Fiziologija bakterijskog soja Lactobacillus amylovorus DSM 20531T uzgojenog na maltozi u laboratorijskom mjerilu

master's thesis

• Title (english): Physiology of strain Lactobacillus amylovorus DSM 20531T cultivated on maltose at laboratory scale
• Author: Ines Inkret
• Mentor: Anita Slavica (mentor)
• Granter: University of Zagreb; Faculty of Food Technology and Biotechnology; (Department of Biochemical Engineering); (Laboratory for Biochemical Engineering, Industrial Microbiology and Malting and Brewing Technology); Zagreb
• Defense date and country: 2017-07-21, Croatia
• Scientific / art field, discipline and subdiscipline: BIOTECHNICAL SCIENCES; Biotechnology

Abstract

Amilolitička bakterija mliječne kiseline Lactobacillus amylovorus DSM 20531T uzgojena je u laboratorijskom mjerilu u modificiranoj MRS podlozi sa maltozom kao izvorom ugljika i energije i induktorom sinteze amilaze. Ovaj soj je uzastopnim precjepljivanjem i uzgojem u podlozi sa maltozom prilagoĎen na korištenje disaharida i, nakon toga, u laboratorijskom mjerilu raste brţe i sintetizira znatno manje biomase nego kada nije prilagoĎen na ovaj supstrat. Homofermentativnom razgradnjom maltoze L. amylovorus DSM 20531T proizvodi mliječnu kiselinu kao jedini krajnji proizvod metabolizma i to znatno učinkovitije nakon prilagodbe. Proizvedena amilaza je izdvojena kroz dva postupka - najprije taloţenjem amonijevim sulfatom i, nakon toga, ionsko-izmjenjivačkom kromatografijom. Tri kromatografski izdvojena proteinska pika pokazuju aktivnost prema škrobu u natrijevom acetatnom puferu (50 mmol L-1, pH 5,5 pri 55C). Ovi su rezultati u skladu sa rezultatima biospecifične kromatografije pomoću koje su izdvojena tri proteinska pika, koja takoĎer pokazuju aktivnost prema škrobu. Najveću specifičnu aktivnost od 300 U mg-1 imaju proteini eluirani tijekom ionsko-izmjenjivačke kromatografije pri 0,6 mol L-1 NaCl u puferu. Sva tri izdvojena pika sadrţe više proteinskih vrpci, kako je to detektirano SDS-PAGE metodom, i mase ovih proteina su izmeĎu 50 i 94 kDa.

Abstract (english)

Cultivation of an amylolytic lactic acid bacterium Lactobacillus amylovorus DSM 20531T was carried out in laboratory scale in a modified MRS medium with maltose as a main carbon and energy source and as an inductor of amylase synthesis. The strain was adapted to use disaccharide by iterative cultivation in the medium with maltose and, afterwards, it grew faster and considerably lower concentration of a biomass was synthesized compared to unadapted conditions. The maltose was homofermentatively degraded by L. amylovorus DSM 20531T and lactic acid was produced more efficiently as a single final product of metabolism after the adaptation. Produced amylase was separated in a two-step procedure which consisted of precipitation by ammonium sulfate and ion-exchange chromatography. Three protein peaks were chromatographycally separated and all of them showed activity toward starch in a sodium acetate buffer (50 mmol L-1, pH 5,5 at 55C). Obtained results are in agreement with results of biospecific chromatography when three protein peaks with activity toward the starch were separated as well. The proteins that were eluted during the ion-exchange chromatography at 0.6 mol L-1 NaCl in the buffer had the highest specific activity of 300 U mg-1. All three separated peaks contained more protein bands, as revealed by SDS-PAGE, and their molecular masses were estimated to be between 50 and 94 kDa.

• Keywords: fiziologija amilolitičke bakterije mliječne kiseline; maltoza; Lactobacillus amylovorus; laboratorijsko mjerilo; amilaza; kromatografija.
• Keywords (english): physiology of amylolytic lactic acid bacterium; maltose; Lactobacillus amylovorus; laboratory scale; amylase; chromatography
• Language: croatian
• URN:NBN: urn:nbn:hr:159:102139
• Study programme: Title: Bioprocess Engineering; Study programme type: university; Study level: graduate; Academic / professional title: magistar/magistra inženjer/inženjerka bioprocesnog inženjerstva (magistar/magistra inženjer/inženjerka bioprocesnog inženjerstva)
• Type of resource: Text
• File origin: Born digital
• Access conditions: Open access
• Created on: 2017-11-07 10:36:14