Glycerin

Polyphenylsulfone (Radel^® R-5000, PPSU, M_w = 50,000 g∙mol⁻¹) was purchased from Solvay Advanced Polymer (Beveren, Belgium), N-methyl-2-pyrrolidinone (NMP, 99%) and glycerin were purchased from Acros Organics (Geel, Belgium), silica nanoparticles (SN) were synthesized based on the Stöber method described in detail elsewhere [75 (link)]. CO₂ (99.99 mol.%) and CH₄ (99.999 mol.%) gases were provided by AGA (Linde group, Vilnius, Lithuania).

Polyvinyl alcohol (PVA), pellets, (average molecular weight M_w = 12.4 × 10⁴ g/mol, degree of hydrolysis DH = 99 ÷ 100%), and glycerin (99.6%) were purchased from Acros Organics (Geel, Belgium). Tert-butyl acrylate (t-BA), 98%, containing 10–20 ppm monomethyl ether hydroquinone as an inhibitor; divinylbenzene (DVB), technical grade 80%, stabilized with the inhibitor monomethyl ether hydroquinone; 3-(triethoxysilyl) propionitrile, 97% (TESPN); aluminum oxide (Al₂O₃); tetraethylorthosilicate, 99% (TEOS); N,N′-dicyclohexylcarbodiimide, 99% (DCC); branched polyethyleneimine (average M_w~25,000 by LS, average M_n~10,000 by GPC); and anhydrous N,N-Dimethylformamide, 99.8% (DMF), were purchased from Sigma-Aldrich (Merck, KGaA, Darmstadt, Germany). All products containing inhibitors were first purified through an Al₂O₃ column prior to their usage.
Pure copper chloride (II) dihydrate p.a. (99.0% CuCl₂∙2H₂O) was bought from ChemPUR Feinchemikalien und Forschung GmbH (Karlsruhe, Germany); hydrochloric acid (HCl), ≥37%, was purchased from Fluka (Honeywell Specialty Chemicals, Seelze, Germany); and ethanol absolute, 99.3% (EtOH), was bought from Chemical Company (Iasi, Romania). An ammonium hydroxide (NH₄OH) solution (28–30%) was purchased from Sigma-Aldrich (Merck KGaA, Darmstadt, Germany) EMSURE^® ACS, Reag. Ph Eur. All aqueous solutions were prepared with freshly distilled water.

A total of eight COL-R A. baumannii and eight COL-R E. coli non-duplicate isolates were isolated from the First Affiliated Hospital of Wenzhou Medical University during 2012–2019. All isolates were cultured on a Columbia blood agar plate (BAP). Strains were identified using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS, BioMerieux, France) and stored at −80℃ in Luria Bertani (LB) broth medium supplemented with 30% glycerin (Thermo Fisher Scientific, Shanghai, China.).

Bacterial cell pellets were adjusted to equal amounts by adding 1× SDS buffer consisting of 62.6 mM Tris–HCl, 2% sodium dodecyl sulfate, 0.01% bromophenol blue, 5% glycerin and reducing agent (Thermo Fisher Scientific), while the supernatants were mixed with 4× SDS buffer. Prior to the separation by SDS–PAGE on 10% polyacrylamide gels, the protein samples were boiled for 10 min at 94°C (Moese et al., 2001 (link)). The separated proteins were then analyzed by staining with Coomassie Brilliant Blue R-250 (Bio-Rad, Munich/Germany). For Western Blotting, the proteins were blotted onto PVDF membranes (Immobilon-P, Merck Millipore, Darmstadt/Germany) and blocked in TBS-T buffer (pH 7.4, 0.2 M Tris, 1.4 M sodium chloride and 1% Tween-20) containing 5% milk powder for 1 h at room temperature or at 4°C overnight (Boehm et al., 2011 (link)). After addition of the primary antibodies for 2 h at room temperature, horseradish peroxidase-conjugates anti-rabbit polyvalent sheep immunoglobulin was applied as secondary antibody (Zhang et al., 2015 (link)). Antibody detection was performed using 1.41 mM luminol in 0.1 Tris–HCl (pH 6.8) supplemented with 0.61 mM p-coumaric acid solved in DMSO and 0.02% hydrogen peroxide. Unless otherwise indicated, all chemicals were obtained from Carl Roth (Karlsruhe/Germany).