Water‐soluble calix[4]resorcinarenes as chiral NMR solvating agents for bicyclic aromatic compounds

Water‐soluble calix[4]resorcinarenes with proline, 3‐hydroxyproline, and 4‐hydroxyproline substituent groups are evaluated as chiral NMR solvating agents on a series of bicyclic aromatic compounds with naphthyl, indole, dihydroindole, and indane rings. The substrates interact with the calixresorcinarene through insertion of the aromatic ring into the cavity. Most of the substrates are analyzed as cationic species, although one anionic species is analyzed. All of the substrates exhibit enantiomeric discrimination in the ¹H‐NMR spectrum with one or more of the calixresorcinarenes. In most cases, the hydroxyproline derivatives are more effective at causing enantiodifferentiation than the corresponding proline derivative. Presumably, the hydroxyl group on the proline moieties is involved in interactions with the substituent groups of the substrate that are important in creating chiral recognition. The enantiomeric discrimination in the ¹H‐NMR spectrum is large enough for many resonances to permit the analysis of enantiomeric purity. Chirality 2009. © 2009 Wiley‐Liss, Inc.     

Study of calix[4]resorcinarene-dopamine complexation in mixed phospholipid monolayers formed at the air-water interface

We have studied the physical properties of monolayers formed by calix[4]resorcinarene and in mixtures with dipalmitoyl phosphatidylcholine (DPPC) in various molar ratios formed at the air-water interface and at presence of dopamine in water subphase by means of measurements of surface pressure and dipole potential. We showed that both calix[4]resorcinarene as well as its mixture with DPPC form stable monolayers at the water subphase. The presence of dopamine resulted in an increase of the mean molecular area and in a decrease of the compressibility modulus of the monolayers. For mixed monolayers at higher content of calix[4]resorcinarene (> 0.2 molar fraction) a deviation from ideal miscibility took place especially for monolayers in a solid state. This can be connected with formation of aggregates of calix[4] resorcinarene. Lowest miscibility and weakest interaction of dopamine with a monolayer was observed for calix[4]resorcinarene molar fraction of 0.33 in the monolayer.     

Study of calix[4]resorcinarene-dopamine complexation in mixed phospholipid monolayers formed at the air-water interface

We have studied the physical properties of monolayers formed by calix[4]resorcinarene and in mixtures with dipalmitoyl phosphatidylcholine (DPPC) in various molar ratios formed at the air-water interface and at presence of dopamine in water subphase by means of measurements of surface pressure and dipole potential. We showed that both calix[4]resorcinarene as well as its mixture with DPPC form stable monolayers at the water subphase. The presence of dopamine resulted in an increase of the mean molecular area and in a decrease of the compressibility modulus of the monolayers. For mixed monolayers at higher content of calix[4]resorcinarene (> 0.2 molar fraction) a deviation from ideal miscibility took place especially for monolayers in a solid state. This can be connected with formation of aggregates of calix[4] resorcinarene. Lowest miscibility and weakest interaction of dopamine with a monolayer was observed for calix[4]resorcinarene molar fraction of 0.33 in the monolayer.     

Embedding calix[4]resorcinarenes in liposomes: Experimental and computational investigation of the effect of resorcinarene inclusion on liposome properties and stability

Two calix[4]resorcinarenes, which differ in the length of the alkyl chain on the methylene bridge between the aromatic rings, have been embedded in unilamellar liposomes prepared from 1-palmitoyl-2-oleoyl-sn-glycerol-3-phosphocholine in three host/guest ratios, following two different procedures. The effect of the insertion of the guests has been evaluated through the measurements of the viscosity and the kinetic stability of the liposomal systems by means of the fluorescent probes pyrene and 5(6)-carboxyfluorescein. The presence of the guests reduces the viscosity of the liposomes, suggesting a modification of the bilayer structure. However, this does not affect liposome stability. A calix[4]resorcinarene cavitand with a more rigid conformation compared to the parent resorcinarene, has been also synthetized and embedded in liposomes. The free energy of the insertion of the substrates in the lipid bilayer has been evaluated through Molecular Dynamics simulations.     

Complex formation of cytochrome C with a calixarene carboxylic acid derivative: a novel solubilization method for biomolecules in organic media

A calixarene carboxylic acid derivative has been found to form a complex with the cationic protein cytochrome c. The solubilized cytochrome c was stable and showed peroxidase activity in chloroform. The calix[6]arene and the calix[8]arene achieved quantitative extraction of the protein. The calix[6]arene, whose cavity is well-fitted to a protonated amino group, exhibited a selectivity to lysine-rich proteins due to the recognition of the epsilon-amino groups in lysine residues on the surface of the protein. This is the first report showing protein extraction by calixarenes. The solubilized cytochrome c could catalyze an oxidative reaction in organic solvents. This host compound functions as a novel solubilization tool for biomolecules and a separation tool for lysine-rich proteins.     

Extraction selectivities of lower rim substituted calix[4]arene hosts induced by variations in the upper rim substituents

The compound 25,26,27,28-tetra-(2-dimethyldithiocarbamoylethoxy)calix[4]arene has been prepared from 25,26,27,28-tetra-(2-bromoethoxy)calix[4]arene by reaction with sodium dimethyldithiocarbamate. As an extractant for heavy metal ions 25,26,27,28-tetra-(2-dimethyldithiocarbamoylethoxy)calix[4]arene is effective for Hg²⁺, Ag⁺, Pd²⁺ and Au³⁺, but much less effective than 5,11,17,23-tetra-tert-butyl-25,26,27,28-tetra-(2-N,N-dimethyldithiocarbamoylethoxy)calix[4]arene for both Hg₂²⁺ and MeHg⁺. Calixarene alcohols also show selectivity as hosts. The alcohol derivative 25,26,27,28-tetra-(2-hydroxyethoxy)calix[4]arene undergoes slow occlusion of iodine into the lower rim, whereas with the alcohol 5,11,17,23-tetra-tert-butyl-25,26,27,28-tetra-(2-hydroxyethoxy)calix[4]arene no interaction is observed.     

Conjugation, immunoreactivity, and immunogenicity of calix[4]arenes; model study to potential calix[4]arene-based Ac3+ chelators.

For the development of calix[4]arene-based radiotherapeutic agents, the conjugation to biomolecules and immunogenicity in mice of potential 225Ac3+-chelating calix[4]arenes were studied. A calix[4]arene triethyl ester isothiocyanate and a bis(calix[4]arene) hexacarboxylic acid, containing a masked thiol functionality, were used in conjugation experiments to a mouse monoclonal antibody and serum albumins. All characterization techniques indicate that only the calix[4]arene carboxylic acid is successfully conjugated to the biomolecules. The immunoreactivity of the conjugates is not impaired when up to 6 equiv of calixarene are bound to the monoclonal antibody. Animal tests indicated that the immunogenicity toward the calix[4]arene is strongly influenced by the nature of the carrier, the dosage, and the injection method. No immune response occurred when a homologous carrier was used or when a heterologous carrier was applied at a dosage of 10 microg per immunization via intravenous injection. Under all other conditions, the presence of antibodies directed against the calix[4]arene was demonstrated. Thus, for the application in radioimmunotherapy, the conjugation of a calix[4]arene to a humanized antibody will probably not lead to an immune response, and the immunoreactivity will not be disturbed.     

Calix[4]arene methylenebisphosphonic acids: the features of alkaline phosphatases inhibition

The inhibition of alkaline phosphatases by calix[4]arenes functionalysed at the macrocyclic upper rim by one or two methylenebisphosphonic acid fragments has been investigated. It is established, that calix[4]arene bismethylenebisphosphonic acid displayed stronger inhibition of alkaline phosphatase from bovine intestine mucosa than calix[4]arene methylenebisphosphonic acid. At the same time, the both inhibitors showed almost similar levels of inhibitory activities in respect of bovine kidney alkaline phosphatase or E. coli alkaline phosphatase. The tested compounds were docked computationally to the active site of the E. coli alkaline phosphatase. On the basis of results obtained the possible binding modes of inhibitors were analysed.     

Modulation of cell proliferation in rat liver cell cultures by new calix[4]arenes

Cell cycle progression is dependent on intracellular iron level and chelators lead to iron depletion and decrease cell proliferation. This antiproliferative effect can be inhibited by exogenous iron. In this work, we present the synthesis of new synthetic calix[4]arene podands bearing two aspartic/glutamic acid, ornithine groups or hydrazide function at the lower rim, designed as potential iron chelators. The synthesis only afforded calix[4]arenes in the cone conformation. We report their effect on cell proliferation, in comparison with the new oral chelator ICL670A (4-[3,5-bis-(2-hydroxyphenyl)-1,2,4-triazol-1-yl]-benzoic acid). The antiproliferative effect of these new compounds was studied in the rat hepatoma cell line Fao by measuring mitochondrial succinate dehydrogenase activity. Their cytotoxicity was evaluated by extracellular LDH activity. Preliminary results indicated that among all tested compounds, monohydrazidocalix[4]arene 2 which is not cytotoxic in Fao cells exhibits interesting antiproliferative activity. This effect, independent on iron depletion, remains to be further explored. Moreover, it also shows that new substituted calix[4]arenes could open the way to new valuable medicinal chemistry scaffolding.     

Calix[4]arenes C-136 and C-137 hyperpolarize myometrium mitochondria membranes

Calixarenes are supramolecular compounds interacting with bioactive molecules and ions, causing changes in biochemical and biophysical processes. The aim of this work was to study the effects of calix[4]arenes C-136, C-137, and C-138 at the level of polarization of the rat myometrium mitochondria membrane. The structure of synthesized calix[4]arene molecules was confirmed by the methods of ¹H NMR and infrared spectroscopy. Calix[4]arenes C-136 and C-137 each possess two chalcone amide moieties at the lower rim, while calix[4]arene C-138, only one. Calix[4]arenes C-136 and C-137 differ by the presence of ether or hydroxyl groups, respectively, at the lower rim of calix[4]arene skeleton, as well as the length of alkyl spacer between chalcone amide group and the macrocycle. It was shown that calix[4]arenes C-136, C-137, and C-138 form micelles in aqueous medium and in dimethylformamide (DMF). Irradiation of micelles with an argon laser on the flow cytometer results in the rise of autofluorescence. In an aqueous medium, calix[4]arene micelles interact with a positively charged voltage-sensitive fluorescent probe TMRM, which can testify to the presence of negative charge in these structures. However, calix[4]arene micelles do not interact with TMRM in DMF solution. The mitochondrial membrane potential was measured using fluorescent dyes MTG and TMRM with confocal microscopy and fluorescent dye TMRM with flow cytometry. Experiments were conducted on myometrium cells in culture and on suspension of digitonin-permeabilized uterus myocytes. It was shown that the fluorescent signal was stable during the time of experiment. Calix[4]arenes C-136 and C-137 (10 μM) hyperpolarize mitochondria membranes. At maximum, the effect was 173% relative to the control. At the same time, calix[4]arene C-138 did not influence the mitochondria membrane potential. The relationship between the structural organization of investigated calix[4]arene molecules and their effect on polarization of the mitochondria membrane is discussed.     

Improvement of catalytic activity of Candida rugosa lipase in the presence of calix[4]arene bearing iminodicarboxylic/phosphonic acid complexes modified iron oxide nanoparticles

In the present study, iron oxide magnetite nanoparticles, prepared through a co-precipitation method, were coated with phosphonic acid or iminodicarboxylic acid derivatives of calix[4]arene to modulate their surfaces with different acidic groups. Candida rugosa lipase was then directly immobilized onto the modified nanoparticles through sol–gel encapsulation. The catalytic activities and enantioselectivities of the two encapsulated lipases in the hydrolysis reaction of (R/S)-naproxen methyl ester and (R/S)-2-phenoxypropionic acid methyl ester were assessed. The results showed that the activity and enantioselectivity of the lipase were improved when the lipase was encapsulated in the presence of calixarene-based additives; the encapsulated lipase with the phosphonic acid derivative of calix[4]arene had an excellent rate of enantioselectivity against the (R/S)-naproxen methyl and (R/S)-2-phenoxypropionic acid methyl esters, with E = 350 and 246, respectively, compared to the free enzyme. The encapsulated lipases (Fe-Calix-N(COOH)) and (Fe-Calix–P) showed good loading ability and little loss of enzyme activity, and the stability of the catalyst was very good; they only lost 6–11% of the enzyme’s activity after five batches.     

Chiral calix[4]arenes bearing amino alcohol functionality as membrane carriers for transport of chiral amino acid methylesters and mandelic acid

Novel chiral calix[4]arene derivatives bearing amino alcohol moieties at the lower rim have been synthesized from the reaction of p-tert-butylcalix[4]arene diester with various amino alcohols. The transport of amino acid esters (phenylglycine, phenylalanine, and tryptophan methyl esters hydrochloride) and mandelic acid were studied through chloroform bulk liquid membrane system using chiral calix[4]arenes 15-20. All these receptors have been found to act as carriers for transport of aromatic amino acid methylesters and mandelic acid from the aqueous source phase to the aqueous receiving phase. The influence of calixarene and guest structures upon transport through liquid membrane is discussed.     

Calix[4]arene C-90 selectively inhibits Ca2+,Mg2+-ATPase of myometrium cell plasma membrane

The supramolecular compound calix[4]arene C-90 (5,11,17,23-tetra(trifluoro)methyl(phenylsulfonylimino)-methylamino-25,26,27,28-tetrapropoxycalix[4]arene) is shown to efficiently inhibit the ATP hydrolase activity of Ca²⁺,Mg²⁺-ATPase in the myometrium cell plasma membrane fraction and also in a preparation of the purified enzyme solubilized from this subcellular fraction. The inhibition coefficient I _0.5 values were 20.2 ± 0.5 and 58.5 ± 6.4 μM for the membrane fraction and the solubilized enzyme, respectively. The inhibitory effect of calix[4]arene C-90 was selective comparatively to other ATPases localized in the plasma membrane: calix[4]arene C-90 did not influence the activities of Na⁺,K⁺-ATPase and “basal” Mg²⁺-ATPase. The inhibitory effect of calix[4]arene C-90 on the Ca²⁺,Mg²⁺-ATPase activity was associated with the cooperative action of four trifluoromethylphenyl sulfonylimine (sulfonylamidine) groups oriented similarly on the upper rim of the calix[4]arene macrocycle (the calix[4]arene “bowl”). The experimental findings seem to be of importance for studies, using calix[4]arene C-90, of membrane mechanisms of regulation of calcium homeostasis in smooth muscle cells and also for investigation of the participation of the plasma membrane Ca²⁺-pump in control of electro- and pharmacomechanical coupling in myocytes.     

Pyrene-p-tert-butylcalixarenes inclusion complexes formation: a surface photochemistry study.

Diffuse reflectance and luminescence techniques were used to study the photophysics and photochemistry of pyrene within p-tert-butylcalix[n]arenes with n = 4, 6, and 8, and to study their ability to form inclusion complexes in heterogeneous media. Evidences for inclusion complex formation were found for the three hosts under study. Ground state diffuse reflectance results have shown the formation of ground state dimers of pyrene inside the cavity of calix[6]arene and calix[8]arene, with this feature much more evident for calix[6]arene. For calix[4]arene, only a monomer fits inside the cavity and the presence of pyrene microcrystals outside the cavity was detected. A luminescence lifetime distribution analysis was performed, revealing the presence of prompt emissions from the pyrene microcrystals outside the cavity in the case of calix[4]arene and from the constrained dimers inside the cavities of calix[6]arene and calix[8]arene. Transient absorption results have shown the presence of pyrene radical cation and also of trapped electrons for the three hosts under study. The formation of the phenoxyl radical of the calixarene following the laser pulsed excitation of pyrene at 355 nm is increased for calix[6]arene and calix[8]arene. This feature is particularly relevant for calix[6]arene, suggesting a very favourable situation for the hydrogen atom abstraction to occur. The analysis of the degradation products revealed the presence of hydroxypyrene as a major photodegradation product for the three hosts. Dihydro-hydroxypyrene was also formed in the case of calix[6]arene and calix[8]arene. The formation of the calixarene's phenoxyl radical and subsequent hydrogen abstraction is consistent with the formation of dihydro-dihydroxypyrene.     

Structural and functional bases of the intermolecular interaction of calix[4]arene C-97 with myosin subfragment-1 of myometrium

Calix[4]arene C-97 (code is shown) is the macrocyclic compound which has lipophilic intramolecular higly-structured cavity formed by four aromatic cycles, one of which on the upper rim is modified by methylene bisphosphonic group. It was shown that calix[4]arene C-97 (100 microM) efficiently inhibits ATPase activity of myosin subfragment-1 from pig myometrium, the inhibition coefficient I(0.5) being 83 +/- 7 microM. At the same time, this compound at 100 microM concentration significantly increases the effective hydrodynamic diameter of myosin subfragment-1, that may be indicative of intermolecular complexation between the calix[4]arene and myosin head. Computer simulation methods (docking, molecular dynamics, involving the Grid) have been used to clarify structural basis of the intermolecular interaction of calix[4]arene C-97 with myosin subfragment-1 of the myometrium; participation of hydrophobic, electrostatic and pi-pi (stacking) interactions between calix[4]arene C-97 and amino acid residues of myosin subfragment-1, some of them being located near the active site of the ATPase has been found out.     

New strapped calix[4]pyrrole based receptor for anions

Calix[4]pyrrole bearing catechol-derived diether strap linked via alkyl chains has been synthesized and characterized for the first time. The strap with 1,2-diether link is providing a relatively constrained geometry on its side of the calix[4]pyrrole moiety. As a result only one isomer (cis-type) of the receptor formed during reaction. The crystal structure reveals two molecules of methanol bound to the host. This calix[4]pyrrole also exhibits enhanced binding towards halide anions compared to simple calix[4]pyrrole apart from showing binding towards dihydrogenphosphate and acetate ions. The association constants are quite similar to that found for orcinol strapped calix[4]pyrrole towards halide anions in general, but having a higher preference for chloride than bromide ion in particular. Further it shows very strong preference towards fluoride ion.     

Carboxymethylated cyclodextrins and their complexes with Pr(III) and Yb(III) as water‐soluble chiral NMR solvating agents for cationic compounds

Cyclodextrins that are indiscriminately carboxymethylated at the 2‐, 3‐, and 6‐positions are used as chiral NMR solvating agents for cationic substrates with phenyl, naphthyl, pyridyl, indoline, and indole rings. Enantiodifferentiation with the α‐, β‐, and γ‐cyclodextrin derivatives is compared. The carboxymethylated derivatives are almost always more effective as chiral NMR solvating agents for cationic substrates than native cyclodextrins. The most effective carboxymethylated cyclodextrin varies for different substrates, and at times even different resonances of the substrate. Addition of paramagnetic praseodymium(III) or ytterbium(III) to mixtures of the carboxymethylated cyclodextrin and substrate often causes enhancements in enantiomeric discrimination and facilitates measurements of enantiomeric purity. The lanthanide ion bonds to the carboxymethyl groups and causes perturbations in the chemical shifts in the NMR spectra of substrate molecules in the cyclodextrin cavity. Chirality, 2010. © 2009 Wiley‐Liss, Inc.     

Calixarene-based phosphinic acids as inhibitors of protein tyrosine phosphatases

In the present work, the derivatives of calix[4]arene, thiacalix[4]arene, and sulfonylcalix[4]arene bearing four methylene(phenyl)phosphinic acid groups on the upper rim of the macrocycle were synthesized and studied as inhibitors of human protein tyrosine phosphatases. The inhibitory capacities of the three compounds towards PTP1B were higher than those for protein tyrosine phosphatases TC–PTP, MEG1, MEG2, and SHP2. The most potent sulfonylcalix[4]arene phosphinic acid displayed K_i value of 32?nM. The thiacalix[4]arene phosphinic acid was found to be a low micromolar inhibitor of PTP1B with selectivity over the other PTPs. The kinetic experiments showed that the inhibitors compete with the substrate for the active site of the enzyme. Molecular docking was performed to explain possible binding modes of the calixarene-based phosphinic inhibitors of PTP1B.     

Designing nanoporous crystalline materials by self-assembly: 2D hydrogen-bonded networks from upper rim calix[4]arene diamide derivatives

The self-assembly properties in the solid state of tectons derived from the reaction of calix[4]arene aminomethyl derivatives with a carboxylic acid or a protected amino acid are described, along with their conformational properties in solution. The X-ray crystal structures show that these compounds self-assemble in the lattice through a network of hydrogen-bonds between the amide groups, which overall results in infinite nanotubes formed by the calixarene aromatic cavities. Small changes in the upper rim substituents do not disturb the basic self-assembly motif which appears general and quite useful for controlling crystal growth by design and obtaining novel nanoporous crystalline materials.     

Inhibition of Yersinia protein tyrosine phosphatase by phosphonate derivatives of calixarenes

Inhibition of Yersinia protein tyrosine phosphatase by calix[4]arene mono-, bis-, and tetrakis(methylenebisphosphonic) acids as well as calix[4]arene and thiacalix[4]arene tetrakis(methylphosphonic) acids have been investigated. The kinetic studies revealed that some compounds in this class are potent competitive inhibitors of Yersinia PTP with inhibition constants in the low micromolar range. The binding modes of macrocyclic phosphonate derivatives in the enzyme active center have been explained using computational docking approach. The results obtained indicate that calix[4]arenes are promising scaffolds for the development of inhibitors of Yersinia PTP.