Optimiranje postupaka ekstrakcije funkcionalnih biomolekula Lactobacillus sojeva

undergraduate thesis

• Title (english): Optimization of extraction of functional biomolecules of Lactobacillus strains
• Author: Belma Karamehmedović
• Mentor: Jasna Novak (mentor)
• Granter: University of Zagreb; Faculty of Food Technology and Biotechnology; (Department of Biochemical Engineering); (Laboratory for Antibiotic, Enzyme, Probiotic and Starter Cultures Technology); Zagreb
• Defense date and country: 2022-09-07, Croatia
• Scientific / art field, discipline and subdiscipline: BIOTECHNICAL SCIENCES; Biotechnology

Abstract

Površinske strukture probiotičkih bakterija su ključne molekule prilikom interakcija s mikrookolišom gastrointestinalnog trakta, te se pojedine istražuju kao funkcionalne biomolekule. Upravo predstavnici ovih biomolekula su egzopolisaharidi (EPS) koji su izvanstanični šećerni polimeri, odnosno S-proteini koji su skupina površinskih proteina koje eksprimiraju pojedini Lactobacillus sojevi. Funkcionalne biomolekule Lactobacillus sojeva se intenzivno istražuju te se prema recentnim istraživanjima definiraju kao postbiotici. Za strukturnu i funkcionalnu analizu ključno je provesti učinkovitu ekstrakciju ciljanih molekula. Upravo u ovom radu provedeno je optimiranje uvjeta ekstrakcije EPS iz odabranih Lactobacillus sojeva mikrobioma humanog mlijeka. EPS je ekstrahiran iz soja Limosilactobacillus fermentum MC1, a S-proteini iz sojeva Levilactobacillus brevis MB1, MB2, MB13, MB20. Dijalizirani uzorci stanično vezanih EPS-b izolirani su primjenom tri različita agensa NaOH, fenol, EDTA. Najveći prinos od YEPS=544,12 mg/L određen je u uzorku EPS-b nakon primjene NaOH. Maksimalna deproteinizacija i koncentracija ekstracelularnih EPS (611,11 mg/L) (EPS-r), eksperimentalno je određena u uzorku ekstrahiranom u postupku pročišćavanja koji uključuje trikloroctenu kiselinu. SDS-PAGEom površinskih proteina ustanovljena je prisutnost S-proteina kod sva četiri soja L. brevis. Metodom po Bradfordu određena je koncentracija ekstrahiranih S-proteina pri čemu je najveća koncentracija (27,286 mg/µL) određena u uzorku površinskih proteina soja L. brevis MB20.

Abstract (english)

The surface structures of probiotic bacteria are key molecules interacting with the microenvironment of the gastrointestinal tract, and some are being investigated as functional biomolecules. The representatives of these biomolecules are extracellular sugar polymers exopolysaccharides (EPS) and Sproteins, a group of surface proteins expressed by certain strains of Lactobacillus. Functional biomolecules of Lactobacillus spp. are intensively investigated and, according to recent research, are defined as postbiotics. Efficient extraction is crucial for the structural and functional analysis of target biomolecules. In this work, the optimization of EPS extraction conditions from selected Lactobacillus strains of the human milk microbiome was carried out. EPS was extracted from Limosilactobacillus fermentum MC1, and S-proteins from Levilactobacillus brevis strains MB1, MB2, MB13, MB20. Dialyzed samples of cell-bound EPS-b were isolated using three different agents: NaOH, phenol, and EDTA. The highest yield of YEPS=544.12 mg/L was determined in the EPS-b sample treated with NaOH. The maximum concentration of extracellular EPS (611.11 mg/L) (EPS-r) was experimentally determined in a sample extracted when trichloroacetic acid (TCA) was applied. SDS-PAGE of surface proteins established the presence of S-protein in all four strains of L. brevis. The concentration of
extracted S-proteins was determined using the Bradford method, with the highest concentration (27.286 mg/µL) being determined in the sample of surface proteins of the strain L. brevis MB20.

• Keywords: Limosilactobacillus fermentum; Levilactobacillus brevis; egzopolisaharidi; S-proteini; ekstrakcija; postbiotici
• Keywords (english): Limosilactobacillus fermentum; Levilactobacillus brevis; exopolysaccharides; S-proteins; extraction; postbiotics
• Language: croatian
• URN:NBN: urn:nbn:hr:159:232810
• Study programme: Title: Biotechnology; Study programme type: university; Study level: undergraduate; Academic / professional title: sveučilišni/a prvostupnik/prvostupnica (baccalaureus/baccalaurea) inženjer/inženjerka biotehnologije (sveučilišni/a prvostupnik/prvostupnica (baccalaureus/baccalaurea) inženjer/inženjerka biotehnologije)
• Type of resource: Text
• File origin: Born digital
• Access conditions: Open access
• Created on: 2024-04-08 09:42:47