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Biomedical subjects

John M Woodley

Publications and source records attributed to John M Woodley.

9 recordsLinked to original sources
PubMed2006

Biocatalysis for pharmaceutical intermediates: the future is now.

Biocatalysis is continuing to gain momentum and is now becoming a key component in the toolbox of the process chemist, with a place alongside chemocatalysis and chromatographic separations. The pharmaceutical industry demands a speed of development that must be on a parallel with conventional chemistry and high optical purity for complex compounds with multiple chiral centres. This review describes how these demands are being addressed to make biocatalysis successful, particularly by the use of micro-scale technology for high-speed catalyst screening and process development alongside discipline integration of biology and engineering with chemistry. Developments in recombinant technology will further expand the repertoire of biocatalysis in the coming years to new chemistries and enable catalyst design to fit the process. Further development of biocatalysis for green chemistry and high productivity processes can also be expected.

Biotransformation↗
PubMed2006

On oxygen limitation in a whole cell biocatalytic Baeyer-Villiger oxidation process.

In this article, a recombinant cyclohexanone monooxygenase (CHMO), overexpressed in Escherichia coli has been used to study the oxidation of bicyclo[3.2.0]hept-2-en-6-one to its two corresponding lactones at very high enantiomeric excess. The reaction is a useful model for the study of biocatalytic oxidations to create optically pure molecules. The major limitations to a highly productive biocatalytic oxidation in this case are oxygen supply, product inhibition, and biocatalyst stability. In this article, we investigate the effects of whole cell biocatalyst concentration on the rate of reaction at a range of scales from shake flasks to 75 L bioreactors. At low cell concentrations (<2 g(dcw)/L) the maximum specific rate (0.65 g/g(dcw).h) is observed. However, at higher cell concentrations (> 2 g(dcw)/L), the reaction becomes oxygen limited and both the specific rate and absolute rate decrease with further increases in cell concentration. The role of oxygen limitation in reducing the rate of reaction with scale was investigated by increasing the maximum oxygen transfer rate in the reactor at a high cell concentration and observing the increase in product formation rate. We propose a qualitative model demonstrating the relationship between oxygen limitation, biocatalyst concentration, and the rate of reaction. This conceptual model will be a useful guide in the industrial scale-up of whole cell mediated Baeyer-Villiger biocatalysis.

Bioreactors↗
PubMed2005

Directed evolution of biocatalytic processes.

The benefits of applying biocatalysts to organic synthesis, such as their high chemo-, regio-, and enantio-specificity and selectivity, must be seriously considered, especially where chemical routes are unavailable, complex or prohibitively expensive. In cases where a potential biocatalytic route is not yet efficient enough to compete with chemical synthesis, directed evolution, and/or process engineering could be implemented for improvements. While directed evolution has demonstrated great potential to enhance enzyme properties, there will always be some aspects of biocatalytic processes that it does not address. Even where it can be successfully applied, the resources required for its implementation must currently be weighed against the feasibility of, and resources available for developing a chemical synthesis route. Here, we review the potential of combining directed evolution with process engineering, and recent developments to improve their implementation. Favourable targets for the directed evolution of new biocatalysts are the syntheses of highly complex molecules, especially where chemistry, metabolic engineering or recombineering provide a partial solution. We also review some of the recent advances in the application of these approaches alongside the directed evolution of biocatalysts.

Animals↗
PubMed2003

Accelerated design of bioconversion processes using automated microscale processing techniques.

Microscale processing techniques are rapidly emerging as a means to increase the speed of bioprocess design and reduce material requirements. Automation of these techniques can reduce labour intensity and enable a wider range of process variables to be examined. This article examines recent research on various individual microscale unit operations including microbial fermentation, bioconversion and product recovery techniques. It also explores the potential of automated whole process sequences operated in microwell formats. The power of the whole process approach is illustrated by reference to a particular bioconversion, namely the Baeyer-Villiger oxidation of bicyclo[3.2.0]hept-2-en-6-one for the production of optically pure lactones.

Automation↗
PubMed2003

Towards large-scale synthetic applications of Baeyer-Villiger monooxygenases.

Biocatalysis is coming of age, with an increasing number of reactions being scaled-up and developed. The diversity of reactions is also increasing and oxidation reactions have recently been considered for scale-up to commercial processes. One important chemical conversion, which is difficult to achieve enantio- or enantiotopo- selectively, is the Baeyer-Villiger (BV) oxidation of ketones. Using cyclohexanone monooxygenase to catalyse the reaction produces optically pure esters and lactones with exquisite enantiomeric excess values. Recently, these enzymes and their many applications in synthetic chemistry have been explored. The scale-up of these conversions has been examined with the idea of implementing the first commercial Baeyer-Villiger monooxygenase-based process. Here, we review the state-of-the-art situation for the scale-up and exploitation of these enzymes.

Bioreactors↗
PubMed2002

The use of microscale processing technologies for quantification of biocatalytic Baeyer-Villiger oxidation kinetics.

Microscale processing techniques would be a useful tool for the rapid and efficient collection of biotransformation kinetic data as a basis for bioprocess design. Automated liquid handling systems can reduce labor intensity while the small scale reduces the demand for scarce materials such as substrate, product, and biocatalyst. Here we illustrate this concept by establishing the use of several microwell formats (96-round, 96-deep square and 24-round well microtiter plates) for quantification of the kinetics of the E. coli TOP10 [pQR239] resting cell catalyzed Baeyer-Villiger oxidation of bicyclo[3.2.0]hept-2en-6-one using glycerol as a source of reducing power. By increasing the biocatalyst concentration until the biotransformation rate was oxygen mass-transfer limited we can ensure that kinetic data collected are in the region away from oxygen limitation. Using a 96-round well plate the effect of substrate (bicyclo[3.2.0]hept-2en-6-one) concentration on the volumetric CHMO activity was examined and compared to data collected from 1.5-L stirred-tank experiments. The phenomenon and magnitude of substrate inhibition, observed at the larger scale, was accurately reproduced in the microwell format. We have used this as an illustrative example to demonstrate that under adequately defined conditions, automated microscale processing technologies can be used for the collection of quantitative kinetic data. Additionally, by using the experimentally determined stoichiometry for product formation and glycerol oxidation, we have estimated the maximum oxygen transfer rates as a function of well geometry and agitation rate. Oxygen-transfer rates with an upper limit of between 33 mmol. L(-1). h(-1) (based solely on product formation) and 390 mmol. L(-1). h(-1) (based on product formation and glycerol oxidation) were achieved using a 96-square well format plate shaken at 1300 rpm operated with a static surface area to volume ratio of 320 m(2). m(-3).

Bioreactors↗
PubMed2002

The search for the ideal biocatalyst.

While the use of enzymes as biocatalysts to assist in the industrial manufacture of fine chemicals and pharmaceuticals has enormous potential, application is frequently limited by evolution-led catalyst traits. The advent of designer biocatalysts, produced by informed selection and mutation through recombinant DNA technology, enables production of process-compatible enzymes. However, to fully realize the potential of designer enzymes in industrial applications, it will be necessary to tailor catalyst properties so that they are optimal not only for a given reaction but also in the context of the industrial process in which the enzyme is applied.

Animals↗
PubMedDate not supplied

Reactor operation and scale-up of whole cell Baeyer-Villiger catalyzed lactone synthesis.

The recombinant whole cell biocatalyst Escherichia coli TOP10 [pQR239], expressing cyclohexanone monooxygenase from Acinetobacter calcoaceticus NCIMB 9871, was used in 1.5- and 55-L fed-batch processes to oxidize bicyclo[3.2.0]hept-2-en-6-one to its corresponding regioisomeric lactones, (-)-(1S,5R)-2-oxabicyclo[3.3.0]oct-6-en-3-one and (-)-(1R,5S)-3-oxabicyclo[3.3.0]oct-6-en-2-one. By employing a bicyclo[3.2.0]hept-2-en-6-one feed rate below that of the theoretical volumetric biocatalyst activity (275 micromol x min(-1) x L(-1)), the reactant concentration in the bioreactor was successfully maintained below the inhibitory concentration of 0.2-0.4 g x L(-1). In this way approximately 3.5 g x L(-1) of the combined regioisomeric lactones was produced with a yield of product on reactant of 85-90%. The key limitation to the process was shown to be product inhibition. This process was scaled up to 55 L, producing over 200 g of combined lactone product. Using a simple downstream process (centrifugation, adsorption to activated charcoal, 5-fold concentration with ethyl acetate elution, and silica gel chromatography), we have shown that the two regioisomeric lactone products could be isolated and purified at this scale.

Acinetobacter calcoaceticus↗