, cylindrospermopsins, paralytic toxins, anatoxins or other neurotoxins are produced during the overgrowth of certain phytoplankton and benthic cyanobacteria, which includes either prokaryotic or eukaryotic microalgae. Although, further studies are necessary to define the biological role of these toxins, at least some of them are known to be poisonous to humans and wildlife due to their occurrence in these aquatic systems. The World Health Organization (WHO) has established as provisional recommended limit 1 μg of microcystin-LR per liter of drinking water. In this work we present a microsphere-based multi-detection method for five classes of freshwater and brackish toxins: microcystin-LR (MC-LR), cylindrospermopsin (CYN), anatoxin-a (ANA-a), saxitoxin (STX) and domoic acid (DA). Five inhibition assays were developed using different binding proteins and microsphere classes coupled to a flow-cytometry Luminex system. Then, assays were combined in one method for the simultaneous detection of the toxins. The IC₅₀'s using this method were 1.9 ± 0.1 μg L⁻¹ MC-LR, 1.3 ± 0.1 μg L⁻¹ CYN, 61 ± 4 μg L⁻¹ ANA-a, 5.4 ± 0.4 μg L⁻¹ STX and 4.9 ± 0.9 μg L⁻¹ DA. Lyophilized cyanobacterial culture samples were extracted using a simple procedure and analyzed by the Luminex method and by UPLC–IT-TOF-MS. Similar quantification was obtained by both methods for all toxins except for ANA-a, whereby the estimated content was lower when using UPLC–IT-TOF-MS. Therefore, this newly developed multiplexed detection method provides a rapid, simple, semi-quantitative screening tool for the simultaneous detection of five environmentally important freshwater and brackish toxins, in buffer and cyanobacterial extracts.     

Characterisation of botulinum toxins type A and B,by matrix-assisted laser desorption ionisation and electrospray mass spectrometry

A method earlier developed for the mass spectrometric (MS) identification of tetanus toxin (TTx) was applied to botulinum toxins type A and B (BTxA and BTxB). Botulinum toxins are extremely neurotoxic bacterial toxins, likely to be used as biological warfare agent. Biologically active BTxA and BTxB are comprised of a protein complex of the respective neurotoxins with specific haemagglutinins (HAs) and non-toxic non-haemagglutinins (NTNHs). These protein complexes are also observed in mass spectrometric identification. The particular BTxA complex, from Clostridium botulinum strain 62A, almost completely matched database data derived from genetic sequences known for this strain. Although no such database information was available for BTxB, from C. botulinum strain okra, all protein sequences from the complex except that of HA-70 were found to match proteins known from other type B strains. It was found that matrix-assisted laser desorption ionisation MS provides provisional identification from trypsin digest peptide maps and that liquid chromatography electrospray (tandem) mass spectrometry affords unequivocal identification from amino acid sequence information of digest peptides obtained in trypsin or pepsin digestion.     

Charge States of y Ions in the Collision-Induced Dissociation of Doubly Charged Tryptic Peptide Ions

Bonds that break in collision-induced dissociation (CID) are often weakened by a nearby proton, which can, in principle, be carried away by either of the product fragments. Since peptide backbone dissociation is commonly charge-directed, relative intensities of charge states of product y- and b-ions depend on the final location of that proton. This study examines y-ion charge distributions for dissociation of doubly charged peptide ions, using a large reference library of peptide ion fragmentation generated from ion-trap CID of peptide ions from tryptic digests. Trends in relative intensities of y²⁺ and y¹⁺ ions are examined as a function of bond cleavage position, peptide length (n), residues on either side of the bond and effects of residues remote from the bond. It is found that y_n-2/b₂ dissociation is the most sensitive to adjacent amino acids, that y²⁺/y¹⁺ steadily increase with increasing peptide length, that the N-terminal amino acid can have a major influence in all dissociations, and in some cases other residues remote from the bond cleavage exert significant effects. Good correlation is found between the values of y²⁺/y¹⁺ for the peptide and the proton affinities of the amino acids present at the dissociating peptide bond. A few deviations from this correlation are rationalized by specific effects of the amino acid residues. These correlations can be used to estimate trends in y²⁺/y¹⁺ ratios for peptide ions from amino acid proton affinities.     

Kinetics and mechanism of Os(VIII)-catalyzed hexacyanoferrate(III) oxidation of α amino acids in alkaline medium

The kinetics of the Os(VIII)-catalyzed oxidation of glycine, alanine, valine, phenylalanine, isoleucine, lycine, and glutamic acid by alkaline hexacyanoferrate(III) reveal that these reactions are zero order in hexacyanoferrate(III) and first order in Os(VIII). The order in amino acid as well as in alkali is 1 at [amino acid] ?2.5 × 10^?2M and [OH^?] ?1.3 × 10^?M, but less than unity at higher concentrations of amino acids or alkali. The active oxidizing species under the experimental conditions is OsO₄(H₂O) (OH)^?. The ferricyanide is merely used up to regenerate the Os(VIII) species from Os(VI) formed during the reaction. The structural influence of amino acids on the reactivity has been discussed. The amino acids during oxidation are shown to be degraded through intermediate keto acids. The kinetic data are accommodated by considering the interaction between the conjugate base of the amino acids and the active oxidizing species of Os(VIII) to form a transient complex in the primary rate-determining step. The catalytic effect of hexacyanoferrate(II) has been rationalized.     

Computer modeling of binding of diverse weak toxins to nicotinic acetylcholine receptors

Weak toxins are the "three-fingered" snake venoms toxins grouped together by having an additional disulfide in the N-terminal loop I. In general, weak toxins have low toxicity, and biological targets have been identified for some of them only, recently by detecting the effects on the nicotinic acetylcholine receptors (nAChR). Here the methods of docking and molecular dynamics simulations are used for comparative modeling of the complexes between four weak toxins of known spatial structure (WTX, candoxin, bucandin, gamma-bungarotoxin) and nAChRs. WTX and candoxin are those toxins whose blocking of the neuronal alpha7- and muscle-type nAChR has been earlier shown in binding assays and electrophysiological experiments, while for the other two toxins no such activity has been reported. Only candoxin and WTX are found here to give stable solutions for the toxin-nAChR complexes. These toxins appear to approach the binding site similarly to short alpha-neurotoxins, but their final position resembles that of alpha-cobratoxin, a long alpha-neurotoxin, in the complex with the acetylcholine-binding protein. The final spatial structures of candoxin and WTX complexes with the alpha7 neuronal or muscle-type nAChR are very similar and do not provide immediate answer why candoxin has a much higher affinity than WTX, but both of them share a virtually irreversible mode of binding to one or both these nAChR subtypes. Possible explanation comes from docking and MD simulations which predict fast kinetics of candoxin association with nAChR, no gross changes in the toxin conformation (with smaller toxin flexibility on alpha7 nAChR), while slow WTX binding to nAChR is associated with slow irreversible rearrangement both of the tip of the toxin loop II and of the binding pocket residues locking finally the toxin molecule. Computer modeling showed that the additional disulfide in the loop I is not directly involved in receptor binding of WTX and candoxin, but it stabilizes the structure of loop I which plays an important role in toxin delivery to the binding site. In summary, computer modeling visualized possible modes of binding for those weak toxins which interact with the nAChR, provided no solutions for those weak toxins whose targets are not the nAChRs, and demonstrated that the additional disulfide in loop I cannot be a sound criteria for joining all weak toxins into one group; the conclusion about the diversity of weak toxins made from computer modeling is in accord with the earlier phylogenetic analysis.     

Molecular recognition properties of acyclic cucurbiturils toward amino acids,peptides, and a protein

The binding of 1 and 2 toward 19 amino acid amides by ¹H NMR and ITC is reported. Hosts 1 and 2 bind to aromatic or hydrophobic residues by cavity inclusion leaving the cationic residues at the C=O portals. K_a values range from 10² to >10⁶ M^?1 with H-Phe-NH₂, H-Trp-NH₂, and H-Tyr-NH₂ displaying sub-micromolar K_d values. Hosts 1 and 2 bind tightly to dicationic H-Lys-NH₂ and H-Arg-NH₂ which are poor guests for CB[7]. Comparison of the affinity of 1 and 2 toward the amino acid amide, N-acetyl-amino-acid amide, and amino acid forms of Phe revealed that the removal of the NH₃⁺ to O=C and SO₃^? electrostatic interactions costs 3.8 kcal/mol whereas the introduction of an unfavourable CO₂^? to O=C and SO₃^? electrostatic interactions costs 2.1 kcal/mol. Hosts 1 and 2 bind to insulin with low micromolar affinity. Acyclic CB[n] display high affinity toward a wider range of N-terminal amino acids residues than CB[n] which suggests a broad range of applications.     

The effect of amino acid backbone length on molecular packing: crystalline tartrates of glycine, β‐alanine, γ‐aminobutyric acid (GABA) and dl‐α‐aminobutyric acid (AABA)

We report a novel 1:1 cocrystal of β‐alanine with dl ‐tartaric acid, C₃H₇NO₂·C₄H₆O₆, (II), and three new molecular salts of dl ‐tartaric acid with β‐alanine {3‐azaniumylpropanoic acid–3‐azaniumylpropanoate dl ‐tartaric acid–dl ‐tartrate, [H(C₃H₇NO₂)₂]⁺·[H(C₄H₅O₆)₂]⁻, (III)}, γ‐aminobutyric acid [3‐carboxypropanaminium dl ‐tartrate, C₄H₁₀NO₂⁺·C₄H₅O₆⁻, (IV)] and dl ‐α‐aminobutyric acid {dl ‐2‐azaniumylbutanoic acid–dl ‐2‐azaniumylbutanoate dl ‐tartaric acid–dl ‐tartrate, [H(C₄H₉NO₂)₂]⁺·[H(C₄H₅O₆)₂]⁻, (V)}. The crystal structures of binary crystals of dl ‐tartaric acid with glycine, (I), β‐alanine, (II) and (III), GABA, (IV), and dl ‐AABA, (V), have similar molecular packing and crystallographic motifs. The shortest amino acid (i.e. glycine) forms a cocrystal, (I), with dl ‐tartaric acid, whereas the larger amino acids form molecular salts, viz. (IV) and (V). β‐Alanine is the only amino acid capable of forming both a cocrystal [i.e. (II)] and a molecular salt [i.e. (III)] with dl ‐tartaric acid. The cocrystals of glycine and β‐alanine with dl ‐tartaric acid, i.e. (I) and (II), respectively, contain chains of amino acid zwitterions, similar to the structure of pure glycine. In the structures of the molecular salts of amino acids, the amino acid cations form isolated dimers [of β‐alanine in (III), GABA in (IV) and dl ‐AABA in (V)], which are linked by strong O—H…O hydrogen bonds. Moreover, the three crystal structures comprise different types of dimeric cations, i.e. (A…A)⁺ in (III) and (V), and A⁺…A⁺ in (IV). Molecular salts (IV) and (V) are the first examples of molecular salts of GABA and dl ‐AABA that contain dimers of amino acid cations. The geometry of each investigated amino acid (except dl ‐AABA) correlates with the melting point of its mixed crystal.     

Molecular dynamics simulation exploration of cooperative migration mechanism of calcium ions in sarcoplasmic reticulum Ca2+‐ATPase

Calcium ATPase is a member of the P‐type ATPase, and it pumps calcium ions from the cytoplasm into the reticulum against a concentration gradient. Several X‐ray structures of different conformations have been solved in recent years, providing basis for elucidating the active transport mechanism of Ca²⁺ ions. In this work, molecular dynamics (MD) simulations were performed at atomic level to investigate the dynamical process of calcium ions moving from the outer mouth of the protein to their binding sites. Five initial locations of Ca²⁺ ions were considered, and the simulations lasted for 2 or 6 ns, respectively. Specific pathways leading to the binding sites and large structural rearrangements around binding sites caused by uptake of calcium ions were identified. A cooperative binding mechanism was observed from our simulation. Firstly, the first Ca²⁺ ion binds to site I , and then, the second Ca²⁺ ion approaches. The interactions between the second Ca²⁺ and the residues around site I disturb the binding state of site I and weaken its binding ability for the first bound Ca²⁺. Because of the electrostatic repulsion of the second Ca²⁺ and the electrostatic attraction of site II , the first bound Ca²⁺ shifts from site I to site II . Concertedly, the second Ca²⁺ binds to site I , forming a binding state with two Ca²⁺ ions, one at site I and the other at site II . Both of Glu908 and Asp800 coordinate with the two Ca²⁺ ions simultaneously during the concerted binding process, which is believed to be the hinge to achieve the concerted binding. In our simulations, four amino acid residues that serve as the channel to link the outer mouth and the binding sites during the binding process were recognized, namely Tyr837, Tyr763, Asn911, and Ser767. The analyses regarding the activity of the proteins via mutations of some key residues also supported our cooperative mechanism. © 2009 Wiley Periodicals, Inc. J Comput Chem 2009     

Synthesis and Regioselective Hydrolysis of Novel Dialkyl 4-Imidazolyl-1,4-Dihydropyridine-3,5-dicaroxlates as Potential Dual Acting Angiotensin II Inhibitors and Calcium Channel Blockers

Most of the known effects of angiotensin II are mediated via AT₁ receptor by increasing intracellular Ca²⁺ by influx of extracellular Ca²⁺. Combination therapies of angiotensin receptor blocker (ARB) with calcium channel blocker (CCB) which act through L-type calcium channel have beneficial therapeutic and protective effects on cardiovascular system. Thus, it was hypothesized that merging the key structural elements present in an AT₁ receptor antagonist (telmisartan) with key structural elements in 1,4-dihydropyridine calcium channel blockers (nifedipine) would yield a compound with dual activity for both receptors. This strategy led to the design and synthesis of dialkyl 1,4-dihydro-2,6-dimethyl-4-[2-n-alkyl-1-[(2′-carboxybiphenyl- 4-yl)methyl]imidazole-4(or 5)-yl]-3,5-pyridinedicarboxylates (4 and 6). The synthesis of compounds 4 and 6 was accomplished through the reaction of 2-n-alkyl-1-[(2′-carbomethoxybiphenyl-4-yl)methyl]imidazole-4(or 5)-carboxaldehydes with alkyl acetoacetate followed by regioselctive hydrolysis of carboethoxybiopheny to carboxybiphenyl that are essential for ARB activity. It is suggested that existence of hindrance by substituted groups prevent hydrolysis of esteric groups on dihydropyridine ring. The structures of the compounds were characterized by ¹H-nuclear magnetic resonance, infrared and mass spectroscopy.     

Adamantylcalixarenes with CMPO groups at the wide rim: synthesis and extraction of lanthanides and actinides

Starting from p-adamantylcalix[4]- and [6]arenes functionalized with carboxylic acid or ester groups at the adamantane nuclei, carbamoylmethylphosphine oxide (CMPO)-containing ligands of a novel type were synthesized. They were studied as extractants for a series of f-block elements including radioactive ¹⁵²Eu(III), ²⁴¹Am(III), ²³³U(VI), and ²³⁹Pu(IV). Tetrameric ligand 4b in which CMPO residues are connected to adamantane nuclei through methylene groups gave the best extraction results for lanthanides and actinides. For all the ligands the extraction efficiency does not decrease at higher nitric acid concentration. Although the discrimination between trivalent actinides and lanthanides is not good, all ligands are highly selective for thorium(IV) with the best separation factor achieved in the case of hexameric ligand 5 (D_Th/D_Ln>24).     

Synthesis of alkyl 6‐methyl‐4‐(2‐pyridyl)‐1,2,3,4‐tetrahydro‐2H‐pyrimidine‐2‐one‐5‐carboxylates for evaluation as calcium channel antagonists

The Bigenelli acid catalyzed condensation of 2‐pyridylcarboxaldehyde ( 1 ), urea ( 2 ) and an alkyl acetoacetate ( 3 ) afforded the respective alkyl (Me, Et, i‐Pr, i‐Bu, t‐Bu) 6‐methyl‐4‐(2‐pyridyl)‐1,2,3,4‐tetrahydro‐2H‐pyrimidine‐2‐one‐5‐carboxylates ( 4a‐e ). The most potent calcium channel antagonist ethyl 6‐methyl‐4‐(2‐pyridyl)‐1,2,3,4‐tetrahydro‐2H‐pyrimidine‐2‐one‐5‐carboxylate ( 4b , IC₅₀ = 1.67 × 10^?5 M) wasa much weaker calcium channel antagonist than the reference drug nifedipine (Adalat®, IC₅₀ = 1.40 × 10^?8 M) on guinea pig ileal longitudinal smooth muscle (GPILSM). The alkyl 6‐methyl‐4‐(2‐pyridyl)‐1,2,3,4‐tetrahydro‐2H‐pyrimidine‐2‐one‐5‐carboxylates did not show any inotropic effect on heart since no increase, or decrease, in the contractile force of guinea pig left atrium was observed. These structure activity studies show that the alkyl 6‐methyl‐4‐(2‐pyridyl)‐1,2,3,4‐tetrahydro‐2H‐pyrimidine‐2‐one‐5‐carboxylates ( 4a‐e ) are partial bioisosteres of nifedipine with respect to calcium channel antagonist activity on guinea pig ileal longitudinal smooth muscle (GPILSM).     

Mixed-Ligand Complex Formation Equilibria of Vanadium(III) with 2,2′-Bipyridine and the Amino Acids Glycine, Proline, α-Alanine and β-Alanine Studied in 3.0 mol?dm?3 KCl at 25 °C

We studied speciation of the mixed-ligand complex formation equilibria of vanadium(III) with both 2,2??-bipyridine (Bipy) and the amino acids glycine (HGly), proline (HPro), ??-alanine (H??Ala), and ??-alanine (H??Ala) by means of electromotive forces measurements emf(H) using 3.0?mol?dm^?3 KCl as the ionic medium at 25 °C. The experimental data were analyzed by means of the computational least-squares program LETAGROP, taking into account the hydrolysis of the vanadium(III) cation, the respective stability constants of the binary complexes, and the acid/base reactions of the ligands which were kept fixed during the analysis. In all four amino acid systems studied we observed the complexes [V₂O(Bipy)(B)]³⁺, [V₂O(Bipy)₂(B)₂]²⁺, [V(OH)(Bipy)(B)₂] and [V(OH)₂(Bipy)(B)], where B represents the deprotonated form of the amino acids studied in this work. The respective stability constants were determined and the species distribution diagrams as a function of pH are briefly discussed.     

Formation Constants for the Ternary Complexes of Vanadium(III), 2,2′-Bipyridine, and the Amino Acids Histidine, Cysteine, Aspartic and Glutamic Acids

In this study we report the stability constants and the speciation of the ternary vanadium(III) complexes with 2,2??-bipyridine (Bipy) and the amino acids histidine (HHis), cysteine (H₂Cys), aspartic acid (H₂Asp) and glutamic acid (H₂Glu) by means of potentiometric titrations employing 3.0 mol?dm^?3 KCl as the ionic medium at 25?°C. The potentiometric data were analyzed taking into account the hydrolysis of the vanadium(III) cation and the respective stability constants of the binary complexes and the acid?Cbase reactions of the ligands, which were kept fixed during the analysis. The complexes detected in the different systems are: in the vanadium(III)?CBipy?CHHis system, [V(HBipy)(HHis)]⁴⁺ and [V(HBipy)(H₂His)]⁵⁺; in the vanadium(III)?CBipy?CH₂Cys system, [V₂O(Bipy)(Cys)]²⁺; in the vanadium(III)?CBipy?CH₂Asp system, [V(Bipy) (Asp)]⁺, [V₂O(Bipy)(Asp)]²⁺, and V₂O(Bipy)₂(Asp)₂; and finally in the vanadium(III)?CBipy?CH₂Glu system, [V(Bipy)(H₂Glu)]³⁺ and [V(Bipy)(Glu)]⁺. The respective stability constants were determined and the specie distribution diagrams as a function of pH are briefly discussed.