Lysozyme from hen egg white

To determine the lysozyme activity, Anderson and Siwicki (34 ) method reported by Ullah et al. (30 (link)) was followed. Briefly, serum (100 μL) was taken with the help of a micropipette in a fresh test tube and mixed with 900 μl of 750 μg mL⁻¹Micrococcus lysodeikticus (Sigma, USA) suspended in saline phosphate buffer (pH 6.2) solution. Bacteria were thoroughly mixed, and the rate of absorbance change was observed by using a spectrophotometer set at 540 nm. The reading was noted after 1 min interval for 10 min. Lysozyme activity was measured by using lysozyme from hen egg white (Sigma-Aldrich) as a standard.

α,α-Trehalose (in
dihydrated form), sucrose (in anhydrous
form), lysozyme from hen egg white, and myoglobin from equine heart
were all purchased from Sigma-Aldrich and used without any further
purification.

A solution of lysozyme was prepared by dissolving lysozyme from hen egg white (80 mg, Sigma-Aldrich) in deionized water (4 mL). The solution pH was adjusted to 2.0 using aqueous HCl (2 M). After filtration using a 0.22 μm syringe-driven filter, the solution was heated at 90 °C for 20 h with stirring at 300 rpm. Large aggregates were removed by centrifugation at 2290 × g for 3 min. Nanofibril concentration was measured using Bio-rad protein assay kit with BSA as a standard.

Natural zeolite of clinoptilolite type (CZ) consisted of ~ 85% CZ was originated from Noyemberyan, Armenia. Synthetic zeolite Y microparticles (particle size ~ 30 µm, CAS No: 1318-02-1), magnetic iron oxide Fe₃O₄ (IO) nanoparticles (particle size 6.5 ± 3 nm, CAS No: 1317-61-9), Insulin, human recombinant, expressed in yeast (E.C. 234-279-7, I2643, ~ 24 IU/mg protein), Glycine (G7126), Lysozyme from hen egg white (E.C. number: 3.2.1.17, lyophilized powder, L 6876, ∼ 50,000 unit/mg protein), NaCl, Thioflavin T (ThT, T3516) were obtained from Sigma-Aldrich Corporation (St. Louis, MO; USA). Photodynamic dyes hypericin (HYP), and temperature sensitive dye rhodamine B (RhB) from Sigma-Aldrich were of analytical grade and used without further purification. Sulfonated aluminum phthalocyanine (SAP), a second-generation synthetic photosensitizer with a light absorption maximum at 675 nm was purchased from NIOPIC, Russia. All other chemicals were of analytical grade.

SWNTs (CG300L35) for this research were purchased from CHASM (Norman, OK, USA) and used as received. Lysozyme from hen egg white, double-stranded DNA sodium salt from salmon testes, and pluronic F108 were purchased from Sigma-Aldrich (St. Louis, MO, USA). Sodium dodecyl sulfate (SDS) was purchased from Fisher Scientific (Hampton, NH, USA). Finally, BD Bacto (Franklin Lakes, NJ, USA) TSB mixture was prepared according to vendor instructions.
The dispersants and SWNT were dispersed using initial concentrations and sonication protocols from the literature (Table 3). After sonication, each sample was centrifuged at 17,000× g for 3 h to remove large SWNT bundles and aggregates. The supernatant was collected and used for the experiments. For normalized concentrations, the SWNT dispersions were diluted with their respective dispersant controls to a corrected concentration of 0.227 mg/mL.

2-Mercaptoethanol, Acetonitrile, Ampicillin sodium salt, Benzydamine hydrochloride, Benzydamine N-oxide hydrogen maleate, Bradford Reagent, d(+)-Glucose, Flavin adenine dinucleotide disodium salt hydrate (FAD), HisTrap HP columns, IGEPAL CA-630, Imidazole, Kanamycin, Nickel(II) sulfate hexahydrate, Lysozyme from hen egg white, Phenylmethylsulfonyl fluoride, and potassium phosphate were purchased from Sigma-Aldrich. NADPH tetra(cyclohexammoninum) and β-NADP sodium salt were purchased from Carbosynth. Isopropyl-beta-d-thiogalactopyranoside (IPTG) was purchased from BIOSYNTH. TRIS Base was purchased from Fisher Molecular Biology. Yeast Extract and Tryptone were purchased from Fisher Bioreagents. PageRuler Unstained Protein Ladder was purchased from Thermo Fisher Scientific Baltics UAB.

Ch was provided by Join Therapeutics S.r.l. (Padova, Italy). Sodium Hyaluronate were purchased from HTL Biotechnology. Sodium azide, sodium nitrate, sodium dihydrogen phosphate monohydrate, sodium hydrogen phosphate dihydrate, trimethylsilyl-3-propionic acid (TSP), Hyaluronidase from bovine testes (400–1000 u/mg) and Lysozyme from hen egg white (93,300 u/mg) were purchased from SigmaAldrich (Milan, Italy). Deionized water (conductivity less than 0.1 μS) was prepared with an inverse osmosis system (Culligan, Milan, Italy). PolyCAL TM Pullulan std-57k (Malvern Instruments LtD, Malvern, United Kingdom). The reagent grades were ≥ 98%.