Fundamental investigations of enantioselective heterogeneous catalysis

Enantioselective catalysis is an increasingly important method of providing enantiomeric compounds for the pharmaceutical and agrochemical industries. To date, heterogeneous catalysts have failed to match the industrial impact achieved by homogeneous systems. One successful approach to the creation of heterogeneous enantioselective catalysts has involved the modification of conventional metal particle catalysts by the adsorption of chiral molecules. This article examines the contribution of effects such as chiral recognition and amplification to these types of system and how insight provided by surface science model studies may be exploited in the design of more effective catalysts.

The enantioselective population pharmacokinetics of intravenous ketorolac in children using a stereoselective assay suitable for microanalysis

Objective: To describe the effect of age and body size on enantiomer selective pharmacokinetic (PK) of intravenous ketorolac in children using a microanalytical assay. Methods: Blood samples were obtained at 0, 15 and 30 min and at 1, 2, 4, 6, 8 and 12 h after a weight-dependent dose of ketorolac. Enantiomer concentration was measured using a liquid chromatography tandem mass spectrometry method. Non-linear mixed-effect modelling was used to assess PK parameters. Key findings: Data from 11 children (1.7–15.6 years, weight 10.7–67.4 kg) were best described by a two-compartment model for R(+), S(−) and racemic ketorolac. Only weight (WT) significantly improved the goodness of fit. The final population models were CL = 1.5 × (WT/46)0.75, V1 = 8.2 × (WT/46), Q = 3.4 × (WT/46)0.75, V2 = 7.9 × (WT/46), CL = 2.98 × (WT/46), V1 = 13.2 × (WT/46), Q = 2.8 × (WT/46)0.75, V2 = 51.5 × (WT/46), and CL = 1.1 × (WT/46)0.75, V1 = 4.9 × (WT/46), Q = 1.7 × (WT/46)0.75 and V2 = 6.3 × (WT/46)for R(+), S(−) and racemic ketorolac. Conclusions: Only body weight influenced the PK parameters for R(+) and S(−) ketorolac. Using allometric size scaling significantly affected the clearances (CL, Q) and volumes of distribution (V1, V2).

Chloroperoxidase catalyzed enantioselective epoxidation of selected olefins and regiospecific degradation of dimethylsulfoniopropionate

Chloroperoxidase (CPO), secreted by marine fungus Caldariomyces fumago, is the most versatile catalyst among known heme enzymes. Chloroperoxidase can catalyze epoxidation reactions with high enantioselectivity and high yield, which makes CPO an attractive candidate for both industrial and medicinal chiral synthesis. Toward this end, we have constructed two CPO mutants, F103A and N74V. Chiral HPLC was used to evaluate the enantioselectivity and yield of CPO and the mutants toward the epoxidation of styrene and its derivatives. Both of the mutants show dramatically changed epoxidation profiles compared to the parent protein. This information provided fresh insight into the mechanism through which CPO achieves its enantioselectivity. Furthermore, effort was made to understand the biological function of CPO through characterization of CPO catalyzed oxidation of dimethylsulfoniopropionate (DMSP), a secondary metabolite of many marine algal species that plays a pivotal role in marine ecology and global climate.^

Mechanisms of chloroperoxidase-catalyzed enantioselective reactions as probed by site-directed mutagenesis and isotopic labeling

Chloroperoxidase (CPO) is a heme-containing glycoprotein secreted by the marine fungus Caldariomyces fumago. Chloroperoxidase contains one ferriprotoporphyrin IX prosthetic group per molecule and catalyzes a variety of reactions, such as halogenation, peroxidation and epoxidation. The versatile catalytic activities of CPO coupled with the increasing demands for chiral synthesis have attracted an escalating interest in understanding the mechanistic and structural properties of this enzyme. In order to better understand the mechanisms of CPO-catalyzed enantioselective reactions and to fine-tune the catalytic properties of chloroperoxidase, asparagine 74 (N74) located in the narrow substrate access channel of CPO was replaced by a bulky, nonpolar valine and a polar glutamine using site-directed mutagenesis. The CPO N74 mutants displayed significantly enhanced activity toward nonpolar substrates compared to wild-type CPO as a result of changes in space and polarity of the heme distal environment. More interestingly, N74 mutants showed dramatically decreased chlorination and catalase activity but significantly enhanced epoxidation activity as a consequence of improved kinetic perfection introduced by the mutation as reflected by the favorable changes in k cat and kcat/KM of these reactions. It is also noted that the N74V mutant is capable of decomposing cyanide, the most notorious poison for many hemoproteins, as judged by the unique binding behavior of N74V with potassium cyanide. Histidine 105 (H105) was replaced by a nonpolar amino acid alanine using site-directed mutagenesis. The CPO H105 mutant (H105A) displayed dramatically decreased chlorination and catalase activity possibly because of the decreased polarity in the heme distal environment and loss of the hydrogen bonds between histidine 105 and glutamic acid 183. However, significantly increased enantioselectivity was observed for the epoxidation of bulky styrene derivatives. Furthermore, my study provides strong evidence for the proposed histidine/cysteine ligand switch in chloroperoxidase, providing experimental support for the structure of the 420-nm absorption maximum for a number of carbon monoxide complexes of heme-thiolate proteins. For the NMR study, [dCPO(heme)] was produced using 90% deuterated growth medium with excess heme precursors and [dCPO(Phe)] was grown in the same highly deuterated medium that had been supplemented with excess natural phenylalanine. To make complete heme proton assignments, NMR spectroscopy has been performed for high-resolution structural characterization of [dCPO(heme)] and [dCPO(Phe)] to achieve unambiguous and complete heme proton assignments, which also allows important amino acids close to the heme active center to be determined.

Investigation of additive effects in enantioselective copper-catalysed C-H insertion and aromatic addition reactions of α-diazocarbonyl compounds

Significant enhancements in enantioselectivities and reaction efficiencies in asymmetric copper-catalysed C-H insertion and aromatic addition reactions of α-diazocarbonyl compounds in the presence of various group I salts are reported. For the first time in carbenoid chemistry, evidence for the critical role of the metal cation is described.

Catalyst and substrate effects in enantioselective C–H insertion reactions of α-diazosulfones

This thesis describes a systematic investigation of the mechanistic and synthetic aspects of intramolecular reactions of a series of α-diazo-β-oxo sulfone derivatives using copper and, to a lesser extent, rhodium catalysts. The key reaction pathways explored were C–H insertion and cyclopropanation, with hydride transfer competing in certain instances. Significantly, up to 98% ee has been achieved in the C–H insertion processes using copper-NaBARF-bisoxazoline catalysts, with the presence of the additive NaBARF critical to the efficiency of the transformations. This novel synthetic methodology provides access to a diverse range of enantioenriched heterocyclic compounds including thiopyrans, sulfolanes, β- and γ-lactams, in addition to carbocycles such as fused cyclopropanes. The synthesis of the α-diazosulfones required for subsequent investigations is initially described. Of the twenty seven diazo sulfones described, nineteen are novel and are fully characterised in this work. The discussion is subsequently focused on a study of the copper and rhodium catalysed reactions of the α-diazosulfones with Chapter Four concentrated on highly enantioselective C–H insertion to form thiopyrans and sufolanes, Chapter Five focused on C–H insertion to form fused sulfolanes, Chapter Six focused on C–H insertion in sulfonyl α-diazoamides where both lactam formation and / or thiopyran / sulfolane formation can result from competing C–H insertion pathways, while Chapter Seven focuses on cyclopropanation to yield fused cyclopropane derviatives. One of the key outcomes of this work is an insight into the steric and / or electronic factors on both the substrate and the catalyst which control regio-, diastereo- and enantioselectivity patterns in these synthetically powerful transformations. Full experimental details for the synthesis and spectral characterisation of the compounds are included at the end of each Chapter, with details of chiral stationary phase HPLC analysis and assignment of absolute stereochemistry included in the appendix.

Enantioselective chemoenzymatic synthesis of 3-hydroxytetrahydrofurans

The research described in this thesis is concerned with the synthesis and stereoselective transformations of 4,5-dihydro-3(2H)-furanones and their 3-hydroxy derivatives. In Chapter 1, a review of synthetic routes to 3-hydroxytetrahydrofurans is presented. This incorporates the wide range of applications for these types of compounds. Preparative routes to and stereoselective transformations of the furanones investigated in this study are discussed in Chapter 2. The bulk of the work centers on stereoselective carbonyl group reductions to generate the 3-hydroxytetrahydrofuran derivatives in racemic form followed by kinetic resolution via lipase mediated esterification, resulting in enantioenriched 3-acetoxy and 3-hydroxytetrahydrofuran derivatives. In many cases, these processes proceed in a highly enantioselective manner. The influence of the lipase species and concentration of enzyme employed on the yield and stereochemical outcome of the reactions is examined in detail. Access to the complementary series of furanone and hydroxytetrahydrofuran derivatives by oxidation or reduction of the enantioenriched compounds was achieved through conventional synthetic methods. Chapter 2 also contains details of a novel synthetic route to a range of 2,3,5-trisubstituted furans from α-hydroxyenones and 4,5-dihydro-3(2H)-furanones. The mechanistic rationale for these transformations and the migratory aptitude of alkyl groups towards the formation of these furans is discussed in detail. Finally, Chapter 2 outlines the synthesis of a series of diarylcyclopentenones that were synthesised as part of our investigations. Chapter 3 contains a description of the synthetic procedures and biotransformations carried out together with key analytical and spectroscopic properties of the compounds studied and where appropriate, their analysis using chiral HPLC analysis.

Catalytic enantioselective electrocyclic cascades

A catalytic enantioselective electrocyclic cascade leads to the construction of topologically complex systems comprising multiple rings with up to three stereocentres. This phase-transfer catalysed process offers a new strategy for the rapid and enantioselective generation of complex products bearing all-carbon quaternary stereogenic centres. © 2012 The Royal Society of Chemistry.

Rhodium-catalysed enantioselective C-H functionalization in asymmetric synthesis

Development of an organocatalytic method for the enantioselective synthesis of rivastigmine

The focus of this thesis was the study of a recently developed class of picolinamide cinchona alkaloid derivatives for the synthesis of Rivastigmine, a biologically active compound used for the treatment of Alzheimer’s disease. Six 9-picolinamide-cinchona alkaloid derivatives were successfully synthesized through simple and effective methods. These catalysts were then applied in the enantioselective reduction of O-protected ketimines (intermediates of Rivastigmine). The hydrosilylation of the N-phenyl ketimines afforded good results with excellent yields and high enantioselectivities, while much lower values, in terms of both enantioselectivity and yield, were obtained in the reduction of N-benzyl ketimines. Preliminary studies on the immobilization of these organocatalysts to different solid supports were conducted, with the purpose of applying them in continuous flow systems, which to date has never been reported; RESUMO: No âmbito deste trabalho, foi estudada a síntese de um composto biologicamente ativo usado para o tratamento da doença de Alzheimer, Rivastigmina, usando uma classe de picolinamidas derivadas de alcaloides de cinchona recentemente desenvolvida. Seis 9-picolinamida derivados de alcaloides de cinchona foram preparados com êxito através de metodologias simples e eficazes. Os organocatalisadores foram posteriormente aplicados na redução enantiosseletiva de cetiminas O-protegidas (intermediários de Rivastigmina). Foram obtidos bons resultados na hidrossililação de N-fenilo cetiminas, com rendimentos excelentes e elevadas enantiosseletividades, enquanto a redução de N-benzilo cetiminas proporcionou valores muito mais baixos, tanto em termos de rendimento como de enantiosseletividade. Com o objetivo de serem aplicados em sistemas de fluxo contínuo, realizaram-se estudos preliminares sobre a imobilização destes organocatalisadores em diferentes suportes sólidos, a qual, ate à data, ainda não foi descrita na literatura.

Mechanisms of Chloroperoxidase-catalyzed Enantioselective Reactions as Probed by Site-directed Mutagenesis and Isotopic Labeling

Chloroperoxidase (CPO) is a heme-containing glycoprotein secreted by the marine fungus Caldariomyces fumago. Chloroperoxidase contains one ferriprotoporphyrin IX prosthetic group per molecule and catalyzes a variety of reactions, such as halogenation, peroxidation and epoxidation. The versatile catalytic activities of CPO coupled with the increasing demands for chiral synthesis have attracted an escalating interest in understanding the mechanistic and structural properties of this enzyme. In order to better understand the mechanisms of CPO-catalyzed enantioselective reactions and to fine-tune the catalytic properties of chloroperoxidase, asparagine 74 (N74) located in the narrow substrate access channel of CPO was replaced by a bulky, nonpolar valine and a polar glutamine using site-directed mutagenesis. The CPO N74 mutants displayed significantly enhanced activity toward nonpolar substrates compared to wild-type CPO as a result of changes in space and polarity of the heme distal environment. More interestingly, N74 mutants showed dramatically decreased chlorination and catalase activity but significantly enhanced epoxidation activity as a consequence of improved kinetic perfection introduced by the mutation as reflected by the favorable changes in kcat and kcat/KM of these reactions. It is also noted that the N74V mutant is capable of decomposing cyanide, the most notorious poison for many hemoproteins, as judged by the unique binding behavior of N74V with potassium cyanide. Histidine 105 (H105) was replaced by a nonpolar amino acid alanine using site-directed mutagenesis. The CPO H105 mutant (H105A) displayed dramatically decreased chlorination and catalase activity possibly because of the decreased polarity in the heme distal environment and loss of the hydrogen bonds between histidine 105 and glutamic acid 183. However, significantly increased enantioselectivity was observed for the epoxidation of bulky styrene derivatives. Furthermore, my study provides strong evidence for the proposed histidine/cysteine ligand switch in chloroperoxidase, providing experimental support for the structure of the 420-nm absorption maximum for a number of carbon monoxide complexes of heme-thiolate proteins. For the NMR study, [dCPO(heme)] was produced using 90% deuterated growth medium with excess heme precursors and [dCPO(Phe)] was grown in the same highly deuterated medium that had been supplemented with excess natural phenylalanine. To make complete heme proton assignments, NMR spectroscopy has been performed for high-resolution structural characterization of [dCPO(heme)] and [dCPO(Phe)] to achieve unambiguous and complete heme proton assignments, which also allows important amino acids close to the heme active center to be determined.

Preparative deracemization of unnatural amino acids

Unnatural amino acids are a growing class of intermediates required for pharmaceuticals, agrochemicals and other industrial products. However, no single method has proven sufficiently versatile to prepare these compounds broadly at scale. To address this need, we have developed a general chemoenzymatic process to prepare enantiomerically pure L- and D-amino acids in high yield by deracemization of racemic starting materials. This method involves the concerted action of an enantioselective oxidase biocatalyst and a non-selective chemical reducing agent to effect the stereoinversion of one enantiomer and can result in an enantiomeric excess of >99% from the starting racemate, and product yields of over 90%. This approach compares very favourably with resolution processes, which have a maximum single-pass yield of 50%. We have developed efficient methods to adapt the process towards new target compounds and to optimize key factors that influence process efficiency and offer competitive economics at scale.

Chemo-enzymatic synthesis of unnatural amino acids

A general chemo-enzymatic process has been developed to prepare enantiomerically pure L- and D-amino acids in high yield by deracemisation of racemic starting materials. The method has been developed from initial academic studies to be a robust, scalable industrial process. Unnatural amino acids, in high optical purity, are a rapidly growing class of intermediates required for pharmaceuticals, agrochemicals and other fine chemical applications. However, no single method has proven sufficiently adaptable to prepare these compounds generally at large scale. Our approach uses an enantioselective oxidase biocatalyst and a non-selective chemical reducing agent to effect the stereoinversion of one enantiomer and can result in an enantiomeric excess of > 99 % from a starting racemate, and product yields over 90 %. The current approach compares very favourably to resolution methods which have a maximum single pass yield of 50 %. Efficient methods have been developed to adapt the biocatalyst used in this process towards new target compounds and to optimise key factors which improve the process efficiency and offer competitive economics at scale.