Substituent effects on the oxidative coupling of phenylpropanoids
Incorporation of phenylpropanoids into the plant polymer lignin occurs by a radical coupling mechanism catalysed by the enzyme peroxidase. Generation of the parent radical requires reduction of the enzyme intermediate compound I, which can be monitored by electron spin resonance (ESR) spectroscopy. It was found that the substituted cinnamyl alcohols were more easily oxidised than their corresponding acids and esters. For all phenylpropanoids, ease of oxidation increased with methoxyl substitution. The dehydrodimers, which represent the products of initial coupling, showed a similar trend with respect to methoxylation but their ability to reduce compound I was significantly lower than the monomers. This indciated that the polymer grows primarily as a result of the monomer reactivity, with polymerisation to high molecular weight restricted to dimethoxylated and possibly monomethoxylated compounds. Phenylpropanoids have many resonance structures and the distribution of the unpaired electron, determined by ESR and computational methods, was used to predict the mode of intermolecular coupling. Predictions were tested by preparation of synthetic polymers. The highest preponderance of C-C linkages was observed for the polymer derived from non-methoxylated units. Increasing methoxylation resulted in an increased proportion of C-O bonds, despite the stereochemical barrier imposed by these substituents. The polymer containing predominately C-C linkages had the most rigid structure, whereas, a greater amount of C-O bonds gave a more flexible and closely packed conformation. These results demonstrated that polymerisation, duplicated synthetically, was indeed controlled by the electronic structure of the monomers. The properties observed for the monomethoxylated cinnamic acid may explain the choice of this precursor for polymer cross-linking within the cell wall. Similarly, the role of the non-methoxylated cinnamate as a terminal unit and the kinetics of 2+2 cycloaddition reactions could be explained. The consequences for polymer formation following generic manipulation of lignin precursor pathways and the subsequent incorporation of cinnamyl aldehydes and polyhydroxylated phenylpropanoids were investigated.