Author:

Abstract:

Amyloid protein fibrils (APF) can be derived from isolated food proteins by heating under acidic (e.g. pH 2) conditions. APF have been found to occur found foie gras. However, their presence in common food products remains to be investigated. APF are highly ordered nanofibers composed of cross β-strands, stacked perpendicularly along the fibril axis, which are stabilized by intermolecular hydrogen bonds. We hypothesized that boiling can induce the formation of amyloid protein fibrils in hen egg white (EW). For that, ovalbumin (OVA, ca. 54% of EW proteins) and EW were boiled for 15 min. A method was developed to extract fibrillar protein structures from heated OVA and EW, based on the fact that compact amyloid structures are less susceptible to chemical or enzymatic cleavage than amorphous protein aggregates. Firstly, a chemical extraction procedure using sodium dodecyl sulfate (SDS) and/or dithiothreitol (DTT) was evaluated. Low concentrations of SDS [≤ 0.3% (w/v)] induced APF formation in unheated OVA and EW, while high concentrations of SDS [> 0.5% (w/v)] disrupted APF derived from heated OVA and EW. The addition of 1.0% (w/w) DTT to the extraction medium enhanced both effects. Secondly, heated EW and OVA were subjected to an enzymatic treatment (37 °C, 48 h, 150 rpm). Trypsin solubilized both fibrillar and amorphous structures, while proteinase K solubilized mainly amorphous regions. Therefore, it was hypothesized that the pellet of heated EW formed after treatment with proteinase K contained mainly APF. This pellet was extracted with 0.01 M HCl (room temperature, 1 h, 150 rpm). Thioflavin T fluorescence, size exclusion chromatography, Fourier transform infrared spectroscopy measurements and transmission electron microscopy images indicated the presence of APF in hard-boiled OVA and EW. These results suggest that APF may be present in common food products containing eggs.