Improving lipase performance in non-aqueous reactions through rational design of treatments

Persistent Link:
http://hdl.handle.net/10150/280178
Title:
Improving lipase performance in non-aqueous reactions through rational design of treatments
Author:
Garcia, Rafael Andres
Issue Date:
2002
Publisher:
The University of Arizona.
Rights:
Copyright © is held by the author. Digital access to this material is made possible by the University Libraries, University of Arizona. Further transmission, reproduction or presentation (such as public display or performance) of protected items is prohibited except with permission of the author.
Abstract:
Enzymes can be used to catalyze reactions in non-aqueous media, which can be particularly useful for the processing of materials with low water solubility. One such reaction is the lipase-catalyzed esterification between citronellol and acetic acid, in a medium of hexane. Both reaction conditions and special pretreatments on the lipase impact the lipase's catalytic performance. This project focuses on improving lipase performance by optimizing its pretreatment. The pretreatment factors studied include pH adjustment, lyophilization time, addition of buffer salts, non-buffer salts, denaturants or active site protectants to the lyophilizate, and flash-freezing in the presence of phase interfaces. The effectiveness of a particular pretreatment is presented in terms of 'relative performance' (RP), which is equal to the number of times faster the pretreated lipase catalyzes the reaction relative to untreated lipase. The individual and interactive effects of the pretreatment factors were studied in detail and compared. Buffer salts had a much stronger performance enhancing effect than non-buffer salts; pretreatment with 90% (w/w) sodium phosphate yielded lipase with an RP of ∼64 (catalyzed the reaction ∼64 times faster that untreated lipase). A strong interaction between the treatments with sodium phosphate and pH adjustment was found. Both these treatments may mitigate the inhibitory effect of acetic acid, one of the substrates. Activating effects of phase interfaces and active site protectants are shown to be complementary to other treatments, demonstrating that they act by a distinct mechanism. An optimization process known as 'the method of steepest ascent' was used to simultaneously optimize the pH and buffer salt pretreatments, as well as the concentration of acetic acid in the reaction mixture, This process succeeded in rapidly developing pretreatment combinations that yielded lipase with RP as high as 170. Studies were also conducted to develop methods for recovery, reprocessing and reuse of lipase from completed reactions.
Type:
text; Dissertation-Reproduction (electronic)
Keywords:
Engineering, Agricultural.; Engineering, Chemical.
Degree Name:
Ph.D.
Degree Level:
doctoral
Degree Program:
Graduate College; Agricultural and Biosystems Engineering
Degree Grantor:
University of Arizona
Advisor:
Riley, Mark R.

Full metadata record

DC FieldValue Language
dc.language.isoen_USen_US
dc.titleImproving lipase performance in non-aqueous reactions through rational design of treatmentsen_US
dc.creatorGarcia, Rafael Andresen_US
dc.contributor.authorGarcia, Rafael Andresen_US
dc.date.issued2002en_US
dc.publisherThe University of Arizona.en_US
dc.rightsCopyright © is held by the author. Digital access to this material is made possible by the University Libraries, University of Arizona. Further transmission, reproduction or presentation (such as public display or performance) of protected items is prohibited except with permission of the author.en_US
dc.description.abstractEnzymes can be used to catalyze reactions in non-aqueous media, which can be particularly useful for the processing of materials with low water solubility. One such reaction is the lipase-catalyzed esterification between citronellol and acetic acid, in a medium of hexane. Both reaction conditions and special pretreatments on the lipase impact the lipase's catalytic performance. This project focuses on improving lipase performance by optimizing its pretreatment. The pretreatment factors studied include pH adjustment, lyophilization time, addition of buffer salts, non-buffer salts, denaturants or active site protectants to the lyophilizate, and flash-freezing in the presence of phase interfaces. The effectiveness of a particular pretreatment is presented in terms of 'relative performance' (RP), which is equal to the number of times faster the pretreated lipase catalyzes the reaction relative to untreated lipase. The individual and interactive effects of the pretreatment factors were studied in detail and compared. Buffer salts had a much stronger performance enhancing effect than non-buffer salts; pretreatment with 90% (w/w) sodium phosphate yielded lipase with an RP of ∼64 (catalyzed the reaction ∼64 times faster that untreated lipase). A strong interaction between the treatments with sodium phosphate and pH adjustment was found. Both these treatments may mitigate the inhibitory effect of acetic acid, one of the substrates. Activating effects of phase interfaces and active site protectants are shown to be complementary to other treatments, demonstrating that they act by a distinct mechanism. An optimization process known as 'the method of steepest ascent' was used to simultaneously optimize the pH and buffer salt pretreatments, as well as the concentration of acetic acid in the reaction mixture, This process succeeded in rapidly developing pretreatment combinations that yielded lipase with RP as high as 170. Studies were also conducted to develop methods for recovery, reprocessing and reuse of lipase from completed reactions.en_US
dc.typetexten_US
dc.typeDissertation-Reproduction (electronic)en_US
dc.subjectEngineering, Agricultural.en_US
dc.subjectEngineering, Chemical.en_US
thesis.degree.namePh.D.en_US
thesis.degree.leveldoctoralen_US
thesis.degree.disciplineGraduate Collegeen_US
thesis.degree.disciplineAgricultural and Biosystems Engineeringen_US
thesis.degree.grantorUniversity of Arizonaen_US
dc.contributor.advisorRiley, Mark R.en_US
dc.identifier.proquest3073223en_US
dc.identifier.bibrecord.b43468196en_US
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