Sodium salicylate

The hydroxyl radical scavenging assay was performed according to the protocol proposed by Sudha et al. [23 ] and based on the previous work of Smirnoff and Cumbes [24 (link)]. Briefly, an aliquot (1 mL) of each homogenized cheese spread (CSExt, CSFos, and CS) was added to 2 mL of a reaction mixture containing 1 mL of 1.5 mM FeSO₄ (Merck, Darmstadt, Germany), 0.7 mL of 6 mM hydrogen peroxide (Merck, Darmstadt, Germany), and 0.3 mL of 20 mM of sodium salicylate (VWR, Leuven, Belgium). After incubation for 1 h at 37 °C, the absorbance of the hydroxylated salicylate complex was measured at 562 nm (indicated as A sample) in a UV-VIS spectrophotometer (Shimadzu, Barueri, SP, Brasil). Three replicates were performed. Using ascorbic acid as the standard (0–1 mg/mL), results were expressed as the equivalent concentration of ascorbic acid (mg ascorbic acid equiv/mL), being the hydroxyl radical scavenging percentage calculated using the following formula: $$\mathrm{Hydroxyl}-\mathrm{radical} \mathrm{Scavenging} \%=\left(1-\frac{\mathrm{A} \mathrm{sample}-\mathrm{A} \mathrm{blank}}{\mathrm{A} \mathrm{control}}\right)\times 100$$
For each sample, the absorbance of 2 mL reaction mixture plus 1 mL of deionized water was measured as the control (indicated as A control), whereas 2 mL of the reaction mixture with sodium salicylate replaced by water plus 1 mL of cheese spread was measured as the blank (indicated as A blank).

To analyse processing and assembly reactions, proteins were resolved through 12% Tris-glycine SDS-PAGE gels (mini-protean II; Bio-rad; 37.5:1 Acrylamide/Bis acrylamide), the gels were soaked in 1 M sodium salicylate (VWR) for 30 min, dried for one hour at 80 °C (DrygelSR, Hoefer, San Francisco, CA, USA), exposed to film at −80 °C overnight and developed using standard photographic reagents.