Zinc sulfate heptahydrate

For each nutraceutical, with the exception of quercetin, a stock solution was prepared. A serial dilution was then done in culture medium to attain desired working concentrations for treatment of cell layers. Prior to supplementation, the media was filter sterilized with a 0.2 μm sterile syringe (Corning). For zinc, a stock solution (100 mM) was made from zinc sulfate heptahydrate (Fisher Chemical) in deionized distilled water. A butyrate stock solution (400 mM) was made from sodium butyrate (Sigma-Aldrich) in deionized distilled water. A berberine (Sigma-Aldrich) stock solution (2.7 mM) was also prepared in deionized distilled water, but was made each day at the time of use. In the case of indole (Sigma-Aldrich), a 400 mM stock solution was prepared in absolute ethanol. With quercetin (Sigma-Aldrich), dry chemical was added directly to complete culture medium to make up a working concentration (400 μM) that was applied directly to cells. Solubilization of quercetin in medium at 400 μM required warming medium to 38°C for 40 minutes with constant stirring. Lower concentrations were prepared simply by serial dilution in complete medium. Proper solvent controls were performed in all experiments.

In total, 100 mL DI water was pre-heated as the solvent to dissolve zinc sulfate heptahydrate (ZnSO₄·7H₂O, Fisher Chemical), copper (II) sulfate pentahydrate (Fisher Chemical), and boric acid (Powder/Certified ACS, Fisher Chemical) under continuous stirring for 20 min until a transparent solution was obtained (noted as Solution C). The Zn-Cu alloys were deposited on Zn substrates using the two-electrode setup in Solution C.

Sodium carboxymethyl cellulose (Mw = 250
kDa; DS= 0.9), n-octylamine, and n-hydroxysuccinimide (NHS) were purchased from Acros Organics (Geel,
Belgium). Acetic acid, pyrene, and ethylenediaminetetraAcetic acid
(EDTA) were purchased from Sigma-Aldrich (Steinheim, Germany). 1-(3-(Dimethylamino)propyl)-3-ethylcarbodiimide
hydrochloride (EDC) was purchased from Alfa Aesar (Lancashire, United
Kingdom). Dextran standards were purchased from PSS Polymer (Mainz,
Germany). Zinc sulfate heptahydrate was purchased from Fisher chemicals
(Dublin, Ireland). Chitosan (Mw = 890 kDa; DD ≥ 90%) was purchased
from Glentham (Corsham, United Kingdom). Ethanol and ethanol absolute
were purchased from Gadot-group (Netanya, Israel). Water (HPLC grade)
was purchased from Bio Lab (Jerusalem, Israel). Deionized water (DW)
was obtained by mechanically filtering them through a Treion TS1173
column. Deuterated solvent for NMR analysis (D₂O) was purchased
from Armar Chemicals (Döttingen, Switzerland). All regents
and solvents were used without further purification.