Inhibitory effect of propafenone derivatives on Pseudomonas aeruginosa biofilm and pyocyanin production

Abstract

Introduction/Objective Biofilm and pyocyanin production are essential components of Pseudomonas aeruginosa virulence and antibiotic resistance. Our objective was to examine inhibitory effect of synthetized propafenone derivatives 3-(2-Fluorophenyl)-1-(2(2-hydroxy-3-propylamino-propoxy)-phenyl)-propan-1-one hydrochloride (5OF) and 3-(2-Trifluoromethyl-phenyl)-1-(2-(2-hydroxy-3-propylamino-propoxy)-phenyl)-propan-1-one hydrochloride (5CF3) on biofilm and pyocyanin in Pseudomonas aeruginosa clinical strains. Methods Effects were tested on nine clinical isolates and one control laboratory strain of P. aeruginosa. In vitro analysis of biofilm growing was performed by incubating bacteria (0.5 McFarland) with 5OF and 5CF3 (500-31.2 µg/ml) and measuring optical density (OD) at 570 nm. Bacteria in medium without compounds were positive control. Blank medium (an uninoculated medium without test compounds) was used as negative control. Pyocyanin production was estimated by OD at 520 nm, after bacteria incubated with 5CF3 and 5OF (250 and 500 mg/ml), treated with chloroform, and chloroform layer mixed with HCl. Results A total of 500 µg/ml of 5OF and 5CF3 completely inhibited biofilm formation in 10/10 and 4/10 strains, respectively. A total of 250 µg/ml of 5OF and 5CF3 strongly inhibited biofilm formation in 7/10 strains, while inhibition with 125 µg/ml of 5OF and 5CF3 was moderate. Lower concentrations had almost no effect on biofilm production. Pyocyanin production was reduced to less than 40% of the control value in 6/9, and less than 50% of the control in 7/9 strains with 500 mg/ml of 5OF and 5CF3, respectively. At 250 µg/ml 5OF and 5CF3, most strains had pyocyanin production above 50% of the control value. Conclusion Synthetized propafenone derivatives, 5OF and 5CF3, inhibited biofilms and pyocyanin production of Pseudomonas aeruginosa clinical strains. Presented results suggest that propafenone derivatives are potential lead-compounds for synthesis of novel antipseudomonal drugs.

Uvod/Cilj Proizvodnja biofilma i piocijanina je važan faktor virulencije i antibiotske rezistencije bakterije Pseudomonas aeruginosa. Cilj rada je bio da se ispita inhibitorni efekat sintetisanih propafenonskih derivata, 3-(2-fluoro-fenil)-1-[2-(2-hidroksi-3-propilamino-propoksi)-fenil]-propan-1-on-hidrohlorid) (5OF) i 3-(2-trifluorometilfenil)-1-[2-(2-hidroksi-3-propilamino-propoksi)-fenil]-propan-1-on-hidrohlorid) (5CF3), na produkciju biofilma i piocijanina kod kliničkih izolata bakterije Pseudomonas aeruginosa. Metode Efekat propafenonskih derivata ispitan je na devet kliničkih izolata i jednom standardnom soju bakterije P. aeruginosa. Uticaj na produkciju biofilma ispitan je in vitro, inkubacijom bakterija (0,5 po Makfarlandu) sa 5OF i 5CF3 (500-31,2 µg/ml), i merenjem optičke gustine na 570 nm. Bakterije u medijumu bez ispitivanih jedinjenja su bile pozitivna kontrola, a sam medijum negativna kontrola. Proizvodeni piocijanin, koji je određivan merenjem optičke gustine na 520 nm, na koinkubacije bakterija sa 5CF3 ili 5OF (250 i 500 mg/ml), tretiran je hloroformom i mešanjem hloroformskog sloja sa HCl. Rezultati Pri koncentraciji od 500 µg/ml 5OF je doveo do potpune inhibicije produkcije biofilma kod svih ispitivanih sojeva (10/10). Inhibicija biofilma sa 500 µg/ml 5CF3 bila je potpuna kod 4/10 sojeva. Pri koncentraciji 5OF i 5CF3 od 250 µg/ml produkcija biofilma kod većine ispitanih izolata bila je slaba, dok je pri koncentraciji 125 µg/ml 5OF odnosno 5CF3 produkcija bila umerena. Niže koncentracije 5OF i 5CF3 nisu imale inhibitorni efekat na formiranje biofilma. U prisustvu 500 mg/ml 5OF u 6/10 ispitivanih sojeva produkcija piocijanina pala je na manje od 40% u odnosu na kontrolnu vrednost. Ista koncentracija (500 mg/ml) 5CF3 snizila je produkciju piocijanina na manje od 50% od kontrole u 7/9 sojeva. Pri koncentraciji 250 µg/ ml 5OF ili 5CF3 većina sojeva produkovala je piocijanin iznad 50% u odnosu na pozitivnu kontrolu. Zaključak Sintetisani propafenonski derivati, 5OF i 5CF3, inhibiraju produkciju biofilma i piocijanina kod kliničkih sojeva bakterije Pseudomonas aeruginosa. Dobijeni rezultati ukazuju na to da propafenonski derivati predstavljaju moguća polazna jedinjenja za sintezu novih antipseudomonasnih agenasa.