J. A@. Food Chem. 1992, 40, 2337-2340 2987 Spectroabsorptiometric Investigations of Complexing Reactions of Polyhydroxylic Flavylium Compounds Dragan S. Veselinovi6,' Jelisaveta M. Baranac, Zoran D. ZujoviC, and Dragana S. Djordjevib Institute of Physical Chemistry, Faculty of Sciences, University of Belgrade, P.O. Box 550, W-11001 Belgrade, Yugoslavia The anhydrobasic form of delphinidin forms with Al(II1) at pH 5.85 a complex, Dp:Al = 3:1, with the relative equilibrium constant of 14.38 f 0.07, at room temperature and at ionic strength I = 0.2 mol/L. Complexationin acetate buffer has a positive effect on delphinidin stabilization,the decomposition rate constant of which is k = 0.052 min-l, at pH 5.85 and room temperature, and which decreases to k = 0.0025 min-l in the presence of Al(II1) because of complexation. INTRODUCTION It is well-known from Bayer's published papers (Bayer, 1958,1959; Bayer et al., 1960) that hydroxylated flavylium compounds, with OH groups in ortho positions in the B-ring, form complex compounds with some metals. Complexesthusly formed in vitro were expected to provide explanation for colorationof some fruits and flower petals in vivo. Within Bayer's investigation of complexation possibilities of different anthocyanidines and anthocyans (glycosidified anthocyanidines), he also paid close attention to the reaction of delphinidin (hexahydroxyflavylium compound) with aluminum. His investigationresulted in the formation of a ligand to the Al(II1) complex of 3:l for which he also proposed a possible structure but without calculating its stability constant. For this reason, by using electronic absorption spectra we made an attempt in our work to define the fist transformations of delphinidin molecules in solutions with different pH values and then to investigate the complexation reaction with aluminum. EXPERIMENTAL PROCEDURES The reagents used were Britton-Robinson (Britton, 1952) (I = 0.2 mol/L)and acetate (Dobos,1978)buffer solutions. Other chemicals used were delphinidin chloride (Fluka), &C13.6&0 p.a., "Zorka". Delphinidin chloride stock solutions (C = 10-3 mol/L)were prepared by diseolving precisely weighed quantities of substance. AlC13 solutionswere also C = lO+mol/L. Solutions were prepared by diluting 0.5 mL of delphinidin stock solution with different buffers up to 10 mL. Delphinidin concentration in all buffer solutions was constant, CD, = 5 X mol/L. Complex formation WBB investigated by using 0.5 mL of del- phinidin and different volumes of AlCls stock solutions and by filling up with buffer solution up to 10 mL. Both stock solution concentrations were C = 10-3 mol/L. Absorption spectra were recorded on a Pye Unicam SP8-100 spedrophotometer, with l-cmquartzcells, 1 min after preparation of the solution, at room temperature using the corresponding solution BB a reference. RESULTS As shown by delphinidin spectra (Figure 1) in Britton- Robinson buffer solutions at pH 2.0-10.0 and at ionic strength of 0.2 mol/L, an increasing pH value results in the change of both the position and the intensity of the basic absorption maximum (Table I). The maximum of the cationic form (A+) of delphinidin at pH 2.0 (Figure 1, curve 1) bathochromically shifts with increasing pH, pointingtothe change of cationicstructure to anhydrobasic (A) during the deprotonization process and finally to the A 0.9 0.8 0.7 0.6 0.5 0.4 0.: 0.2 0.' C NO 1 2 3 L 5 6 7 8 9 PH 2.00 3.00 4.00 5.00 6.00 7.00 8.00 9.00 10.00 Figure 1. Absorption spectra of delphinidin in Britton-Robinson buffer solutions. CD, = 10-5 mol/L. formation of anhydrobase (A) at pH 7.0, as illustrated by the given scheme. At further increase, up to pH 10.0, a new bathochromicshift indicates further deprotonization and formation of the anionic form of this chromophore (A-) (Figure 1, curve 9). The transformational sequence of delphinidinstructurewith changingpH made it possible first to determine the range of pH values characteristic of the formation of the anhydrobaseform of this chromophore (pH 5.0-6.0) and to determine, in accordance with the literature (Bayer, 1959), the optimum values for the formation of complex compounds. To be able to choose the optimum buffer system for complexation, measure- ments were also performed with delphinidin in acetate buffer solutions in the same pH region (pH 5.0-6.0). It is seen in Figure 2 that the percentage of the anhydrobase form is highest at pH 5.85 (the most intensive maximum), and this appears to be the most suitable medium for complexation reaction. Figure 3 shows the spectra of delphinidin and the aluminum complex in acetate buffer solutions at different pH values. The highest bathochro- Q 1992 American Chemlcal Soclety