Synthesis of cholaphanes by ring closing metathesis

The synthesis of cholaphanes by ring closing metathesis (RCM) of 3α,7α,12α,24-tetraol allyl derivatives, obtained from cholic acid, was attempted. The reactions of tetraol 3,24-diallyl ether or 3,24-diacrylate were not satisfactory. However, diallyl derivatives of disteroidal 3,3′- or 24,24′-ortho-phthalates reacted smoothly affording cyclic dimers in good yields. In all the reactions studied, the E isomers of the macrocycles were obtained in excess.     

Rapid formation and slow collapse of a carbocation-anion pair to a neutral molecule

The 4,4',4' '-trimethoxytrityl cation (TMT(+)) was observed to react with acetate ion in acetic acid reversibly to give the corresponding ester (TMT-OAc). The rate of the reaction was found to be independent of [NaOAc] over a 25000-fold range. Similar results were observed in the presence of Bu(4)N(+) in acetic acid as well as in HOAc/AN (1/1). It was concluded that {TMT(+) (HOAc/AcO(-))} is an ion pair that forms essentially completely from free TMT(+) and HOAc/AcO(-) during the time of mixing under stopped-flow conditions. The process which was studied kinetically is the intramolecular collapse of the ion pair to TMT-OAc which takes place in two steps involving a kinetically significant intermediate. The remarkably close resemblance of this reaction to the Winstein scheme for solvolysis reactions is noted. In analogy to the Winstein scheme, it was proposed that the intermediate could be an intimate ion pair formed upon extrusion of solvent from the solvent separated ion pair. The product-forming step could then correspond to the intimate ion pair reacting further to form a covalent bond between the two moieties within the complex. The values of the thermodynamic and the activation parameters as well as the apparent rate constants for the reaction in the presence of either sodium or tetrabutylammonium ions suggest that these counterions play insignificant roles in the reactions. However, the equilibrium constant for the intramolecular step (K(4)) was observed to be two times greater in the presence of Bu(4)N(+) than in the presence of Na(+). The rate of the reaction in HOAc was observed to be about four times as great as that in HOAc/AN (1/1).     

Novel Deep Cavity Calix[4]pyrroles Derived from Steroidal Ketones

Novel steroidal calix[4]pyrroles were prepared in excellent yields from commercially available cholic acid derivatives using an efficient synthetic sequence. Once in hand, it was found that these calix[4]pyrroles exist in the form of four different configurational isomers. Separation of these isomers was achieved readily using normal phase HPLC techniques. Once purified, the steroidal calix[4]pyrroles were screened via -FABMS analyses to judge their utility in effecting the enantioselective recognition of appropriate organic anions. Results that provided support for antipodal R > S selectivity were obtained in the case of both tartaric acid and mandelic acid. Direct extraction studies were then carried out and these confirmed the pattern of R > S selectivity observed by -FABMS.     

Determination of critical micellar concentrations of cholic acid and its keto derivatives

The critical micellar concentration (CMC) values of keto derivatives of cholic acid (3,12-dihydroxy-7-oxo-5β-cholanoic acid, 3,7-dihydroxy-12-oxo-5β-cholanoic acid, 12-hydroxy-3,7-dioxo-5β-cholanoic acid, 3-hydroxy-7,12-dioxo-5β-cholanoic acid, 3,7,12-triketo-5β-cholanoic acid) and cholic acid itself, were determined. Replacement of hydroxyl groups in cholic acid molecule with keto groups yields the derivatives whose CMC values increase with increase in the number of keto groups introduced. The CMCs of derivatives with the same number of keto groups but at different positions do not differ significantly. The relationship between the number of keto groups in the molecule of cholic acid keto derivatives and CMC value can be described by the following equation: CMC = 43 number of keto groups + 14.667. The effect of NaCl concentration on CMC increases with increase in the number of keto groups.     

Oxandrastins: Antibacterial Meroterpenes from an Australian Mud Dauber Wasp Nest-Associated Fungus,Penicillium sp. CMB-MD14

The ethyl acetate extract of an ISP-2 agar cultivation of the wasp nest-associated fungus Penicillium sp. CMB-MD14 exhibited promising antibacterial activity against vancomycin-resistant enterococci (VRE), with a bioassay guided chemical investigation yielding the new meroterpene, oxandrastin A (1), the first andrastin-like metabolite with an extra oxygenation at C-2. A culture media optimisation strategy informed a scaled-up rice cultivation that yielded 1, together with three new oxandrastins B–D (2–4), two known andrastins C (5) and F (6), and a new meroterpene of the austalide family, isoaustalide F (7). Structures of 1–7 were assigned based on detailed spectroscopic analysis and chemical interconversion. A GNPS molecular networking analysis of the rice cultivation extract detected the known austalides B (8), H (9), and H acid (10), tentatively identified based on molecular formulae and co-clustering with 7. That the anti-VRE properties of the CMB-MD14 extract were exclusively attributed to 1 (IC₅₀ 6.0 µM, MIC₉₉ 13.9 µM), highlights the importance of the 2-OAc and 3-OAc moieties to the oxandrastin anti-VRE pharmacophore.     

Simultaneous detection of cholyl adenylate and coenzyme A thioester utilizing liquid chromatography/electrospray ionization mass spectrometry.

It has been proposed that acyl adenylate is first formed during activation of the carboxy group into the acyl CoA thioester, an intermediate in the formation of amino acid conjugates. Acyl CoA synthetases may be responsible for this acyl adenylate formation. Recently, we hypothesized the preferential formation of cholic acid adenylate, a major bile acid, preceding production of the corresponding CoA thioester in incubations with rat liver microsomal fractions. To verify this biosynthetic mechanism, monitoring of the incubation mixture of acyl adenylate together with both substrate and acyl CoA thioester is needed. We have developed a detection method for the simultaneous detection of these cholic acid derivatives utilizing liquid chromatography/electrospray ionization mass spectrometry. The CoA thioester of cholic acid forms a chelation complex with the divalent cations remaining on the silica gel packed into the analytical column. Both the addition of a chelating agent, such as EDTA, to the mobile phase and an adjustment of the mobile phase pH to a weak alkaline effectively removed such chelate formation, producing a sharp CoA thioester peak. For a simultaneous mass spectrometric analysis of cholic acid, the corresponding adenylate and CoA thioester, the combined use of a 300 A particle diameter ODS column and 20 mM ammonium acetate buffer (pH 9.0)/2-propanol/acetonitrile as the mobile phase have been proved to be preferable. To avoid any degradation of the chemically unstable adenylate produced in the incubation, we employed a direct injection of the sample onto a preconcentration column. The obtained results indicated a high sensitivity of this method.     

Synthese d'antibiotiques triterpeniques a partir d'acides biliaires-II : Introduction du methyle 14α associe á Δ-8,9

The synthesis of methyl 14α-methyl-15-oxo-5β-chol-8-en-24-oate derivatives from cholic acid is described, as are the spectroscopic data of the various intermediates. A route to tetracyclic triterpene acids related to polyporenic acid is thus opened.     

Studies directed toward synthesis of quassinoids. VI—mass spectral fragmentation of D-ring seco esters and δ-lactone derivatives of cholic acid

Analogy and deuterium labeling demonstrate that considerable remote group interaction occurs between functional groups at positions 7 and 16 in 16, 17-secosteroids. It is proposed that ion-dipole interact?ions separations of distances of 10 to 2 Å will involve energies of 1–30 kcal mol^?1 and can play a significant role in fragmentation processes. Successive loss of one, two and three HOAc molecules from a milecular or daughter ion increase progressively in relative intensity; irregularity in this pattern indicates the operation of special factors or multiple genesis of daughter ions having the same m/e. Evidence is presented which suggests that facile ketene ejection from the acetate functional group only occurs from a radical ion and must be preceded by transfer of a hydrogen radical from the acetate group to the site of charge localization.     

Mechanism of nucleophilic substitution in chromium(III) complexes. II. Formation of Acetatopentamminechromium(III)-ion from aquopentamminechromium(III)-ion in acetic acid — sodium acetate buffer media

The rate of formation of the acetatopentamminechromium(III)ion from the aquopentammine complex in HOAc? NaOAc buffer media has been investigated spectrophotometrically. The results suggest that the reaction occurs by two concurrent paths one of which is independent of acetate ion while the other is first order with respect to acetate ion concentration. The values of the rate constants for both the steps and the corresponding activation parameters, ΔH^≠ and ΔS^≠, have been evaluated. The results are consistent with an S_N1 mechanism for the acetate independent path and an S_N1 IP mechanism for the acetate dependent path. Evidence for ion-pair formation is given.     

A Fluorescence Polarization Assay of Erythrocyte Membrane Fluidity on the Abbott TDx Analyzer

Four fluorescein dye derivatives (attached to cholic acid, deoxycholic acid, lithocholic acid, and 5-cholenic acid-3β-ol) were studied as a means of detecting alterations in erythrocyte membrane fluidity. In addition, membrane fluidity using these novel dye derivatives was correlated to nonenzymatic glycation of erythrocyte membrane ghosts. These dyes have excitation and emission wavelengths suitable for analysis on an Abbott TDx analyzer. Both the lithocholic acid and the 5-cholenic acid-3β-ol fluorescein derivatives were capable of detecting erythrocyte membrane fluidity with an increase in polarization of approximately 34% over the range of glycation studied, and the fluidity was strongly correlated with the degree of glycation. The cholic acid and deoxycholic acid fluorescein derivatives were unable to enter the erythrocyte membrane since no alteration in fluorescence intensity or polarization was observed upon incubation with the membrane preparations. The results obtained indicate that the lithocholic acid and 5-cholenic acid-3β-oI fluorescein derivatives are potential new reagents for the development of routine assays for membrane fluidity on the Abbott TDx analyzer.     

Asymmetric ring opening of terminal epoxides via kinetic resolution catalyzed by chiral (salen)Co mixture

The highly enantioselective hydrolytic kinetic resolution (HKR) of racemic terminal epoxides by bimetallic chiral (salen)Co and (salen)Co(III)-OAc mixture provides a simple and effective method for the synthesis of enantiomerically enriched terminal epoxides (ee > 99%) and diols. At the equimolar amounts of bimetallic chiral (salen)Co and (salen)Co(II)-OAc, the catalytic activity increases more than two times in comparison with (salen)Co(III)-OAc used alone. The mixed catalytic system can be recycled and reused. No significant loss of catalytic activity was observed after three runs.     

Studies on steroids. CCXXVIII Trace analysis of bile acids by gas chromatography-mass spectrometry with negative ion chemical ionization detection

A suitable derivatization method for the trace analysis of bile acids by gas chromatography (GC) in combination with negative ion chemical ionization (NICI) mass spectrometry is described. Of various derivatives for the carboxyl group, the pentafluorobenzyl (PFB) ester provided the highest value of the ratio of the negative to the positive ion current. A characteristic carboxylate anion [M - 181]- was produced as the most abundant ion by the loss of the PFB group in NICI. The PFB esters were further derivatized to the dimethylethylsilyl (DMES) ethers, whereby lithocholic acid, deoxycholic acid, chenodeoxycholic acid, ursodeoxycholic acid and cholic acid were distinctly separated by GC on a cross-linked methyl silicone fused-silica capillary column. The detection limit for the PFB-DMES derivatives of dihydroxylated bile acids was 2 fg when the fragment ion was monitored at m/z 563 in the NICI mode using isobutane as a reagent gas.     

Synthesis and spectroscopic studies of new bile acid derivatives linked by a 1,2,3-triazole ring

A novel method for the synthesis of cholic acid derivatives has been developed using ‘click chemistry’. Intermolecular 1,3-dipolar cycloaddition of the propargyl ester and azide groups of 3α-azidoacetoxy-7α,12α-diformyloxy-5β-cholan-24-oate gave a new dimer and oligomer linked by a 1,2,3-triazole ring. The structures of the products were confirmed by spectral (¹H NMR, ¹³C NMR, and FT-IR) analysis, mass spectrometry and PM5 semiempirical methods. Estimation of the pharmacotherapeutic potential has been accomplished for the synthesized compounds on the basis of Prediction of Activity Spectra for Substances (PASSs).     

Hydrogen bonding of single acetic acid with water molecules in dilute aqueous solutions

In separation processes, hydrogen bonding has a very significant effect on the efficiency of isolation of acetic acid (HOAc) from HOAc/H₂O mixtures. This intermolecular interaction on aggregates composed of a single HOAc molecule and varying numbers of H₂O molecules has been examined by using ab initio molecular dynamics simulations (AIMD) and quantum chemical calculations (QCC). Thermodynamic data in aqueous solution were obtained through the self-consistent reaction field calculations and the polarizable continuum model. The aggregation free energy of the aggregates in gas phase as well as in aqueous system shows that the 6-membered ring is the most favorable structure in both states. The relative stability of the ring structures inferred from the thermodynamic properties of the QCC is consistent with the ring distributions of the AIMD simulation. The study shows that in dilute aqueous solution of HOAc the more favorable molecular interaction is the hydrogen bonding between HOAc and H₂O molecules, resulting in the separation of acetic acid from the HOAc/H₂O mixtures with more difficulty than usual.     

The mass spectrometric fragmentation of n-heptane

The electron impact fragmentation of n-heptane has been investigated using ¹³C labelled derivatives. A mechanism is proposed for the loss of alkyl radicals where the cleavage of a C? C bond is coupled with the rearrangement of a hydrogen atom, thus yielding a secondary alkyl ion that eventually fragments further by a subsequent loss of olefin. For alkyl ions with less than six carbon atoms this consecutive pathway is in competition with formation directly from the molecular ion. The consecutive pathway contributes about 15% to the intensity of the alkyl ions with four and five carbon atoms and 80% for smaller ions. The electron energy dependence was studied.     

Effect of Quinones on the Cobalt(II) Acetate–Catalyzed Mild Oxidation of Cyclohexane with Hydrogen Peroxide in Acetic Acid

The effects of free-radical reaction inhibitors (InH), hydroquinone (HQ) and quinone (Q), on the oxidation of cyclohexane catalyzed by cobalt(II) acetate Co(OAc)₂ · 4H₂O and on the decomposition of hydrogen peroxide in acetic acid (HOAc) at 303 K were studied. It was found that an increase in the concentration of HQ in the starting reaction mixture containing cyclohexane, the catalyst, and H₂O₂ dissolved in HOAc resulted in an exponential decrease in the yields of the target products of oxidation: cyclohexanol, cyclohexanone, and cyclohexyl hydroperoxide. In the presence of Q, the dependence of the yield of the target products on the initial inhibitor concentration exhibited a maximum at (1.8–2.5) × 10^–2 M Q. At (2.2–2.4) × 10^–2 M Q concentrations, the yield of the target products was 55–60% of that in an uninhibited process. Based on kinetic, spectrometric, and quantum-chemical data, the effect found was explained by the fact that under the experimental conditions highly active hydroxyl derivatives of radicals rather than a hydroxy quinolide hydroperoxide (the homolysis of which can produce species with a free valence, which are capable of initiating free-radical reactions) were largely formed from Q.     

Metal coordination as a tool for controlling the self-assembling and gelation properties of novel type cholic amide-phenanthroline gelating agent

(3α,7α,12α)-Trihydroxy-N-[1,10-phenanthrolin-5-yl]-5β-cholan-24-amide was found to be a powerful gelating agent for methanol-water in gelator to solvent ratio starting from 0.1% in absence of metal ion. Formation of phenanthroline-zinc (II) 2:1 complex changes dramatically gelating properties; when stored, it dissolves into clear solution without Tyndall effect and this solution, when heated to ca. 70°C reversibly forms a gel again, without chemical change as proven by NMR spectrometry. Structures of the gels of cholic amide-phenanthroline and its Zn²⁺ complex were studied by SEM.     

Transport behavior of four bile salt micelles and cholesterol solubilized by their micelles across porous membrane

 The transport behavior of bile salts (BSs) solubilizing cholesterol (Ch) or none across an artificial membrane was investigated for sodium salts of deoxycholic acid (NaDC), chenodeoxycholic acid (NaCDC), ursodeoxycholic acid (NaUDC) and cholic acid (NaC) in tetraborate–carbonate buffer solution at pH 10.0 and 37 °C. The study demonstrated that the surfactant properties such as critical micellization concentration (CMC) and micellar size or diffusion coefficient were determinable from the flux or permeability measurements. The comparison among the respective pure systems of BSs led to a conclusion that the micellar size was in the order of NaDC>NaCDC>NaUDC>NaC and determined CMC values were in agreement with those in literature. The magnitude of solubilizing power (capacity) of BS for Ch was found to decrease in the order of NaDC>NaCDC>NaC>NaUDC; this order is in accordance with that of the empirical hydrophobicity index. The hydrodynamic radii for the singly dispersed species and the micellar species of the respective BSs and of Ch-solubilizing micelles were estimated from the permeability data; the radii of the Ch-solubilizing micelles are approximately 12–15 Å and interestingly, smaller than those of the respective BS alone micelles ranging from 14 to 22 Å. Received: 15 April 1997 Accepted: 25 July 1997     

A single glucose derivative suitable for gas chromatography/mass spectrometry and gas chromatography/combustion/isotope ratio mass spectrometry

The incorporation of stable isotopes improves the assessment of glucose metabolism and, with some researchers using two tracers, (2)H-glucose assessed by gas chromatography/mass spectrometry (GC/MS) and (13)C-glucose by gas chromatography/combustion/isotope ratio mass spectrometry (GC/C/IRMS), a common derivative for both is advantageous. The most commonly used derivatives for GC/MS are inappropriate for GC/C/IRMS as additional functional groups dilute the label. We therefore considered the suitability of six derivatives for both GC/MS and GC/C/IRMS. Glucose alkylboronates were prepared by adding the appropriate alkylboronic acid (butyl- or methylboronic acid) in pyridine to desiccated glucose. The derivatisation was completed by reacting this with either (a) acetic anhydride or trifluoroacetic anhydride (acetate derivatives) or (b) bis(trimethylsilyl)trifluoroacetamide BSTFA (TMS derivatives). All six derivatives were assessed using GC/MS and (13)C GC/C/IRMS.Neither TMS derivative exhibited any signal intensity in the molecular ion, although a M-15 ion showed good agreement between experimental and theoretical data and, whilst still low in intensity, could be suitable for isotope work. Similarly, none of the acetate derivatives showed any intensity at the molecular ion although three key fragmentation series were identified. The most attractive sequence, initiated by the loss of 1,2 cyclic boronate, resulted in the main fragment ion of interest, m/z 240, corresponding to the fluorinated methylboronate derivate. Minimal carbon and hydrogen atoms are added to this derivative making it an excellent choice for stable isotope work, while proving suitable for analysis by both GC/MS and GC/C/IRMS.     

De Novo and Divergent Synthesis of Highly Functionalized Furans by Cascade Reactions of 2-Hydroxy-1,4-diones with Nucleophiles

Herein, a divergent synthesis of a variety of 2α- and 5α-substituted furan derivatives from 2-hydroxy-1,4-diones is reported. By using appropriate substrates and an acid catalyst, the reactions occurred selectively through cyclization/1,6-conjugate addition or cyclization/Friedel–Crafts-type cascade reactions. A broad range of nucleophilic reagents (>10 types for the 1,6-conjugate addition for 5α substitution and >20 types for the Friedel–Crafts-type cascade reaction for 2α substitution), including alcohols, amides, furan, thiophene, pyrrole, indole, phenols, and many others, can successfully participate in the reactions, providing a universal strategy for a diversity-oriented synthesis of α-substituted furan derivatives. Deuteriation experiments and DFT calculations were carried out to support the proposed reaction mechanisms. Antifungal activity experiments revealed that products with an indole or 4-hydroxycoumarin core substituted at the 2α position showed moderate activities against Rhizoctorzia solani and Botrytis cinerea, respectively.