Enantiomeric Separation of New Chiral Azole Compounds.

Twelve new azole compounds were synthesized through an ene reaction involving methylidene heterocycles and phenylmaleimide, producing four oxazoles, five thiazoles, and one pyridine derivative, and ethyl glyoxal for an oxazole and a thiazole compound. The twelve azoles have a stereogenic center in their structure. Hence, a method to separate the enantiomeric pairs, must be provided if any further study of chemical and pharmacological importance of these compounds is to be accomplished. Six chiral stationary phases were assayed: four were based on macrocyclic glycopeptide selectors and two on linear carbohydrates, i.e., derivatized maltodextrin and amylose. The enantiomers of the entire set of new chiral azole compounds were separated using the three different mobile phase elution modes: normal phase, polar organic, and reversed phase. The most effective chiral stationary phase was the MaltoShell column, which was able to separate ten of the twelve compounds in one elution mode or another. Structural similarities in the newly synthesized oxazoles provided some insights into possible chiral recognition mechanisms.

Application of capillary electrophoresis technique for the enantioseparation of bioactive ferrocene-based compounds versus DFT calculated data.

Herein, a series of bioactive ferrocene-modified N-heterocycles with alkyl linkers was prepared in good to quantitative yields starting from easy accessible ferrocene alcohols and heterocycles under acidic or neutral (for imidazole) conditions in racemic forms. The analytical resolution of a number of bioactive racemic ferrocene azoles 1-6 (where azole = imidazole, pyrazole, and benzotriazole derivatives) into enantiomers was first carried out by CE using sulfobuthylether-ß-CD (captisol) as a chiral selector. The analytical approaches to highly enantiomeric-enriched ferrocene derivatives are based on the formation of their inclusion complexes. The best chiral separation was achieved using zone CE in a quartz capillary. The ACE was used to evaluate the stability constants of captisol complexes with enantiomeric forms of two ferrocene derivatives 1, FcCHMe-imidazole, and 6, FcCHMe-benzotriazole. The optimal conditions for the resolution of the studied (R, S)-ferrocene compounds 1, 2, and 6 were predicted on the basis of the performed quantum chemical calculations and then implemented by the electrophoretic method. A high correlation between density functional theory calculation results and experimental electrophoresis data were obtained. Successful enantioseparation of racemic mixtures is of great importance for the characterization and further applications of drug candidates in enantiopure forms and in the development of clinical treatment. The advantages of the CE procedure make it possible to have important practical value and significance for determining the purity and enantiomeric excess of other ferrocene-containing compounds.

Hydroxypropyl ß-cyclodextrin nanohybrid monoliths for use in capillary electrochromatography with UV detection: application to the enantiomeric separation of adrenergic drugs, anticholinergic drugs, antidepressants, azoles, and antihistamine.

Two kinds of hydroxypropyl ß-cyclodextrin nanohybrid monoliths were synthesized and applied in capillary electrochromatography with UV detection. One column was fabricated by concurrently using glycidyl methacrylate-bonded hydroxypropyl ß-cyclodextrin (GMA-HP-ß-CD), sodium 3-mercaptopropanesulphonate, and alkoxysilanes in the "one-pot" process. The other was prepared by free radical polymerization of GMA-HP-ß-CD, vinylmethylcyclosiloxane, ethylene dimethacrylate, and 2-acrylamido-2-methyl propane sulfonic acid. Compared to the former hybrid monolith, the latter one displayed improved enantiomeric separation. For ten adrenergic drugs, six anticholinergic drugs, two antidepressants, six azoles, and one antihistamine enantiomeric separation was obtained on the monolith synthesized by free radical polymerization. Twelve out of twenty-five drugs were baseline-separated. Especially, anisodamine with two chiral centers was successfully separated with resolution values of 3.06, 2.11, and 2.17. The nanohybrid monoliths were characterized by optical microscopy, scanning electron microscopy, FT-IR, nitrogen adsorption analysis, and thermogravimetric analysis. Relative standard deviation values less than 5% were obtained through run-to-run, day-to-day, and column-to-column investigations (n = 3). Graphical abstract Schematic representation of two kinds of hydroxypropyl ß-cyclodextrin nanohybrid monoliths based on "one-pot" approach (route I) and free radical polymerization approach (route II), respectively.

Dummy molecularly imprinted microspheres prepared by Pickering emulsion polymerization for matrix solid-phase dispersion extraction of three azole fungicides from fish samples.

Dummy molecularly imprinted microspheres (DMIMs) were synthesized by Pickering emulsion polymerization and used as the matrix solid-phase dispersion extraction (MSPD) sorbent for sample pre-treatment of azole fungicides in fish samples. Alpha-(2,4-Dichlorophenyl)-1H-imidazole-1-ethanol (DCE) was used as the fragment dummy template for the imprinting of climbazole (CBZ), clotrimazole (CMZ) and miconazole (MNZ). The morphology of the microspheres were characterized by scanning electron microscopy (SEM) and nitrogen adsorption measurements, narrow diameter distribution (20-50 µm) with regular spherical shape and high surface area (SBET = 408.91 ± 6.72 m2 g-1) were achieved. Good class-selectivity of the DMIMs was found for CBZ, CMZ and MNZ by static adsorption experiments. The imprinted microspheres as MSPD sorbent was then evaluated for the extraction and purification of CBZ, CMZ and MNZ in fish samples. The extracted azole fungicides were detected by HPLC-DAD analysis at 225 nm. MSPD conditions including elution, mass ratio of sample/sorbent and washing were carefully evaluated. The optimized MSPD method have good recoveries (89.2-101.5%) and reproducibility (RSDs 1.6-4.8%, n = 5) for fish samples spiked at 0.5, 2.5 and 12.5 µg g-1. The limits of detection (LODs) were 0.045, 0.036 and 0.033 µg g-1 for CBZ, CMZ and MNZ, respectively. The results show that this method has a good application prospect for the pretreatment of azole fungicides in fish samples.

A fully derivatized 4-chlorophenylcarbamate-ß-cyclodextrin bonded chiral stationary phase for enhanced enantioseparation in HPLC.

This paper reports an effective approach for the fabrication of a per-4-chlorophenylcarbamate-ß-cyclodextrin (ß-CD) bonded chiral stationary phase (CPCDP) in high-performance liquid chromatography. The morphology and structure of the ligand and the chiral stationary phase (CSP) were characterized by scanning electron microscopy, transmission electron microscopy, solid state 13C nuclear magnetic resonance spectra, fourier transform infrared spectra, elemental analysis and thermogravimetric analysis. Because CPCDP was a kind of multimode enantioseparation materials, the enantioseparation of chiral compounds including twelve azole antifungal agents, five proton pump inhibitors and five dihydropyridine calcium antagonists were studied in both reversed-phase and normal-phase chromatography. All analytes were obtained enantiomeric separation. Especially, the resolution of azoles was excellent. The selectivity and resolution of voriconazole reached 15.41 and 16.80, which was an exciting achievement for the enantioseparations by ß-CD based chiral stationary phases. Compared with the commercial 3,5-dimethylphenyl carbamate-ß-CD based chiral stationary phase (DMP), enhanced enantioselectivities for all the above compounds (except ilaprazole) were obtained on CPCDP column, which indicated that the 4-chlorophenylcarbamate group was conducive to the chiral recognition. Chromatographic studies elucidated that enhancement of analyte-chiral substrate interactions were attributed to the inclusion complexation, π-π stacking interaction, hydrogen-bonding, dipole-dipole interaction and steric hindrance. For further study, we also prepared semi-preparative chromatographic columns to obtain a single enantiomer. In addition to excellent chromatographic performance, the prepared CD-based column is stable and much cheaper than commercial columns, which can reduce the cost of test and has a good application prospect in chiral drug analysis.

Synergistic Anti-Candida Activity of Bengazole A in the Presence of Bengamide A †.

Bengazoles A⁻G from the marine sponge Jaspis sp. exhibit potent in vitro antifungal activity against Candida spp. and other pathogenic fungi. The mechanism of action (MOA) of bengazole A was explored in Candida albicans under both liquid culture and surface culture on Mueller-Hinton agar. Pronounced dose-dependent synergistic antifungal activity was observed with bengazole A in the presence of bengamide A, which is also a natural product from Jaspis sp. The MOA of bengazole A was further explored by monitoring the sterol composition of C. albicans in the presence of sub-lethal concentrations of bengazole A. The GCMS of solvent extracts prepared from liquid cultures of C. albicans in the presence of clotrimazole-a clinically approved azole antifungal drug that suppresses ergosterol biosynthesis by the inhibition of 14α-demethylase-showed reduced cellular ergosterol content and increased concentrations of lanosterol and 24-methylenedihydrolanosterol (a shunt metabolite of ergosterol biosynthesis). No change in relative sterol composition was observed when C. albicans was cultured with bengazole A. These results eliminate an azole-like MOA for the bengazoles, and suggest that another as-yet unidentified mechanism is operative.

Repurposing existing drugs: identification of irreversible IMPDH inhibitors by high-throughput screening.

Inosine 5'-monophosphate dehydrogenase (IMPDH) is an essential enzyme for the production of guanine nucleotides. Disruption of IMPDH activity has been explored as a therapeutic strategy for numerous purposes, such as for anticancer, immunosuppression, antiviral, and antimicrobial therapy. In the present study, we established a luciferase-based high-throughput screening system to identify IMPDH inhibitors from our chemical library of known bioactive small molecules. The screening of 1400 compounds resulted in the discovery of three irreversible inhibitors: disulfiram, bronopol, and ebselen. Each compound has a distinct chemical moiety that differs from other reported IMPDH inhibitors. Further evaluation revealed that these compounds are potent inhibitors of IMPDHs with kon values of 0.7 × 104 to 9.3 × 104 M-1·s-1. Both disulfiram and bronopol exerted similar degree of inhibition to protozoan and mammalian IMPDHs. Ebselen showed an intriguing difference in mode of inhibition for different IMPDHs, with reversible and irreversible inhibition to each Cryptosporidium parvum IMPDH and human IMPDH type II, respectively. In the preliminary efficacy experiment against cryptosporidiosis in severe combined immunodeficiency (SCID) mouse, a decrease in the number of oocyst shed was observed upon the oral administration of disulfiram and bronopol, providing an early clinical proof-of-concept for further utilization of these compounds as IMPDH inhibitors.

Comparative studies of immobilized chiral stationary phases based on polysaccharide derivatives for enantiomeric separation of 15 azole compounds.

Immobilized polysaccharide-based columns showed excellent enantioselectivity in normal phase separation mode. In this work, enantioseparation abilities of four immobilized polysaccharide-derived chiral stationary phases (Chiralpak IA, Chiralpak IB, Chiralpak IC, and Chiralpak ID) toward 15 azole compounds were evaluated. Separation was carried out using n-hexane as mobile phase with ethanol, 1-propanol, 1-butanol, and 2-propanol as modifiers. And twelve compounds have achieved baseline separation with the resolutions ranging between 2.05 and 21.73. The enantioseparation on the four polysaccharide-based chiral columns using different alcohol modifiers was compared. In general, the best separation performance was identified as Chiralpak IC, which was able to resolve 11 compounds to baseline and two partially under the screening conditions. Separation on Chiralpak IB was not satisfactory, because only four compounds were baseline separated.

Cyclic azole-homologated peptides from Marine sponges.

This review discusses the chemistry of cyclic azole-homologated peptides (AHPs) from the marine sponges, Theonella swinhoei, other Theonella species, Calyx spp. and Plakina jamaicensis. The origin, distribution of AHPs and molecular structure elucidations of AHPs are described followed by their biosynthesis, bioactivity, and synthetic efforts towards their total synthesis. Reports of partial and total synthesis of AHPs extend beyond peptide coupling reactions and include creative construction of the non-proteinogenic amino acid components, mainly the homologated heteroaromatic and α-keto-ß-amino acids. A useful conclusion is drawn regarding AHPs: despite their rarity, exotic structures and the potent protease inhibitory properties of some members, their synthesis is under-developed and beckons solutions for outstanding problems towards their efficient assembly.

Construction of a hydrazone-linked chiral covalent organic framework-silica composite as the stationary phase for high performance liquid chromatography.

Covalent organic frameworks (COFs), as an emerging class of crystalline porous organic polymers, have great potential for applications in chromatographic separation owning to their fascinating crystalline structures and outstanding properties. However, development of COF materials as novel stationary phases in high performance liquid chromatography (HPLC) is just in its infancy. Herein, we report the design and construction of a new hydrazone-linked chiral COF, termed BtaMth COF, from a chiral hydrazide building block (Mth) and present a one-pot synthetic method for the fabrication of BtaMth@SiO2 composite for HPLC separation of isomers. The as-synthesized BtaMth chiral COF displays good crystallinity, high porosity, as well as excellent chemical stability. Meanwhile, the fabricated HPLC column by using BtaMth@SiO2 composite as the new stationary phase exhibits high resolution performances for the separation of positional isomers including nitrotoluene and nitrochlorobenzene, as well as cis-trans isomers including beta-cypermethrin and metconazole. Additionally, some effects such as the composition of the mobile phase and column temperature for HPLC separations on the BtaMth@SiO2 packed column also have been studied in detail. The successful applications indicate the great potentials of hydrazone-linked chiral COF-silica composite as novel stationary phase for the efficient HPLC separation.

Total synthesis of the azolemycins.

The first total syntheses of newly isolated polyazole natural products azolemycins A-D, along with the synthesis of the tetra-oxazole non-natural analogue, are described.

Effects of organic modifier and temperature on the enantiomeric separation of several azole drugs using supercritical fluid chromatography and the Chiralpak AD column.

The effects of organic modifier and temperature on the enantioseparation of 10 triazoles and eight imidazoles, using supercritical fluid chromatography with the Chiralpak AD column, have been investigated in this work. For this purpose four different organic modifiers (methanol, ethanol, 2-propanol and acetonitrile) were evaluated. Only in the case of two compounds could the enantiomeric separation not be achieved with any of the modifiers tested; the rest of compounds were baseline or partially resolved with at least one of the modifiers. The alcohol-type modifiers provided the best results in terms of retention time and resolution. In general, retention increased in the order methanol < ethanol < 2-propanol; moreover it was possible to establish a relationship between the retention and the number of aromatic rings and dioxolane groups in the molecule, that is, the higher the number is, the higher the retention time. From the study of the temperature effect, the enthalpy-entropy compensation was demonstrated for all the compounds, except for bifonazole using methanol and miconazole using acetonitrile. This suggested that both analytes are enantiomerically resolved through different mechanisms.

Combined use of hydroxypropyl-ß-cyclodextrin and ionic liquids for the simultaneous enantioseparation of four azole antifungals by CE and a study of the synergistic effect.

A CE method employing a dual system of hydroxypropyl-ß-cyclodextrin (HP-ß-CD) and ionic liquids (ILs) has been developed for the simultaneous enantioseparation of four azole antifungals for the first time. In this study, three different types of ILs were employed as modifiers and among them dodecyl trimethyl ammonium chloride was found to be the most effective. The effects of the concentration, cations, and anions of ILs on the enantioseparation were investigated. With the developed dual system, all the enantiomers were well separated in resolutions of 3.8, 3.5, 2.8, and 2.5 for miconazole, econazole, ketoconazole, and itraconazole, respectively. The interactions between dodecyl trimethyl ammonium chloride and HP-ß-CD were also studied using a neutral polyacrylamide coated capillary and (1) H NMR spectroscopy to further explore the synergistic effect involved. It was found that ILs improved the enantioseparation not only by changing the EOF, but also by interactions with HP-ß-CD that could change its ability of forming inclusion complex with the enantiomers.

Prediction of the lipophilicity of nine new synthesized selenazoly and three aroyl-hydrazinoselenazoles derivatives by reversed-phase high performance thin-layer chromatography.

Using reversed-phase high-performance thin-layer chromatography and a methanol-water mixture as the mobile phase, the lipophilicity of 12 new synthesized derivatives is studied. The first eight compounds have as a basic chemical structure aryliden-hydrazino-selenazoles, and the second group of the three compounds belongs to aroyl-hydrazinoselenazoles. The linear correlation between R(Mw) and the methanol-water ratios showed high values for the correlation coefficient. The chromatographic hydrophobic index is determined by using the ratio -R(Mw)/S, and the obtained values ranged between 99 and 73. A good linear correlation is obtained between R(Mw) and the slope. The log P values are calculated using ACD/Labs Software. The matrices are formed with R(Mw) and log P and are subjected to a principal component analysis (PCA). The best way to extract information from PCA is graphically, by plotting the obtained matrices. By analyzing the scores, the compounds can be grouped as follows: a group containing nine compounds, and a second one containing three compounds. Each group of compounds has the same basic chemical structure.

Selenium accumulation in mycelia of Flammulina velutipes during fermentation determined by RP-HPLC.

A method to estimate the content of selenium in organics was introduced based on reversed phase-high performance liquid chromatography (RP-HPLC).The maximum absorption peak of piazselenol was at 330 nm and the optimized temperature and pH value were 40 degrees C and 2.8, respectively. The minimum detection concentration of selenium(IV) was 0.06 microg/mL and the measurable range was 0.12-12.0 microg/mL. The organic selenium accumulation in golden needle mushroom (Flammulina velutipes) mycelia was obtained by subtracting the amount of inorganic selenium from that of total selenium. The organic selenium accumulation of various inoculation amounts showed that organic selenium accumulation in a unit volume of the fermentation broth was positively related the inoculation amount. Compared with the methods reported previously, the method used here is simple, reliable and less toxic.

In search of the holy grail of antifungal therapy.

The ideal antifungal agent remains an elusive goal for treatment of life-threatening systemic fungal infections. Such an agent would have broad antifungal activity, low rates of resistance, flexible routes of administration, few associated adverse events, and limited drug-drug interactions. Only three of the seven classes of antifungal agents currently available are suitable for treatment of systemic infection: the polyenes, the azoles, and the echinocandins. None match all the characteristics of an ideal agent, the Holy Grail of antifungal therapy. Academia and industry need to collaborate in the search for new lead antifungal compounds using traditional screening methods as well as the new pharmacogenomics methods. Enhancing efficacy and reducing toxicity of the currently available therapeutic agents is also another important avenue of study. As an example, the Mycosis Research Center at the University of Mississippi Medical Center has identified pyogenic polyenes in commercial preparations of amphotericin B deoxycholate which correlate with infusion related toxicities. A highly purified formulation of amphotericin B appears promising, with a better therapeutic index compared to its parent compound as evidenced by results of in vitro and in vivo studies reviewed in this presentation.

Enantioselective separation of azole compounds by EKC. Reversal of migration order of enantiomers with CD concentration.

The enantioselective separation of a group of six weak base azole compounds was achieved in this work using EKC with three neutral beta-CDs as chiral selectors. The native beta-CD and two other beta-CD derivatives with different types and positions of the substituents on the CD rim ((2-hydroxy)propyl-beta-CD (HP-beta-CD) and heptakis-2,3,6-tri-O-methyl-beta-CD (TM-beta-CD)) were employed. Apparent binding constants for each pair compound-CD were determined in order to study analyte-CD interactions. The best enantiomeric resolutions for miconazole, econazole, and sulconazole were observed with HP-beta-CD whereas for the separation of the enantiomers of ketoconazole, terconazole, and bifonazole, TM-beta-CD was the best chiral selector. The enantioseparations obtained were discussed on the basis of the structure of the compounds taking into account that inclusion into the hydrophobic CD cavity occurred through the phenyl ring closer to the azole group. In addition, a change in the migration order for the enantiomers of two of the compounds studied (ketoconazole and terconazole) with the concentration of HP-beta-CD was observed for the first time.

Enantiomeric separation of several antimycotic azole drugs using supercritical fluid chromatography.

The chiral resolution of four antifungal compounds, three imidazoles (miconazole, econazole and sulconazole) and one triazole (itraconazole) using supercritical fluid chromatography on the amylose-based chiral stationary phase Chiralpak AD, is presented in this work. The influence of pressure, type and percentage of organic modifier and temperature on retention times and resolution was studied. The enantiomeric separation of the three imidazoles was achieved with resolutions higher than two and analysis times lower than 10 min, obtaining the best results using methanol as modifier. However, the analysis time of the triazole was higher than 80 min due to the existence of a high number of functional groups that were able to interact with the chiral stationary phase. In this case, the resolution of the four stereoisomers was achieved only partially with mixtures of ethanol and 2-propanol as modifier. The isoenantioselective temperatures were obtained from the study of the influence of the temperature, they were above the range of temperatures assayed, except for sulconazole using 2-propanol.

Marine metabolites: metal binding and metal complexes of azole-based cyclic peptides of marine origin.

Azole-based cyclic peptides found in ascidians ("sea squirts") of the genus Lissoclinum have a high propensity to chelate metal ions. This Highlight summarises the current evidence for marine cyclic peptide-metal congruence, and the structural and stereochemical features in cyclic peptides which seem necessary to facilitate metal complexation. The biological relevance of the metal ions in these associations, including their possible role in the assembly of cyclic peptides in the marine milieu, is also briefly considered. Finally, the synthesis of natural, and some novel non-natural, azole-based cyclic peptides from the cyclooligomerisation and assembly of azole-based amino acids, including in the presence of metal ions, is presented.