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Title:Structure optimisation of biopigment prodigiosin from Serratia marcescens ATCC 27117 and antimicrobial and anticancer properties of novel halogenated derivatives
Authors:ID Nikodinović-Runić, Jasmina (Mentor) More about this mentor... New window
ID Lazić, Jelena (Author)
Files:.pdf Jelena_Lazic.pdf (3,39 MB)
MD5: F78E3608FDD6454A6A6F44889C10968A
 
Language:English
Work type:Doctoral dissertation
Typology:2.08 - Doctoral Dissertation
Organization:FPŠ - Graduate School
Abstract:Prodigiosins (PGs) are a class of bacterial secondary metabolites with remarkable biological activities and colour. In this study, optimised fermentative production of prodigiosin (PG) using waste processed meat as a substrate has been achieved to levels of 83.1 ± 3.0 mg/L from a commercially available Serratia marcescens ATCC 27117 strain within 12 h. Methods were established for the reliable PG extraction from both the bacterial cell pellet and the culture supernatant, while gravitation column chromatography was used to obtain pure bacterial PG. The structure of the isolated PG was optimised by environmentally acceptable oxidative bromination reactions, obtaining mono- and dibrominated derivatives (PG-Br and PG-Br2). Chemical structures were confirmed by structural characterisation using nuclear magnetic resonance (NMR) spectroscopy and mass spectrometry (MS), showing that PG-Br is a mixture of two monobrominated isomers in approximately equal ratios, while PG-Br2 was afforded as a pure derivative. PG and its brominated derivatives (Br-derivatives) showed anticancer potential with half-maximal inhibitory concentration (IC50) values ranging from 0.62 to 17.00 μg/mL on four tested cancer cell lines (A549 lung, A375 skin, MDA-MB-231 breast, HCT116 colon) and an induction of early apoptosis, but low selectivity against healthy cell lines (MRC-5 lung fibroblasts and HaCaT skin keratinocytes). All three PG, PG-Br and PG-Br2 compounds did not affect roundworms (Caenorhabditis elegans) at concentrations up to 50 μg/mL. However, an improved toxicity profile of Br-derivatives in comparison to the parent PG was observed in vivo using zebrafish (Danio rerio) model system, when 10 μg/mL applied at 6 h post fertilisation caused death rate of 100, 30 and 0% by PG, PG-Br and PG-Br2, respectively, which is a significant finding for further structural optimisations of bacterial PGs.
Keywords:prodigiosin, Serratia marcescens, meat waste, halogenation, novel derivatives, anticancer activity, apoptosis, zebrafish embryotoxicity
Place of publishing:Nova Gorica
Year of publishing:2022
PID:20.500.12556/RUNG-7664 New window
COBISS.SI-ID:126628867 New window
NUK URN:URN:SI:UNG:REP:PBQYMKIK
Publication date in RUNG:21.10.2022
Views:1802
Downloads:32
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Secondary language

Language:Slovenian
Title:Strukturna optimizacija biopigmenta prodigiozin iz bakterije Serratia marcescens ATCC 27117 ter antimikrobne in antitumorske lastnosti novih halogeniranih derivatov
Abstract:Prodigiozini (PG) so razred obarvanih sekundarnih bakterijskih metabolitov z izjemno biološko aktivnostjo. V doktorski raziskovalni nalogi smo za proizvodnjo prodigiozina uporabili komercialni sev bakterije Serratia marcescens ATCC 27117, kot substrat za fermentacijo pa odpadne produkte procesiranja mesa. Na ta način smo pridobili do 83.1 ± 3.0 prodigiozina v 12-ih urah fermentacije. V nadaljevanju smo vzpostavili metode za ekstrakcijo PS tako iz peleta bakterijskih celic kot tudi iz supernatanta bakterijske kulture. Za pridobitev čistega bakterijskega PG smo uporabili gravitacijsko kolonsko kromatografijo. Strukturo izoliranega PG smo optimizirali z okolju prijazno metodo oksidativnega bromiranja, pri čemer smo dobili mono- in di-bromirane derivate (PG-Br in PG-Br2). Strukturna karakterizacija bromiranih spojin s pomočjo jedrske magnetne resonance (NMR) in masne spektrometrije (MS) je pokazala, da je PG-Br zmes dveh mono-bromiranih izomer v približno enakih razmerjih, medtem ko je bil PG-Br2 pridobljen kot čisti derivat. PG in njegove bromirane derivate smo testirali na štirih rakavih celičnih linijah (A549 – pljučne celice, A375 – kožne celice, MDA-MB-231 – celice raka na prsih in HCT116 – črevesne celice). Potrdili smo antitumorske lastnosti PG in njegovih bromiranih derivatov s polovičnimi maksimalnimi inhibitornimi koncentracijami (IC50) v razponu od 0,62 do 17,00 μg/mL ter indukcijo zgodnje apoptoze, vendar hkrati tudi nizko selektivnost proti zdravim celičnim linijam (MRC-5 - pljučni fibroblasti in HaCaT - kožni keratinociti). Nobena od testiranih PG, PG-Br in PG-Br2 spojin ni imela opaznega vpliva na gliste Caenorhabditis elegans pri koncentracijah do 50 μg/mL. Ugotovili pa smo manjšo toksičnost bromiranih derivatov PG v primerjavi z matičnim PG v modelnem sistemu rib cebric (Danio rerio) in vivo. Pri cebricah, ki so bile 6 ur po oploditvi izpostavljene 10 μg/mL PG, PG-Br oziroma PG-Br2, smo ugotovili 100%, 30% in 0% smrtnost, kar je pomembna ugotovitev za nadaljnjo strukturno optimizacijo bakterijskih PG.
Keywords:prodigiozin, Serratia marcescens, mesni odpadki, halogeniranje, novi derivati, antitumorsko delovanje, apoptoza, embriotoksičnost cebric


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