Masters Theses

Marc Redus

Date of Award

12-1996

Degree Type

Thesis

Degree Name

Master of Science

Major

Life Sciences

Major Professor

Donald K. Dougall

Committee Members

Elizabeth Howell, David Baker, Beth Mullin

Abstract

In this study, a number of non-natural anthocyanins were biosynthesized by the addition of various organic acids to suspension cultures of wild carrot. This resulted in the incorporation of the organic acid residue in a specific place on the anthocyanin molecule, and the resulting novel anthocyanin was purified and characterized. The color of anthocyanins generally decreases with increasing pH due to the conversion of the flavylium ion into a hemiacetal. In addition, the flavylium ion can be converted into blue quinonoid forms at increased PH. Because these reactions result in loss of or changed color of the anythocyanins, their rate constants and equilibrium constants are a measure of color retention of the anthocyanin; if the factors which determine this were understood, this could result in the eventual marketing of anthocyanin preparations with increased color retention. For each anthocyanin, Kh, the equilibrium constant for the (colorless) hemiacetal formation, k1 and k2, the forward and reverse rate constants for this reaction respectively, and Ka, the equilibrium constant for formation of the quinonoid forms, were measured.

Anthocyanins with no acyl group had the largest Kh, smallest k2, and a k1 which was among the largest in the series of 21 compounds measured, and thus retained the least amount of color.

For anthocyanins acylated with benzoic acids, the log Kh, log k1, and log k2 were linearly related to the electronegativity of the substituents showing that the effects on these constants were linearly related to the electron density of the aromatic system. Log Ka was not linearly related to the electron donating capacity of the substituents, so another factor was affecting formation of quinonoid forms.

The Kh and k1 was larger in the benzoyl compounds than in the cinnamoyl series, and k2 was smaller, indicating that benzoylated anthocyanins retained less color than did the cinnamoylated ones.

For the anthocyanins acylated with cinnamic acids, a plot of log Kh and log k1 vs. electronegativity of the substituents showed that these compounds were scattered about a straight line. This suggests that both the electron density in the acyl aromatic ring and another factor were both important in determining color retention by a cinnamoylated anthocyanin.

The log Ka and log k2 in the 4-substituted cinnamoyl series were linearly related to the electronegativity of the substituents on the acyl group, indicating that electron density in the acyl aromatic ring was the primary factor controlling formation of the quinonoid form as well as the reverse reaction from hemiacetal to flavylium ion in these compounds.

A comparison of the compounds acylated with 4- hydroxycinnamic acid, 4-hydroxybenzoic acid, and 3-(4- hydroxyphenyl) propionic acid shows that the 2-carbon chain in acyl groups, as well as the presence of the double bond in cinnamic acids, affected the Kh's and k2's significantly. Removal of the carbon chain also increased k1, but removal of the double bond did not have as much affect.

The k2's appear to be a major factor that influences stability. Rather than showing an increase in k1 as stability decreases, many of the least stable compounds instead showed a decrease in k2.

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