Biokatalitička sinteza (R)-4-kloro-3-hidroksibutironitrila katalizirana s halogenhidrin-dehalogenazom

master's thesis

• Title (english): Biocatalytic synthesis of (R)-4-chloro-3-hydroxybutyronitrile catalysed by halohydrin dehalogenase
• Author: Kristina Matić
• Mentor: Zvjezdana Findrik Blažević (mentor)
• Committee member: Zvjezdana Findrik Blažević (predsjednik povjerenstva)
• Committee member: Dragana Vuk (član povjerenstva)
• Committee member: Marija Vuković Domanovac (član povjerenstva)
• Granter: University of Zagreb; Faculty of Chemical Engineering and Technology; Zagreb
• Defense date and country: 2022-07-08, Croatia
• Scientific / art field, discipline and subdiscipline: TECHNICAL SCIENCES; Chemical Engineering

Abstract

Enzimi su biokatalizatori koji kataliziraju reakcije u živim organizmima. Njihova primjena omogućuje niz prednosti, poput odvijanja reakcija u blažim uvjetima te smanjenje nastanka štetnih nusprodukata. Prirodna uloga halogenhidrin-dehalogenaza je kataliza halohidrina u odgovarajuće epokside (reakcija dehalogenacije), odnosno kataliza reakcije zatvaranja epoksidnog prstena pri čemu dolazi do otpuštanja halogenih iona. U ovom radu je ispitana reakcija biokatalitičke sinteze (R)-4-klor-3-hidroksibutironitrila uz B-tip halogenhidrin-dehalogenaze. Sinteza optički čistih tvari izuzetno je važan dio farmaceutske industrije s obzirom na to da dva enantiomera mogu imati potpuno različita svojstva, a vrlo često jedan od njih može pokazivati štetno i toksično biološko djelovanje. Epoksidi su visoko vrijedni spojevi u organskoj sintezi zbog toga što se epoksidni prsten može kemijski transformirati u brojne sintetski korisne intermedijere. Reakcije u kojima su uključeni enzimi potrebno je pomno istražiti i okarakterizirati kako bi se mogle provoditi u industrijskom mjerilu. Reakcija biokatalitičke sinteze (R)-4-klor-3-hidroksibutironitrila provedena je u kotlastom reaktoru uz natrijev cijanid kao nukleofil, a uzorci su analizirani uporabom plinske kromatografije. Ispitan je utjecaj koncentracija reakcijskih komponenti na brzinu enzimske reakcije te je pokazan nizak afinitet enzima prema supstratu i nukleofilu kod oba enzima, a također je uočen značajan inhibicijski učinak produkta. Aktivnost enzima praćena je spektrofotometrijski, a rezultati su pokazali da aktivnost kod oba enzima pada s vremenom, odnosno dolazi do njihove deaktivacije.

Abstract (english)

Enzymes are biocatalysts that catalyze reactions in living cells. Their usage provides a number of advantages, such as conducting a reaction in milder conditions and a reduction in the occurrence of harmful by-products. The natural role of halohydrin-dehalogenases is the catalysis of halohydrins to the corresponding epoxides (dehalogenation). Halohydrin-dehalogenase catalyzes the epoxy ring closure in which halogen ions are released. In this work, the reaction of biocatalytic synthesis of (R)-4-chloro-3-hydroxybutyronitrile with type B halohydrin-dehalogenase was investigated. The synthesis of optically pure substances is an extremely important part of the pharmaceutical industry since two enantiomers can have completely different properties and very often one of them can show harmful and toxic biological effects. Epoxides are highly valuable compounds in organic synthesis because the epoxy ring can be chemically transformed into a number of synthetic beneficial intermediates. Reactions involving enzymes need to be carefully investigated and characterized in order to be carried out on an industrial scale. The biocatalytic synthesis of (R)-4-chloro-3-hydroxybutyronitrile was carried out in a batch reactor with sodium cyanide as a nucleophile. The samples were analyzed using gas chromatography. The influence of the concentration of the reaction components on the rate of the enzymatic reaction was tested. Low enzyme affinity for the substrate and nucleophile was observed in reactions with both enzymes. A significant inhibitory effect of the product was also shown. The activity of the enzyme was monitored by spectrophotometer and it was observed that in both enzymes there is a decrease in activity over time.

• Keywords: biokataliza; enzimi; halogenhidrin-dehalogenaza; epoksid; (R)-4-klor-3-hidroksibutironitril
• Keywords (english): biocatalysis; enzymes; halohydrin-dehalogenase; epoxide; (R)-4-chloro-3-hydroxybutyronitrile
• Language: croatian
• URN:NBN: urn:nbn:hr:149:185891
• Project: Number: KK.01.1.1.04.0013; Title: Inovativna rješenja u katalitičkim proizvodnim procesima za potrebe farmaceutske industrije; Title: Innovative solutions in catalytic processes in the pharmaceutical industry; Acronym: CAT PHARMA; Leader: Srećko Kirin; Jurisdiction: eu; Funder: EC; Funding stream: EFRR
• Study programme: Title: Chemical Engineering - Graduate study; Study programme type: university; Study level: graduate; Academic / professional title: magistar/magistra inženjer/inženjerka kemijskog inženjerstva (magistar/magistra inženjer/inženjerka kemijskog inženjerstva)
• Type of resource: Text
• File origin: Born digital
• Access conditions: Embargoed access; Embargo expiration date: 2024-07-15
• Created on: 2023-02-26 16:34:36