Antimicrobical activity of phytols and mechanisms of action on Gram-negative bacteria

Cilj ovog rada je utvrđivanje spektra antimikrobne aktivnosti kao i mehanizama djelovanja diterpenskog alkohola fitola na široki spektar Gram-pozitivnih i Gram- negativnih bakterija uključujući vrste Enterococcus faecalis, Bacillus cereus, Escherichia coli, Staphylococcus aureus, Aeromonas hydrophila, Klebsiella pneumoniae i Pseudomonas aeruginosa.. Primjenom metode disk difuzije utvrđene su inhibicije rasta na ispitivane vrste bakterija. Najmanja zona inhibicije rasta zabilježena je kod vrste Staphylococcus aureus i iznosila je 13,1 mm dok je najveću osjetljivost pokazala vrsta Pseudomonas aeruginosa sa zonom inhibicije rasta od 29,9 mm. Osjetljivost na antibiotike uz dodatak različitih koncentracija fitola i bez fitola kod Klebsiella pneumoniae ESBL producirajućih sojeva određena je primjenom metode disk difuzije na Mueller-Hinton agru. U istraživanju je korišten široki spektar klinički značajnih antibiotika. Fitol je dodan na diskove s antibioticima u različitim koncentracijama te je zabilježena inhibicija rasta pri koncentraciji od 10 μg/ml u kombinaciji s polimiksinom B i ceftazidinom. Kod viših ispitivanih koncentracija nije zabilježena značajna razlika u inhibiciji rasta što upućuje na to da povećana koncentracije fitola ne utječe na povećanje antibakterijske aktivnosti. Rezultati ovog rada ukazuju da fitol, kao diterpenski alkohol, povećava aktivnost antibiotika kao što je polimiksin B djelujući na propustljivost vanjske membrane. Također je utvrđena i povećana aktivnost ceftazidina što ukazuje na moguću inhibiciju β-laktamaza na istraživanim ESBL sojevima Klebsiella pneumoniae.The aim of this study is to determine the spectrum of antimicrobial activity as well as the mechanisms of the action of diterpene alcohol, phytol, on a wide spectrum of Gram-positive and Gram-negative bacteria including species of Enterococcus faecalis, Bacillus cereus, Escherichia coli, Staphylococcus aureus, Aeromonas hydrophila, Klebsiella pneumoniae and Pseudomonas aeruginosa. Growth inhibition was determined against the examined species of bacteria by using the disc diffusion method. The lowest growth inhibition zone was recorded against the Staphylococcus aureus species in the range of 13.1 mm, while the highest sensitivity showed Pseudomonas aeruginosa species with a 29.9 mm growth zone inhibition. Antibiotic susceptibility with the addition of different phytic and non-phytic concentrations to Klebsiella pneumoniae ESBL producing strains was determined by using a disc diffusion method at Mueller-Hinton agar. A wide range of clinically relevant antibiotics were also used in the study. Fitol was added to antibiotic disks at different concentrations and growth inhibition was recorded at 10 μg/ml in combination with polymyxin B and ceftazidine. At higher investigated concentrations there was no significant difference in growth inhibition indicating that increased phytol concentrations did not influence the increase in antibacterial activity. The results of this study indicate that phytol, as a diterpene alcohol, increases the activity of antibiotics such as polymyxin B by acting on the permeability of the outer membrane. Ceftazidine activity was also demonstrated, indicating the possible inhibition of βlactamase in the investigated ESBL strains of Klebsiella pneumoniae