The aim of the current study was to thoroughly investigate the structural changes of anthocyanins at pH 3.5 in purified fractions from black carrot, elderberry and strawberry heated over 6 h at 95°C. Degradation products were monitored by HPLC-DAD-MS3 to elucidate the prevailing degradation pathways. In addition, alterations of color and antioxidant properties observed upon heating were scrutinized. Most interestingly, the degradation pathways at pH 3.5 were found to differ from those at pH 1. Among others, chalcone glycosides were detected at 320 nm in heat-treated elderberry and strawberry pigment isolates, and opening of the pyrylium ring initiated anthocyanin degradation. In the case of acylated anthocyanins, acyl-glycoside moieties were split off from the flavylium backbone, first. Finally, for all pigment isolates, phenolic acids and phloroglucinaldehyde were the terminal degradation products as remainders of the B- and A-ring, respectively. Maximum and minimum antioxidant stabilizing capacities were found in black carrot and strawberry, respectively, which was explained by the high degree of acylation in the former. After heating, decline of trolox equivalent antioxidant capacity (TEAC) was observed in all samples, which was attributed to both anthocyanins and their colorless degradation products following thermal exposure. As deduced from the ratio of TEAC value and anthocyanin content, the loss of anthocyanin bioactivity could not be compensated by the antioxidant capacity of newly formed colorless phenolics upon heating.