Mixtures of mono-, bi-, and tridecarboxylated betacyanins together with their corresponding neobetacyanins obtained from Beta vulgaris L. root juice as heating degradation products of betacyanins were identified by high-performance liquid chromatography with tandem mass spectrometry (LC-MS/MS) and diode-array (LC-DAD) detection. Two monodecarboxy-betacyanin pairs of diastereomers were detected after the decarboxylation in ethanolic and aqueous solutions. Generation of 17-decarboxy-betacyanins and 2-decarboxy-betacyanins was suggested, the latter so far never having been attributed to betacyanin thermal degradation products. Other main products of decarboxylation were 2,17-bidecarboxybetanin, its isoform, and 14,15-dehydrogenated (neobetacyanin) derivatives of all the decarboxylated betacyanins. The results of this research are crucial in determining betacyanin degradation mechanisms in juices or extracts of B. vulgaris L. roots and other products containing these pigments.