Testing the Durability of Polymyxin B Immobilization on a Polymer Showing Antimicrobial Activity: A Novel Approach with the Ion-Step Method

Abstract

A polymyxin B (PMB) modified copolymer of ethyl acrylate and 2hydroxyethyl methacrylate is reported that shows antibacterial activity towards E. coli. The polymer is designed for biosensor interfaces to provide protection against microbial contamination. However, leaching of a biomolecule from its solid support is a well-known problem in immobilization chemistry. It has been proposed that antimicrobial activity results from release of the antibiotic from the polymer matrix, but in the work reported here, bactericidal action was seen for tresyl activated polymers even when no polymyxin B could be detected in the supernatant and where covalent attachment of PMB was indicated. Further evidence for the in situ action of PMB on the polymer was provided using a novel method of investigation, subjecting the polymer, deposited on an Ion-Sensitive Field-Effect Transistor (ISFET) to a step change in ion-concentration (Ion-step). With the ion-step method activation of the polymer and the grafting of Polymyxin-B were followed. A comparison of absorbed PMB on a nonactivated polymer and PMB on the tresyl activated polymer suggested that if leaching of PMB from the latter occurred, it was below the detection limit and well below the minimum inhibitory concentration (MIC) for antimicrobial action. Bactericidal action was thus attributed to PMB attached to the polymer.

A review of immobilized antimicrobial agents and methods for testing

“…This led them to conclude that the elution in solution would be much less than the MIC for PMB. 58 They then determined the antimicrobial efficacy using an optical density measured growth curve lag between inoculated media that was or was not exposed to PMB immobilized glass. The bacteria in the media exposed to PMB glass had a delayed growth curve.…”

“…8,9 A wide range of antimicrobial agents have been immobilized, including small molecules (e.g., quaternary ammonium silanes), [10][11][12][13][14][15][16][17] quaternary ammonium polymers, [18][19][20][21][22][23][24][25][26][27][28][29][30][31][32][33][34][35] polyamines, [36][37][38][39][40][41][42] chitosan, [43][44][45][46][47][48][49] enzymes, [50][51][52][53][54][55] peptides, and peptide mimetics. [56][57][58]…”

Layered Double Hydroxides as the Unique Product of Target Ionic Construction for Energy, Chemical, Foods, Cosmetics, Medicine and Ecology Applications

“…One of the methods for regulating the pharmacological activity of such drugs is their immobilization on various carriers, which can increase the stability of the molecule, protect it from the effects of certain factors (for example, gastric juice when administered orally), and also provide controlled release into the environment, which allows prolonging the effect. Thus, many examples of successful immobilization of antibiotics, [202,203] enzymes, [204,205] peptides, [206,207] halogen-active, [208][209][210] and peroxide [211] preparations on polymeric carriers of various nature have been described. Low toxicity, large surface area, sorption properties, and the possibility of diffusion of the intercalated anion make SDGs promising containers for various microbiocides instead of hard-to-produce polymer bases.…”

Non-leaching surfaces capable of killing microorganisms on contact

“…The mode of action of antimicrobial peptides includes formation of pores in the cell membrane resulting in the disruption of the membrane potential with eventual lysis of the cell. 67,68 The immobilization of polymyxin B, a cyclic polycationic peptide with a molecular weight of $1,200 on an alkyl acrylate polymer has been reported and the conjugate showed antibacterial action against E. coli. The authors demonstrated that the bioactivity was indeed due to the immobilized AMP and not AMP leached from the polymer.…”

“…The authors demonstrated that the bioactivity was indeed due to the immobilized AMP and not AMP leached from the polymer. 68 A recent study has reported the immobilization of cathelin LL37, a cryptic antimicrobial peptide obtained by enzymic cleavage of its precursor cathelicidin which is stored in granulocytes. 69 The peptide was covalently immobilized onto a titanium metal surface by means of silanization.…”