Convergent synthesis, chiral HPLC, and vitamin D receptor affinity of analogs of 1,25-dihydroxycholecalciferol

A series of analogs of 1,25-dihydroxycholecalciferol was obtained with an additional chiral center at the terminus of the aliphatic side chain (C-25). The analogs were obtained from (+)-(R)- and (-)-(S)-2-methylglycidols, by opening of the oxirane ring with the carbanions derived from vitamin D C23a,24- or C22-sulfones. The diastereomeric purity of the analogs was determined by high-performance liquid chromatography on a chiral stationary phase. The binding affinity of analogs for the calf thymus intracellular vitamin D receptor (VDR) was two orders of magnitude lower than that of the lead compound of this group, 24a,24b-dihomo-1,25-dihydroxycholecalciferol, and it was comparable to the affinity of analogs of 24-nor-1,25-dihydroxycholecalciferol. However, a twofold difference was observed for analogs diastereomeric at C-25 in their affinity for VDR. The diastereodifferentiation of the binding affinity was found to be specific for vitamin D vicinal 25,26-diols as it disappears for analogs where 26-hydroxyl, neighboring the C-25 chiral center, is replaced with methyl.     

Chiral discrimination of branched-chain fatty acids by reversed-phase HPLC after labeling with a chiral fluorescent conversion reagent

Anteiso fatty acids having 16 to 29 carbon atoms were labeled with the chiral fluorescent conversion reagents, (1R,2R)- and (1S,2S)-2-(2,3-anthracenedicarboximido)cyclohexanol. The diastereomeric esters of anteiso acids having up to 20 carbon atoms were separated into two peaks in an ODS column under low column-temperature conditions, while those having more than 21 carbon atoms were not separated. A C30 column made it possible to separate diastereomeric esters up to C29 anteiso acid. It was possible to predict the absolute configuration of each acid by the elution order of the derivatives.     

Diphenylethanediamine (DPEDA) derivatives as chiral selectors: IV. A comparison of 3,5-dinitrobenzoylated (S,S)- and (S,R)-DPEDA-derived chiral stationary phases with Pirkle's standard (R)-phenylglycine-derived phase in normal phase HPLC

Undecanoyl bound 3,5-dinitrobenzoyl-(S,R)-1,2-diphenylethane-1,2-diamine [(1S,2R)-DNB-DPEDA] as chiral selector (SO) has been synthesized and used as a chiral stationary phase (CSP II) for normal-phase enantioselective HPLC. It is compared with the already published diastereomeric (1S,2S)-DNB-DPEDA-derived CSP I and with the “standard” Pirkle DNB-(R)-phenylglycine-derived CSP III. Chromatographic data for about 100 racemic analytes reveal that CSP II is able to separate especially well enantiomers of derivatized aromatic carboxylic acids and analytes having a benzyl substituent bound at the chiral center. However, CSP I was found to be superior to CSP II and III in its general applicability and its ability to resolve enantiomers of heterocyclic drugs. © 1994 Wiley-Liss, Inc.     

Deracemization of an axially chiral biphenylic derivative as a tool for investigating chiral recognition in self-assemblies

Two diastereomeric cationic surfactants derived from L-proline, in which the second chiral center is a quaternary nitrogen, have been separated and fully characterized. The recognition properties of the aggregates formed by the two diastereomeric surfactants have been investigated by circular dichroism and (1)H NMR through deracemization of racemic 2-carboxy-2'-dodecyloxy-6-nitrobiphenyl.     

Phospholipids chiral at phosphorus. Absolute configuration of chiral thiophospholipids and stereospecificity of phospholipase D

Separate diastereomers of 1,2-dipalmitoyl-sn-glycero-3- thiophosphoethanolamine ( DPPsE ) were prepared in 97% diastereomeric purity and characterized by 31P, 13C, and 1H nuclear magnetic resonance (NMR). The isomers hydrolyzed by phospholipases A2 and C specifically were designated as isomer B (31P NMR delta 59.13 in CDCl3 + Et3N ) and isomer A (59.29 ppm), respectively, analogous to the isomers B and A of 1,2-dipalmitoyl-sn-glycero-3- thiophosphocholine ( DPPsC ) [ Bruzik , K., Jiang , R.-T., & Tsai, M.-D. (1983) Biochemistry 22, 2478-2486]. Phospholipase D from cabbage was shown to be specific to isomer A of DPPsC in transphosphatidylation . The product DPPsE was shown to be isomer A. The absolute configuration of chiral DPPsE at phosphorus was elucidated by bromine-mediated desulfurization in H2 18O to give chiral 1,2-dipalmitoyl-sn-glycero-3-[18O]phosphoethanolamine ( [18O]DPPE) followed by 31 P NMR analysis [ Bruzik , K., & Tsai, M.-D. (1984) J. Am. Chem. Soc. 106, 747-754]. The absolute configuration of chiral DPPsC was elucidated by desulfurization in H2 18O mediated by bromine or cyanogen bromide to give chiral 1,2-dipalmitoyl-sn-glycero-3-[18O]phosphocholine ( [18O]DPPC), which was then converted to [18O]DPPE by phospholipase D with retention of configuration [ Bruzik , K., & Tsai, M.-D. (1984) Biochemistry (preceding paper in this issue)]. The results indicate that isomer A of both DPPsE and DPPsC is SP whereas isomer B is RP.     

Evaluation of chiral separation efficiency of a novel OTPTHE derivatization reagent: Applications to liquid‐chromatographic determination of DL‐serine in human plasma

A novel chiral derivatization reagent, the N‐[1‐oxo‐5‐(triphenylphosphonium)pentyl]‐ (R)‐1,3‐thiazolidinyl‐4‐N‐hydroxysuccinimide ester bromide salt (OTPTHE), was developed for the separation and selective detection of chiral DL‐amino acids by RP‐HPLC analysis. The OTPTHE reacted with DL‐amino acids at 60°C maintained for 30 minutes in the presence of 100 mM borate buffer (pH 9.5). The separability of the diastereomeric derivatives was evaluated in terms of the resolution value (Rs) using 13 kinds of DL‐amino acids, which were completely separated by reversed‐phase chromatography using C18 column at 254 nm. The Rs of the DL‐amino acids varied from 1.62 to 2.51. As for the application of the DL‐amino acids, the determination of DL‐Ser in the human plasma of healthy volunteers was performed based on our developed method. It was shown that linear calibrations were available with high coefficients of correlation (r² > 0.9997). The limit of detection (S/N = 3) of the DL‐Ser enantiomers was 5.0 pmol; the relative standard deviations of the intraday and interday variations were below 4.56%; the accuracy ranged between 95.40%‐110.06% and 95.45%‐109.80%, respectively; the mean recoveries (%) of the DL‐Ser spiked in the human plasma were 99.49%‐103.74%. The amounts of DL‐Ser in the human plasma of healthy volunteers were determined.     

Guest-dependent conformation of 18-crown-6 tetracarboxylic acid: relation to chiral separation of racemic amino acids

(+)-18-crown-6 tetracarboxylic acid (18C6H(4)) has been used as a chiral selector for various amines and amino acids. To further clarify the structural scaffold of 18C6H(4) for chiral separation, single crystal X-ray analysis of its glycine(+) (1), H3O+ (2), H5O2+ (3), NH4+ (4), and 2CH3NH3+ (5) complexes was performed and the guest-dependent conformation of 18C6H(4) was investigated. The crown ether ring of 18C6H4 in 3, 4, and 5 took a symmetrical C2 or C2-like conformation, whereas that in 1 and 2 took an asymmetric C1 conformation, which is commonly observed in complexes with various optically active amino acids. The overall survey of the present and related complexes suggests that the molecular conformation of 18C6H4 is freely changeable within an allowable range, depending on the molecular shape and interaction mode with the cationic guest. On the basis of the present results, we propose the allowable conformational variation of 18C6H4 and a possible transition pathway from its primary conformation to the conformation suitable for chiral separation of racemic amines and amino acids.     

The determination of enantiomer composition of 1‐((3‐chlorophenyl)‐(phenyl)methyl) amine and 1‐((3‐chlorophenyl)(phenyl)‐methyl) urea (Galodif) by NMR spectroscopy,chiral HPLC,and polarimetry

For the first time, a method for enantiomer resolution of the anticonvulsant Galodif (1‐((3‐chlorophenyl)(phenyl)methyl) urea) by chiral HPLC was developed, whereas the enantiomeric composition of 1‐((3‐chlorophenyl)(phenyl)methyl) amine—precursor in Galodif synthesis—cannot be resolved by this method. However, starting 1‐((3‐chlorophenyl)(phenyl)methyl) amine quantitatively forms diastereomeric N‐((3‐chlorophenyl)(phenyl)methyl)‐1‐camphorsulfonamides in reaction with chiral (1R)‐(+)‐ or (1S)‐(?)‐camphor‐10‐sulfonyl chlorides. The diastereomeric ratio of obtained camphorsulfonamides can be easily determined by NMR ¹H and ¹³C spectroscopy. The DFT calculations of specific rotation of Galodif enantiomers showed good agreement with experimental data. The absolute configuration of enantiomers was proposed for the first time.     

Kinetically controlled synthesis of monoglyceryl esters from chiral and prochiral acids methyl esters catalyzed by immobilized Rhizomucor miehei lipase

Partial acylation of only one primary hydroxyl group of glycerol generates a chiral center at position 2. Rhizomucor miehei lipase (RML) catalyzes the kinetically controlled transesterification of different aromatic carboxylic acids methyl esters with glycerol. High synthetic yields of glyceryl esters (around 70-80%) were obtained even in the presence of significant concentrations of water (from 5% to 20%). After a long incubation of the reaction mixture in the presence of the biocatalyst only pure free acid was obtained. Other lipases (from Geobacillus thermocatenulatus and from Thermomyces lanuginose) also catalyzed similar kinetically controlled transesterifications although less efficiently. RML immobilized on Sepharose-Q showed a high activity and specificity, compared to the immobilization by other techniques, only producing monoglyceryl esters with all substrates. In particular, monoglyceryl-phenylmalonate product was synthesized in 82% overall yield and >99% diastereomeric excess at pH 7.0 and 37 °C and 90% glycerol.     

Effect of the mobile phase on the retention behavior of optical isomers of the mandelic acid derivative methyl 2-phenyl-2-(tetrahydropyranyloxy) acetate by chiral HPLC

Conditions for separation of enantiomers of a mandelic acid derivative, methyl 2-phenyl-2-(tetrahydropyranyloxy) acetate (the analyte) were studied. Because of the presence of two chiral carbons, the analyte consists of four stereoisomers stable at ambient temperature. Chiral HPLC of the analyte resulted in four peaks, using an (S,S)-Whelk-O1 column with the mobile phase consisting of hexane and the t-butyl methyl ether (TBME). It was found that TBME dramatically changed the retention of the isomers, though it produced the best enantioseparation on (S,S)-Whelk-O1. The amount of TBME in the mobile phase influenced the degree of retention shift; 5% (v/v) TBME gave a bigger shift than 8% (v/v) and 10% (v/v). 2-Propanol did not produce the same results. The chiral separation was also tried on cellulose tris (3, 5-dimethyl phenylcarbamate) (CDMPC), but only three peaks were seen, indicating some but not full enantiomer resolution.     

1,3,5-Triazine multiselector systems: New tools for chiral discrimination

2-Hexylamino-4-[(S)-1-(1-naphthyl)ethylamino]-6-L-valyl-L-valyl-L-valine isopropylester-1,3,5-triazine (1), a molecule characterized by two different chiral selectors, and 2-hexylamino-4,6-bis-L-valyl-L-valyl-L-valine isopropylester-1,3,5-triazine (2) and 2-ethoxy-4-hexylamino-6-[(S)-1-(1-naphthyl) ethylamino]-1,3,5-triazine (3), systems in which a single kind of chiral selector is present, have been prepared. The enantiodiscriminating ability in solution of the three compounds toward the N-3,5-dinitrobenzoyl derivatives of 1-phenylethylamine (4) or valine methylester (5) has been investigated by ¹H nuclear magnetic resonance (NMR) spectroscopy: 1 shows an improved versatility, relative to 2 and 3, as a chiral solvating agent for NMR spectroscopy. On the basis of the indications obtained, the usefulness of 2-chloro-4-[(S)-1-(1-naphthyl)ethylamino]-6-L-val-L-val-L-valine isopropylester-1,3,5-triazine (1a), a direct precursor of 1, as chiral solvating agent for the determination by NMR of the enantiomeric compositions of derivatives of amines, amino alcohols, amino acids, and carboxyl acids bearing a 3,5-dinitrophenyl moiety, has been demonstrated. Chirality 9:113–121, 1997. © 1997 Wiley-Liss, Inc.     

Proton and carbon-13 nuclear magnetic resonance studies on methyl (methyl d-galactosid)uronates and their per-O-acetyl derivatives

The ¹H- and ¹³C-n.m.r. spectra of the anomeric methyl (methyl d-galactosid)uronates, as well as the ¹H-n.m.r. spectra of their acetyl derivatives, were analyzed. The spectra of the unacetylated d-galactopyranosiduronates showed good correlation with those of the corresponding anomeric d-galactopyranuronic acids and their methyl esters, and with those of the anomeric methyl d-galactopyranosides. From the values of the chemical shifts and coupling constants, it was concluded that the anomeric methyl (methyl d-galactopyranosid)uronates and their corresponding peracetates are in the ⁴C₁(d) conformation. The chemical shifts in the ¹³C-n.m.r. spectra show good correlation with those of the methyl d-galactosides. The signals of the furanose derivatives appear at fields lower than those of the corresponding pyranose compounds.     

Relaxation kinetics of E. coli ribosomes.

Relaxation kinetics measurements on two types of ribosome preparations were parformed by the pressure-jump and temperature-Jump techniques, using light scattered at 90° as detector. For freshly prepared tibosomes isolated as 70S tight coupled from 26 000 RPM sucrose gradint sedimentation in 10 mM Mg²⁺, surprisingly large reaction amplitudes were found in 10 mM Mg²⁺ wilh both techniques, leading to an overall formation constant for 70S couples approximately three orders of magnitude smaller than that reported fot tight couples. For pelleted, two-tunes salt-washed ribosomes, amplitude titration versus Mg²⁺ in the pressure-jump apparatus showed an amplitude maximum near 10 mM Mg²⁺ with a relaxation time near 20 ms, and a second amplitude maximum near 2.5 mM Mg²⁺ with a relaxation time near 25 s. Both types of preparation on reanalysis on sucrose gradients at 5 mM Mg²⁺ showed approximately 15% of subunits, with a distinct zone in the 50S region. 70S light couples recovered from a sucrose density gradient separation at 5 mM Mg²⁺ on pelleted two-times salt-washed ribosomes behaved in the same way as the original sample in pressure-jump experiments at 10 mM Mg²⁺. These findings have been interpreted as follows (I) the processes observed at 10 mM Mg²⁺ are due entirety to the relatively small loose couple content of the samples, even in the case of material isolated as 70S tight couples, (2) the processes observed at 2.5 mM Mg²⁺ are due almost entirely to the preponderant tight couple population of the material, and (3) samples isolated as 70S tight couples from sucrose gradients at 5 mM Mg²⁺ spontaneously revert within hours into micro-heterogeneous material containing about 15% loose couples, for both types of ribosomes.     

Direct screening of chiral discrimination abilities of chiral hosts using mass spectrometry

A simultaneous estimation of the chiral discrimination abilities of several chiral hosts was demonstrated on the basis of one mass spectrum. The chiral host mixture, including H(1), H(2), H(3) ..., and H(m) (m: number of hosts) was prepared by etherification of several chiral alcohols with bistosylate of diethylene glycol. An equimolar mixture of a deuterium-labeled (S)- and unlabeled (R)-enantiomer of an amino acid isopropyl ester hydrochloride (G(S-dn) (+)Cl(-) and G(R) (+)Cl(-), n: number of deuterium atoms) was added to the chiral host mixture, and the FAB mass spectrum was measured to evaluate the chiral discrimination ability of each host in the mixture without isolation. The chiral discrimination ability of each host toward the guest is represented by the relative peak intensity of the diastereomeric complex ion pair, I(H(m) + G(R)((+)/I(H(m) + G(S-dn))(+) (=I(R)/I(S-dn) value). Several new hosts showed good chiral discrimination toward the guest.     

Separation of enantiomeric amines and acids using chiral ion-pair chromatography on porous graphitic carbon

The application of porous graphitic carbon as adsorbing phase for direct separation of enantiomeric acids and amines using chiral ion-pair chromatography is described. The enantiomeric amines were separated as diastereomeric ion pairs with N-benzyloxycarbonylglycyl-L -proline, N-benzyloxycarbonylglycylglycyl-L -proline, or captopril as the chiral counterion. High enantioselectivities were obtained for amines having a hydrogen bonding function in the vicinity of the asymmetrical carbon atom. Quinine was the chiral counterion used to separate the enantiomeric acids. The strongly UV-absorbing quinine improved detection of solutes having low UV-absorbing properties, e.g., (R,S)-2-chloropropionic acid, by “indirect detection.” Retention and stereoselectivity of enanticmeric acids were regulated by the quinine concentration and by the addition of carboxylic acids as well as polar modifiers, e.g., methanol and 2-propanol, to the mobile phase. © 1992 Wiley-Liss, Inc.     

Conformational spaces of the gastrointestinal antisecretory chiral drug omeprazole: stereochemistry and tautomerism

A study of the conformational spaces of the chiral proton pump inhibitor (PPI) drug omeprazole by semiempirical, ab-initio, and DFT methods is described. In addition to the chiral center at the sulfinyl sulfur atom, the chiral axis at the pyridine ring (due to the hindered rotation of the 4-methoxy substituents) was considered. The results were analyzed in terms of the 5-methoxy and 6-methoxy tautomers and the two pairs of enantiomers (R,P)/(S,M) and (R,M)/(S,P). Five torsion angles were systematically explored: the backbone rotations defined by D1 (N3-C2-S10-O11), D2 (C2-S10-C12-C13), and D3 (S10-C12-C13-N14) and two methoxy rotations defined by D4 (C6-C5-O8-C9) and D5 (C16-C17-O19-C20). Significant energy differences were revealed between the 5- and 6-methoxy tautomers, the extended and folded conformations, and the (S,M) and (S,P) diastereomers. The "extended M" conformation of the 6-methoxy tautomer of (S)-omeprazole was found to be the most stable conformer.     

Identification of methyl (methylO-acetyl-O-methylhexopyranosid)uronates by mass spectrometry

Fragmentation of methyl (methylO-acetyl-O-methyl-α-d-gluco- and -galactopyranosid, uronates has been studied at 70 and 12 eV. At 12 eV, the production of ions resulting from secondary and further processes is greatly diminished and the spectra are simpler and easier to interpret. The energy required for the elimination of acetic acid and ketene has been calculated from the ion-appearance potentials. The number and location of methyl groups in methyl (methylO-acetyl-O-methyl-hexopyranosid)uronates can be determined. The procedure is particularly suitable for g.l.c.-m.s. of the uronic acid portion of methanolysates of methylated biopolymers and other substances containing uronic acids. From the presence or absence of peaks for molecular ions in the 12-eV spectra,gluco andgalacto isomers which do not contain a methoxyl group at C-4 can be distinguished. The synthesis of methyl (methyl 2,3-di-O-methyl-α-d-galactopyranosid)uronate is also described.     

Covalent modification of subtilisin Bacillus lentus cysteine mutants with enantiomerically pure chiral auxiliaries causes remarkable changes in activity

Methanethiosulfonate reagents may be used to introduce virtually unlimited structural modifications in enzymes via reaction with the thiol group of cysteine. The covalent coupling of enantiomerically pure (R) and (S) chiral auxiliary methanethiosulfonate ligands to cysteine mutants of subtilisin Bacillus lentus induces spectacular changes in catalytic activity between diastereomeric enzymes. Amidase and esterase kinetic assays using a low substrate approximation were used to establish kcat/KM values for the chemically modified mutants, and up to 3-fold differences in activity were found between diastereomeric enzymes. Changing the length of the carbon chain linking the phenyl or benzyl oxazolidinone ligand to the mutant N62C by a methylene unit reverses which diastereomeric enzyme is more active. Similarly, changing from a phenyl to benzyl oxazolidinone ligand at S166C reverses which diastereomeric enzyme is more active. Chiral modifications at S166C and L217C give CMMs having both high esterase kcat/KM's and high esterase to amidase ratios with large differences between diastereomeric enzymes.     

Enthalpy change on mixing a couple of S- and R-enantiomers of some chiral compounds at 298.15 K

Enthalpy change on the mixing of R- and S-enantiomers of chiral liquid compounds such as dimethyl malate (1), methyl 3-hydroxylbutanoate (2), 2-butanol (3), ethyl 4-chloro-3-hydroxylbutanoate (4), 1,3,3-trimethylbicycle-[2.2.1]heptan-2-one (5), 3,7-dimethyl-6-octenal (6), and 8-bromo-2,6-dimethyl-2-octene (7) is measured over the entire range of mole fractions at 298.15 K, albeit very small values. The mixing of chiral liquids of R-1 + S-1, R-2 + S-2, R-3 + S-3, R-6 + S-6, and R-7 + S-7 produces enthalpic destabilization over the entire range of mole fractions, while that of R-4 + S-4 and R-5 + S-5 shows enthalpic stabilization over entire compositions. Enthalpy change on mixing at an equimolar concentration and the intermolecular interaction obtained by the molecular mechanics calculations show a linear correlation, except for a few compounds measured.     

First systematic chiral syntheses of two pairs of enantiomers with 3,5-dihydroxyheptenoic acid chain, associated with a potent synthetic statin NK-104

First systematic chiral syntheses of two pairs of enantiomers with 3,5-dihydroxyheptenoic acid chain, associated with a potent synthetic statin NK-104 are reported. A pair of syn diol isomers (NK-104 and its enantiomer) was obtained efficiently by diastereomeric resolution. The synthesis of a pair of anti diol isomers (3-epimer and 5-epimer) was accomplished effectively by the asymmetric aldol reaction followed by anti stereoselective reduction as key steps. Their purity determinations were effected by chiral HPLC analysis.