Formation of di-Tyrosine in pasteurized milk during shelf storage

Formation of the protein crosslink di-Tyrosine was studied in PET-bottled pasteurized milk exposed to fluorescent light in a commercial display cabinet. An HPLC method with fluorescence detection was developed and intra-laboratory validated using pure di-Tyrosine synthesized on purpose. Di-Tyrosine was detected after 1-day lightening and increased up to 7 days, reaching around 250 and 320 mu g/g protein in whole and partly skimmed milk, respectively. Afterward, a progressive decrease occurred. By transmission electron microscopy with specific immune gold labelling, presence of di-Tyrosine was observed for the first time on the surface of casein micelles of lightened milk. The crosslink formation, however, did not bring to protein aggregation phenomena detectable by laser light scattering measurements. Exposure to light also induced degradation of riboflavin and decrease of yellowness index. Di-Tyrosine proved to be a suitable indicator to evaluate the progress of protein oxidation in pasteurized milk during storage on the market.