Enantioselectivity in the Rabbit Liver Microsomal Biotransformation of Styrene and Phenylpropenes

The rabbit liver microsomal P-450 catalyzed oxidation of styrene (1a) and isomeric phenylpropenes, trans-1-phenylpropene (1b), cis-1-phenylpropene (1c) and 3-phenylpropene (1d), was investigated and the enantioselectivity of the epoxidation of the olefinic double bond was determined by checking the enantiomeric excesses of the corresponding first formed epoxides (2). These enantiomeric excesses were always modest, ranging between 7% of (1S,2S)-(2b) and 22% of (1R,2R)-(2c). In the case of (1d) a nonenantioselective hydroxylation at the benzylic-allylic C(3) was also oberved. The ratio between this hydroxylation and olefin epoxidation of (Id) was 1:2.     

Stereochemical Difference in Secoisolariciresinol Formation between Cell-free Extracts from Petioles and from Ripening Seeds of Arctium lappa L.

Cell-free extracts from ripening seeds of Arctium lappa L. catalyzed the enantioselective formation of (-)-pinoresinol, (-)-lariciresinol and (-)-secoisolariciresinol from achiral coniferyl alcohol in the presence of NADPH and H₂O₂. The enantioselectivity of the lignan formation was opposite to that of the (+)-secoisolariciresinol formation catalyzed by cell-free extracts from petioles of the same plant species.     

Synthesis and pKa determination of new enantiopure dimethyl‐substituted acridino‐crown ethers containing a carboxyl group: Useful candidates for enantiomeric recognition studies

New enantiopure dimethyl‐substituted acridino‐18‐crown‐6 and acridino‐21‐crown‐7 ethers containing a carboxyl group at position 9 of the acridine ring [(S,S )‐ 8 , (S,S )‐ 9 , (R,R )‐ 10 ] were synthesized. The pK _a values of the new crown ethers [(S,S )‐ 8 , (S,S )‐ 9 , (R,R )‐ 10 ] and of an earlier reported macrocycle [(R,R )‐ 2 ] were determined by UV‐pH titrations. Crown ether (S,S )‐ 8 was attached to silica gel by covalent bonds and the enantiomeric separation ability of the newly prepared chiral stationary phase [(S,S )‐CSP‐ 12 ] was studied by high‐performance liquid chromatography (HPLC). Homochiral preference was observed and the best separation was achieved for the enantiomers of 1‐NEA. Ligands (S,S )‐ 9 and (R,R )‐ 10 are precursors of enantioselective sensor and selector molecules for the enantiomers of protonated primary amines, amino acids, and their derivatives.     

Asymmetric bioreduction of p-haloacetophenones by Mucor

Abstract

A number of p-haloacetophenones were asymmetrically bioreduced to their corresponding (S)-alcohols by Mucor sp. CG10 with good conversion and excellent enantioselectivity. The results showed that the electronic effects of the halogen substituent (X-group) affected the conversion of the substrates and the enantioselectivity of the reaction. The trend observed was as the X-group at the para-position became more electron donating from F, to Cl, Br and I, the conversion of substrates decreased, while the enantioselectivity increased.     

Asymmetric hydrolysis of dimethyl 3-phenylglutarate catalyzed by Lecitase Ultra: Effect of the immobilization protocol on its catalytic properties

The asymmetric hydrolysis of dimethyl 3-phenylglutarate (1) by different immobilized preparations of a phospholipase A₁ (Lecitase Ultra (LECI)) at pH 7 and 25 °C has been studied. Agarose beads coated with octyl, cyanogen bromide (CNBr), polyethylenimine (PEI) or glyoxyl groups were used as supports for the immobilization of LECI. The different derivatives behaved very differently in terms of activity, discrimination between 1 and methyl 3-phenylglutarate (2) resulting from the hydrolysis of 1, enantioselectivity (in the hydrolysis of 1 to produce R or S-2) and enantiospecificity in the hydrolysis of R-2 and S-2. Using 1 mM of 1, CNBr-LECI showed the highest activity (13 × 10⁻³ μmol/min mg protein) while octyl-LECI was about 20 times less active. All the enzyme preparations mainly produced (S)-2, but with different enantioselectivity. CNBr-Lecitase was the most enantioselective, producing the S-2 10 fold more rapidly than the R-2, while octyl-Lecitase gave only half of that difference.LECI adsorbed on octyl-agarose allowed to get a yield up to 99% of S-2 (ee was 66%). The reaction stopped in the monoester and no isomer of this compound was further hydrolyzed by the enzyme. However, when the reaction was catalyzed by the other immobilized LECI preparations, the enzyme was able to hydrolyze mainly the minority isomer, permitting to improve the ee of the remaining S-2. The best results were obtained using CNBr-LECI, which gave (S)-methyl-3-phenylglutarate with a yield of 80% and an ee exceeding 99%.     

Stereodesign of chiral para-substituted calix[4]arenes

According to the hypothesis that the chirality of molecule hosts is a cause of their enantioselectivity, the chirality of para-substituted calix[4]arenes was analyzed quantitatively. The relationship between types of para-substituents and the dissymmetry of the 2D (two-dimensional) entrance into the cavity and the whole 3D (three-dimensional) cavity of calix[4]arenes was studied by means of the enantiomer dissimilarity factor (EDF) method for quantitative evaluation of molecular chirality. The design of the most chiral, and probably enantioselective, para-substituted calix[4]arenes was planned such that all four substituents should be different and the two largest should be near each other (adjacent). It was, on the other hand, shown that the 2D chiral entrance determines chirality of the whole 3D structures of these molecules. This phenomenon is interpreted as an example of the chirality transition from 2D into 3D space.Electronic Supplementary Material available.     

Resolution of 4-(1-hydroxy-2,2,2-trifluoroethyl)phenol and its derivatives by lipase-catalyzed enantioselective alcoholysis

Lipase LIP from Pseudomonas aeruginosa,one of nine commercially available hydrolytic enzymes, catalyzed the enantioselective alcoholysis of racemic 4-(1-acetoxy-2,2,2-trifluoroethyl)phenyl acetate with n-butanol, affording (S)-4-(1-hydroxy-2,2,2-trifluoroethyl)phenol at >99% e.e. (E = >100). Moreover, it also showed high enantioselectivity (E = >100) for the alcoholysis of the racemic o-substituted isomer, 2-(1-acetoxy-2,2,2-trifluoroethyl)phenyl acetate.     

Stereochemical Structure–Activity Relationship of N-(2,3-Epoxypropyl)-N-(α-methylbenzyl)benzenesulfonamide Derivatives

The absolute configurations of two asymmetric centers in four stereoisomers of N-(2,3-epoxypropyl)-N-(α-methylbenzyl)benzenesulfonamide were determined and their biological activities were tested. Consequently, N-[(R)-2,3-epoxypropyl]-N-[(R)-α-methylbenzyl]benzenesulfonamide was found to be the most active isomer and the stereochemistry of the benzyl position was found to be more important than that of C₂ in the epoxypropyl group for biological activity.     

A magnetically separable biocatalyst for resolution of racemic naproxen methyl ester

Candida rugosa lipase (CRL) was encapsulated via the sol–gel method, using 5, 11, 17, 23-tetra-tert-butyl-25,27-bis(2-aminopyridine)carbonylmethoxy-26, 28-dihydroxy-calix[4]arene-grafted magnetic Fe₃O₄ nanoparticles (Calix-M-E). The catalytic activity of encapsulated lipase (Calix-M-E) was tested both in the hydrolysis of p-nitrophenyl palmitate (p-NPP) and the enantioselective hydrolysis of racemic naproxen methyl ester. The present study demonstrated that the calixarene-based compound has the potential to enhance both reaction rate and enantioselectivity of the lipase-catalyzed hydrolysis of racemic naproxen methyl ester. The encapsulated lipase (Calix-M-E) had great catalytic activity and enantioselectivity (E > 400), as well as remarkable reusability as compared to the encapsulated lipase without supports (E = 137) for S-Naproxen.     

Hydrolytic kinetic resolution of the enantiomers of the structural isomers trans-1-phenylpropene oxide and (2,3-epoxypropyl)benzene by yeast epoxide hydrolase

Kinetic resolution of the enantiomers of trans -1-phenylpropene oxide and (2,3-epoxypropyl)benzene was achieved by yeasts from the genus Rhodotorula. The resolution of trans -1-phenylpropene oxide by Rhodotorula glutinis UOFS Y-0123 yielded (1R,2R)-epoxide (ee >98%, yield 30%) and (1R,2S)-diol (ee 95%, yield 40%). The highest enantio- and regioselectivity toward (2,3-epoxypropyl)benzene resided in Rhodotorula sp. UOFS Y-0448 (E = 6.16), yielding (S)-epoxide (ee 64%, yield 33%) and (R)-diol (ee 67%, yield 28%). This confirms the superiority of yeasts from the Basidiomycetes genera in the enantioselective hydrolysis of epoxides from different structural classes.     

Enantioselectivity in the Rabbit Liver Microsomal Biotransformation of Styrene and Phenylpropenes

The rabbit liver microsomal P-450 catalyzed oxidation of styrene (1a) and isomeric phenylpropenes, trans-1-phenylpropene (1b), cis-1-phenylpropene (1c) and 3-phenylpropene (1d), was investigated and the enantioselectivity of the epoxidation of the olefinic double bond was determined by checking the enantiomeric excesses of the corresponding first formed epoxides (2). These enantiomeric excesses were always modest, ranging between 7% of (1S,2S)-(2b) and 22% of (1R,2R)-(2c). In the case of (1d) a nonenantioselective hydroxylation at the benzylic-allylic C(3) was also oberved. The ratio between this hydroxylation and olefin epoxidation of (Id) was 1:2.     

Synthesis of mixed N,N′-(Ar,Ar′-diaryl)iminoisoindolines for applications in palladacycle formations

The synthesis of N,N′-(Ar,Ar′-diaryl)iminoisoindolines containing different aryl groups bound to the two nitrogen atoms is described. The iminoisoindolines were obtained by a three component, one-pot reaction of phthalaldehyde with 1 equivalent p-NO₂-aniline and 1 equivalent p-R-aniline, where R = H, Me, MeO or ⁱPr, resulting in formation of non-symmetrically substituted (mixed) iminoisoindolines, 1-p-nitrophenylimino-2-p-R-phenylisoindoline (R = H (1), Me (2), MeO (3), and ⁱPr (4)), as analytically pure precipitates requiring no further purification. Only one isomer precipitates from solution wherein the nitro group resides exclusively at the imine position while the more electron donating substituent ends up on the isoindoline ring position. Further reaction with Pd(OAc)₂ in dichloromethane at room temperature results in formation of six-membered [C,N] dinuclear cyclopalladated complexes with the general formula [(Ar,Ar′-diaryliminoisoindoline)Pd{μ-OAc}]₂.     

Directed evolution of metagenome-derived epoxide hydrolase for improved enantioselectivity and enantioconvergence

We performed a directed evolution study with a metagenome-derived epoxide hydrolase (EH), termed Kau2. Homology models of Kau2 were built; we selected one of them and used it as a guide for saturation mutagenesis experiments targeted at specific residues within the large substrate binding pocket. During the molecular evolution process, we found several enzyme variants with higher enantioselectivity or enhanced enantioconvergence toward para-Chlorostyrene oxide. Improved enantioselectivities by up to a factor of 5, reaching an E-value of up to 130 with the R-enantiomer as the residual epoxide, were achieved by replacing amino acid pairs at the positions 110 and 113, or 290 and 291, which are positions located in the vicinity of two presumed binding sites for the epoxide enantiomers. The (R)-para-Chlorophenylethane-1,2-diol product exhibited a high enantiomeric excess (ee) of 97% at 50% conversion of the racemic epoxide for the most enantioselective variant. Further, five amino acid substitutions were sufficient to substantially increase the degree of enantioconvergence and to lower the E-value to 17 for the final evolved EH variant, enabling the production of the R-diol with an ee-value of 93% at 28 °C in a complete conversion of the racemic epoxide. Higher ee_p-values of up to 97% were determined in enantioconvergent reactions using lower temperatures. The EH activities of whole cells were found to be within the range of 74–125% of the wild-type activity for all investigated variants. We show in this report that the metagenome-derived Kau2 EH is amenable to the redesign of its enantioselectivity and regioselectivity properties by directed evolution using a homology model as a guide. The generated enzyme variants should be useful for the production of the chiral building blocks (R)-para-Chlorostyrene oxide and (R)-para-Chlorophenylethane-1,2-diol.     

Room temperature and highly enantioselective additions of alkyltitanium reagents to aldehydes catalyzed by a titanium catalyst of (R)‐h8‐binol

Three alkyltitanium reagents of RTi(O‐i‐Pr)₃ (R = Cy ( 1a ), i‐Bu ( 1b ), and n‐Bu ( 1c )) were prepared in good yields. The high‐resolution mass spectroscopy showed that 1b and 1 c in the gas phase are monomeric species. However, the solid state of 1a revealed a dimeric structure. Asymmetric additions of 1a , 1b , 1c to aldehydes catalyzed by a titanium catalyst of (R)‐H₈‐BINOL were studied at room temperature. The reactions produced desired secondary alcohols in good yields with good to excellent enantioselectivities of up to 94% ee. Reactivity and enantioselectivity differences, in terms of steric bulkiness of the R nucleophiles, are herein described. The addition reactions of secondary c‐hexyl to aldehydes were slower than the reactions of primary i‐butyl or n‐butyl nucleophiles. For the primary alkyls, lower enantioselectivities were obtained for products from addition reactions of the linear n‐butyl as compared with the enantioselectivities of products from the addition reactions of the branched i‐butyl group. The same stereochemistry of RTi(O‐i‐Pr)₃ addition reactions as the addition reactions of organozinc, organoaluminum, Grignard, or organolithium reagents directly supports the argument of that titanium‐catalyzed addition reactions of aldehydes involve an addition of an organotitanium nucleophile. Chirality, 2011. © 2011 Wiley‐Liss, Inc.     

Palladium(II) complexes of a bis-2-aminobiphenyl N-heterocyclic carbene: Synthesis, structural studies and catalytic activity

A new imidazolinium [(SIBiphen)H](BF₄) was synthesized in three steps from 2-aminobiphenyl. The reaction of the salt with Pd(OAc)₂, NaI and t-BuOK gave a dimeric Pd(II) complex [(SIBiphen)PdI₂]₂, which was analyzed by an X-ray diffraction study. The reaction of [Pd(allyl)Cl]₂, the imidazolinium salt and t-BuOK in THF at −78 °C gave the monomeric Pd complex, in which the N-heterocyclic carbene was bound to the metal centre, as confirmed by a single-crystal X-ray diffraction study. A preliminary catalytic study showed that these new systems were moderately active in the Suzuki-Miyaura coupling of aryl halides.     

Catalysis of regioselective reduction of NAD+ by ruthenium(II) arene complexes under biologically relevant conditions

Ruthenium(II) arene anticancer complexes [(η ⁶-arene)Ru(en)Cl]PF₆ (arene is hexamethylbenzene, p-cymene, indan; en is ethylenediamine) can catalyse regioselective reduction of NAD⁺ by formate in water to form 1,4-NADH, at pD 7.2, 37 °C, and in the presence of air. The catalytic activity is markedly dependent on the arene, with the hexamethylbenzene (hmb) complex showing the highest activity. For [(η ⁶-hmb)Ru(en)Cl]PF₆, the rate of reaction is independent of NAD⁺ concentration and shows saturation kinetics with respect to formate concentration. A K _m value of 58 mM and a turnover frequency at saturation of 1.46 h⁻¹ were observed. Removal of chloride and performing the reaction under argon led to higher reaction rates. Lung cancer cells (A549) were found to be remarkably tolerant to formate even at millimolar concentrations. The possibility of using ruthenium arene complexes coadministered with formate as catalytic drugs is discussed.Electronic Supplementary Material Supplementary material is available for this article at and is accessible for authorized users.     

Hydroformylation of alkenes catalyzed by new dinuclear aryloxide- and carboxylate-bridged rhodium complexes

The new dinuclear aryloxide- and carboxylate-bridged rhodium complexes bis[μ-(2-methylphenolato)]bis[(η²:η²-cycloocta-1,5-diene)rhodium], di(μ-docosanoato)bis[(η²:η²-norborna-2,5-diene)rhodium] and bis[(μ-(adamant-1-yl)carboxylato]bis[(η²:η²-norborna-2,5-diene)rhodium] have been prepared. The complexes were tested as catalysts in the hydroformylation of styrene with a total pressure of CO/H₂ (1:1) of 1000 psi, at 25 and 60 °C, and displayed a regioselectivity towards the branched aldehyde of up to 97%. The same complexes as catalysts in the hydroformylation of 1-octene displayed a regioselectivity for the linear aldehyde of up to 55%.     

The direct determination of the enantiomers of ketorolac and parahydroxyketorolac in plasma and urine using enantioselective liquid chromatography on a human serum albumin-based chiral stationary phase

An enantioselective assay has been developed for the determination of the enantiomers of ketorolac and its metabolite p-hydroxyketorolac in plasma and urine. The analytical method utilizes a coupled achiral–chiral HPLC system where the initial separation of ketorolac from p-hydroxyketorolac and matrix interferences was achieved on a C₁₈-stationary phase and the enantioselective separations of the two target solutes were accomplished on a human serum albumin-based chiral stationary phase. The two columns were attached in sequence and the assay was carried out without the necessity of column-switching techniques. The method has been validated for use in pharmacokinetic and metabolic studies and represents the initial report of the determination of ketorolac and p-hydroxyketorolac enantiomers in urine. The results of the study indicate that after the administration of racemic ketorolac there was an enantioselective distribution of ketorolac enantiomers in plasma [(R)-ketorolac: (S)-ketorolac = 3.89 ± 0.93 (n = 6) and urine (R)-ketorolac: (S)-ketorolac = 1.26 ± 0.09 (n = 7)]. The mean ratio of the p-hydroxyketorolac enantiomers was 1.77 ± 0.46 (n = 7). Both ketorolac and p-hydroxyketorolac are glucuronized in the acyl carboxyl moiety and the results of this study indicate that this process is not enantiospecific. © 1994 Wiley-Liss, Inc.     

Asymmetric hydrolysis of dimethyl 3-phenylglutarate catalyzed by Lecitase Ultra®: Effect of the immobilization protocol on its catalytic properties

The asymmetric hydrolysis of dimethyl 3-phenylglutarate (1) by different immobilized preparations of a phospholipase A₁ (Lecitase Ultra (LECI)) at pH 7 and 25 °C has been studied. Agarose beads coated with octyl, cyanogen bromide (CNBr), polyethylenimine (PEI) or glyoxyl groups were used as supports for the immobilization of LECI. The different derivatives behaved very differently in terms of activity, discrimination between 1 and methyl 3-phenylglutarate (2) resulting from the hydrolysis of 1, enantioselectivity (in the hydrolysis of 1 to produce R or S-2) and enantiospecificity in the hydrolysis of R-2 and S-2. Using 1 mM of 1, CNBr-LECI showed the highest activity (13 × 10⁻³ μmol/min mg protein) while octyl-LECI was about 20 times less active. All the enzyme preparations mainly produced (S)-2, but with different enantioselectivity. CNBr-Lecitase was the most enantioselective, producing the S-2 10 fold more rapidly than the R-2, while octyl-Lecitase gave only half of that difference.LECI adsorbed on octyl-agarose allowed to get a yield up to 99% of S-2 (ee was 66%). The reaction stopped in the monoester and no isomer of this compound was further hydrolyzed by the enzyme. However, when the reaction was catalyzed by the other immobilized LECI preparations, the enzyme was able to hydrolyze mainly the minority isomer, permitting to improve the ee of the remaining S-2. The best results were obtained using CNBr-LECI, which gave (S)-methyl-3-phenylglutarate with a yield of 80% and an ee exceeding 99%.     

Ruthenium(II) arene complexes containing four- and five-membered monoanionic O,O-chelate rings

Optimization of the design of half-sandwich organometallic Ru^II arene complexes as anticancer agents depends on control of ligand exchange reactions. We have studied the aqueous chemistry of complexes containing O,O-chelate rings. The presence of the four-membered O,O-chelate ring from acetate (AcO) in [η⁶-p-cymene)Ru(AcO)Cl] was confirmed by X-ray crystallography, but in solution the acetate ligand was labile and the hydroxo-bridged dimer [((η⁶-p-cymene)Ru)₂(μ-OH)₃]⁺ readily formed. The dimer was relatively unreactive towards 9-ethyl guanine. The tropolonato (trop) complex [(η⁶-p-cymene)Ru(trop)Cl] was stable in aqueous media and the X-ray crystal structure of the aqua adduct [(η⁶-p-cymene)Ru(trop)(H₂O)]CF₃SO₃, containing a five-membered O,O-chelate ring from trop, was determined. [(η⁶-p-cymene)Ru(trop)Cl] reacted with guanosine to form N7 adducts and with adenosine to form both N7 and N1 adducts. Competitive reactions with guanosine and adenosine gave rise to guanosine:adenosine adducts in a ca. 1.3:1 mol ratio.