Rate and Equilibrium Constants of Dehydration of Acylated Cyanidins
J. Agric. Food Chem., Vol. 47, No. 8, 1999 3453
color by hydration. This conclusion parallels that of
Dangles and Elhajji (1994), who examined the effects
of substituents attached to the flavylium nucleus.
In the anthocyanins studied here the substituents are
not directly attached to the flavylium nucleus but are
at the opposite end of a disaccharide chain from the
flavylium nucleus. The explanation of the electronic
effects of the acyl group substituents on the reactions
of the flavylium nucleus is not immediately obvious.
There is no direct pathway through the carbon frame-
work by which electronic effects from the acyl group can
exert an influence on the flavylium nucleus, and, as a
result, other explanations are needed. However, the acyl
groups in these molecules are physically near the
flavylium nucleus (at least part of the time) as shown
by the chemical shifts in the 1H NMR spectra of the
acylated versus the nonacylated anthocyanins (Dougall
et al., 1998; Gakh et al., 1998). This has been inter-
preted in terms of “intramolecular stacking” (Dangles
et al., 1993; Goto and Kondo, 1991) with the acyl group
near the flavylium nucleus in the stacked form, the acyl
group away from the flavylium nucleus in the unstacked
form, and these two forms in equilibrium with each
other (Dangles et al., 1993). This model of the mono-
acylated anthocyanin molecule is inadequate because
it always leaves one face of the flavylium ion available
to attack by water. We are developing models of monoa-
cylated anthocyanins using NMR and quenched molec-
ular dynamics to attempt to understand the effects of
the acyl groups on color retention and other properties
of the anthocyanins.
F igu r e 6. Relationship between log average rates of the
dehydration reactions of some cinnamoylated anthocyanin
hemiacetals and the electron-donating capacity of the substit-
uents on the acyl group. The circles identify anthocyanins
acylated with para-substituted cinnamic acids. The triangles
identify those acylated with meta- or meta- + para-substituted
cinnamic acids. The double asterisk indicates that the coef-
ficient was statistically significant (P < 0.01). 9 on the graph
refers to compound 9, etc.
um between quinonoid forms and the flavylium ion does
not change with changing electron density in the cin-
namoyl ring system.
The intercepts of the regression lines for log Kh, k1,
and the average of the Kavalues for the benzoylated
compounds are numerically larger than those for the
cinnamoylated compounds, whereas the intercepts for
the regressions of the log k2 values are very similar. This
shows that the benzoylated compounds have less fla-
vylium ion in equilibrium with the other species than
do the cinnamoylated compounds; that is, the benzoyl-
ated compounds retain less color under the influence of
pH than do the cinnamoylated compounds. This con-
firms the importance of the exocyclic double bond in the
color retention of the acylated anthocyanins.
ACKNOWLEDGMENT
We thank Dr. Elizabeth Howell, Biochemistry De-
partment, for access to the UV-vis spectrometer and
for help and guidance with the measurements.
LITERATURE CITED
Bailey, N. T. J . Regression analysis. In Statistical Methods in
Biology; English Universities Press: London, U.K., 1959;
Chapter 10, pp 91-99.
Brouillard, R. Chemical structure of anthocyanins. In Antho-
cyanins as Food Colors; Markakis, P., Ed.; Academic Press:
New York, 1982; Chapter 1, pp 1-40.
Dangles, O.; Elhajji, H. Synthesis of 3-methoxy- and 3-(â-D-
glucopyranosyloxy)flavylium ions. Influence of the flavylium
substitution pattern on the reactivity of anthocyanins in
aqueous solution. Helv. Chim. Acta 1994, 77, 1595-1610.
Dangles, O.; Saito, N.; Brouillard, R. Kinetic and thermody-
namic control of flavylium hydration in the pelargonidin-
cinnamic acid complexation. Origin of the extraordinary
flower color diversity in Pharbitis nil. J . Am. Chem. Soc.
1993, 115, 3125-3132.
Dougall, D. K.; Baker, D. C.; Gakh, E. G.; Redus, M. A.;
Whittemore, N. A. Anthocyanins from wild carrot suspen-
sion cultures acylated with supplied carboxylic acids. Car-
bohydr. Res. 1998, 310, 177-189.
Gakh, E. G.; Dougall, D. K.; Baker, D. C. Proton nuclear
magnetic resonance studies of monoacylated anthocyanins
from the wild carrot: Part 1. Inter- and intramolecular
interactions in solution. Phytochem. Anal. 1998, 9, 28-34.
Goto, T.; Kondo, T. Structure and molecular stacking of
anthocyaninssflower color variation. Angew. Chem., Int. Ed.
Engl. 1991, 30, 17-33.
J affe´, H. H. A reexamination of the Hammett equation. Chem.
Rev. 1953, 53, 191-261.
DISCUSSION
The anthocyanins acylated with benzoic acids display
a greater tendency to form hemiacetals than do the
anthocyanins acylated with the corresponding cinnamic
acids, which identifies the importance of the exocyclic
1
double bond in color retention. In their H NMR spectra,
anthocyanins with both types of acyl groups display an
upfield shift of the H-4 of the flavylium nucleus relative
to the nonacylated anthocyanins, but only the antho-
cyanins acylated with cinnamic acids display an upfield
shift of the H-8 resonance of the flavylium nucleus
(Dougall et al., 1998). These observations suggest that
the two types of acyl groups interact differently with
the flavylium nucleus.
The rates and the equilibrium constant for the hydra-
tion reaction and thus color retention by these antho-
cyanins are altered by the electron-donating capacity
of substituents on the acyl group and by at least one
additional factor associated with meta- and ortho-
substitution on the acyl group. The latter may be steric
effects.
For both the benzoylated and the cinnamoylated
anthocyanins, the higher the electron density on the
aromatic ring, the smaller the Kh for that compound and
the smaller the tendency of that compound to lose its