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Oxidation of Ketones: A (Chemo-) Enzymatic Approach Using Oxygenases and Hydrolases

Chávez, Georgina LU (2013)
Abstract (Swedish)
Popular Abstract in English

The thesis that you are reading at this moment was printed using climate compensated paper, a sign that consciously or not you are contributing somehow to control the current environmental issues in our planet. It would not be nice if every single article used in your daily life contains information about how environmentally friendly that product is? We have to think “big” and start to work for a real solution. It is of common knowledge that the chemical industry has changed the world by providing us with innumerable products but at the same time has been one of the major contributors to pollution on our planet. Investments on research to allow us to make a green process along with policies... (More)
Popular Abstract in English

The thesis that you are reading at this moment was printed using climate compensated paper, a sign that consciously or not you are contributing somehow to control the current environmental issues in our planet. It would not be nice if every single article used in your daily life contains information about how environmentally friendly that product is? We have to think “big” and start to work for a real solution. It is of common knowledge that the chemical industry has changed the world by providing us with innumerable products but at the same time has been one of the major contributors to pollution on our planet. Investments on research to allow us to make a green process along with policies regulating the production of chemicals can be a real solution to several of our environmental problems. Oxidations are important reactions in chemistry. Oxidations in industry are done generally using chlorinated solvents, large amounts of chemicals some of which pose risk of explosion during transportation and storage. A greener alternative is the use of enzymes which are able to oxidise compounds with molecular oxygen (as monooxygenases) or enzymes able to produce oxidants during the process (lipases and perhydrolases). Enzymes usually work in mild conditions, making the process not just greener but also safer as avoid the risk of explosion. In addition, green reactions should also consider changing traditional solvents to greener alternatives, especially considering that solvents per se constitute around 80% of the generated organic waste. This thesis concerns work with the above-mentioned enzymes with a target to produce lactones, the products used in flavors, fragrances, pharmaceuticals and polymers. To contribute to the efforts of green chemistry, the work covers the development of a new colorimetric method for the search of oxidizing enzymes from bacteria, the description of one such enzyme from a bacteria isolated from a soda lake, and development of an enzymatic process to produce caprolactone. Who knows, maybe in the future if you decide to read this thesis again, in the corner of your glasses in very tiny letters you can read something like: this product was made using materials according to green chemistry regulation (Less)
Abstract
Oxidation reactions are important in organic chemistry as well as in nature. In industry, oxidations are commonly used for the synthesis of chemicals and pharmaceuticals, however such processes have a number of limitations, they use chlorinated solvents, stoichiometric oxidation reagents, and in some cases the reagents that have risks of explosion during transportation and storage. This has called for more environment-friendly alternative technologies for oxidation reactions. Baeyer-Villiger oxidation is a reaction in which a ketone is oxidized to an ester or a cyclic ketone to a lactone by treatment with peroxyacids. Lactones constitute an important group of chemicals used in flavors, fragrances, pharmaceutical intermediates and polymer... (More)
Oxidation reactions are important in organic chemistry as well as in nature. In industry, oxidations are commonly used for the synthesis of chemicals and pharmaceuticals, however such processes have a number of limitations, they use chlorinated solvents, stoichiometric oxidation reagents, and in some cases the reagents that have risks of explosion during transportation and storage. This has called for more environment-friendly alternative technologies for oxidation reactions. Baeyer-Villiger oxidation is a reaction in which a ketone is oxidized to an ester or a cyclic ketone to a lactone by treatment with peroxyacids. Lactones constitute an important group of chemicals used in flavors, fragrances, pharmaceutical intermediates and polymer building blocks. The work presented in this thesis concerns enzymes, including Baeyer-Villiger monooxygenases (BVMOs) that catalyse the Baeyer-Villiger oxidation using molecular oxygen as an oxidant, and perhydrolytic enzymes that can be used for in situ generation of peracid for oxidation of cyclic ketones. A simple colorimetric method was developed for detection of BVMO activity and was based on the formation of a purple colored product between an enolizable ketone and 3,5-dinitrobenzoic acid in an alkaline solution. The method was shown to have potential for screening of both wild type and recombinant microbial cells as well as for quantitative measurement of BVMO activity. Further, a recombinant BVMO from a strain of Dietzia was characterized. The sequence of the enzyme suggested that it is related to Ethionamide monooxygenases. The recombinant enzyme was active in whole cells and crude lysate but lost activity on purification. The enzyme was shown to have high activity towards several linear alkenes, and was also moderately active towards cyclobutanone, phenylacetone and thioanisole. Two perhydrolytic enzymes able to produce peracids from a carboxylic ester and hydrogen peroxide were studied for oxidation of cyclohexanone to caprolactone, a chemical of immense importance. The enzymes were immobilized as cross-linked enzyme aggregates (CLEAs). The well-studied lipase B from Candida antarctica (CaLB) gave a maximal caprolactone yield of 80% with ethyl acetate as acyl donor. The perhydrolase was able to produce peracids in an aqueous medium with ethylene glycol diacetate and hydrogen peroxide, and gave caprolactone yield of 70%. In both cases the formation as acetic acid as a coproduct showed to be an important factor for the deactivation of the enzyme.



The use of monooxygenases, lipases and perhydrolases for the Baeyer-Villiger reaction constitutes a greener alternative to traditional chemical processes but the problem of enzyme stability remains to be solved. (Less)
Please use this url to cite or link to this publication:
author
supervisor
opponent
  • Prof. Clarsund, Mats, Enzymatica AB, Lund, Sweden
organization
publishing date
type
Thesis
publication status
published
subject
keywords
cross linked enzyme aggregates., caprolactone, perhydrolase, Lipase B from Candida antarctica
pages
120 pages
publisher
Lund University (Media-Tryck)
defense location
Lecture Hall B, at the Center of Chemistry and Chemical Engineering, Sölvegatan 39, Lund
defense date
2013-05-30 10:30
ISBN
978-91-89627-93-2
language
English
LU publication?
yes
id
3c01d967-3248-4dc2-81ed-8b4fffcb52b3 (old id 3735323)
date added to LUP
2013-05-07 08:37:44
date last changed
2016-09-19 08:45:05
@phdthesis{3c01d967-3248-4dc2-81ed-8b4fffcb52b3,
  abstract     = {Oxidation reactions are important in organic chemistry as well as in nature. In industry, oxidations are commonly used for the synthesis of chemicals and pharmaceuticals, however such processes have a number of limitations, they use chlorinated solvents, stoichiometric oxidation reagents, and in some cases the reagents that have risks of explosion during transportation and storage. This has called for more environment-friendly alternative technologies for oxidation reactions. Baeyer-Villiger oxidation is a reaction in which a ketone is oxidized to an ester or a cyclic ketone to a lactone by treatment with peroxyacids. Lactones constitute an important group of chemicals used in flavors, fragrances, pharmaceutical intermediates and polymer building blocks. The work presented in this thesis concerns enzymes, including Baeyer-Villiger monooxygenases (BVMOs) that catalyse the Baeyer-Villiger oxidation using molecular oxygen as an oxidant, and perhydrolytic enzymes that can be used for in situ generation of peracid for oxidation of cyclic ketones. A simple colorimetric method was developed for detection of BVMO activity and was based on the formation of a purple colored product between an enolizable ketone and 3,5-dinitrobenzoic acid in an alkaline solution. The method was shown to have potential for screening of both wild type and recombinant microbial cells as well as for quantitative measurement of BVMO activity. Further, a recombinant BVMO from a strain of Dietzia was characterized. The sequence of the enzyme suggested that it is related to Ethionamide monooxygenases. The recombinant enzyme was active in whole cells and crude lysate but lost activity on purification. The enzyme was shown to have high activity towards several linear alkenes, and was also moderately active towards cyclobutanone, phenylacetone and thioanisole. Two perhydrolytic enzymes able to produce peracids from a carboxylic ester and hydrogen peroxide were studied for oxidation of cyclohexanone to caprolactone, a chemical of immense importance. The enzymes were immobilized as cross-linked enzyme aggregates (CLEAs). The well-studied lipase B from Candida antarctica (CaLB) gave a maximal caprolactone yield of 80% with ethyl acetate as acyl donor. The perhydrolase was able to produce peracids in an aqueous medium with ethylene glycol diacetate and hydrogen peroxide, and gave caprolactone yield of 70%. In both cases the formation as acetic acid as a coproduct showed to be an important factor for the deactivation of the enzyme. <br/><br>
 <br/><br>
The use of monooxygenases, lipases and perhydrolases for the Baeyer-Villiger reaction constitutes a greener alternative to traditional chemical processes but the problem of enzyme stability remains to be solved.},
  author       = {Chávez, Georgina},
  isbn         = {978-91-89627-93-2},
  keyword      = {cross linked enzyme aggregates.,caprolactone,perhydrolase,Lipase B from Candida antarctica},
  language     = {eng},
  pages        = {120},
  publisher    = {Lund University (Media-Tryck)},
  school       = {Lund University},
  title        = {Oxidation of Ketones: A (Chemo-) Enzymatic Approach Using Oxygenases and Hydrolases},
  year         = {2013},
}