Lipase-catalyzed synthesis of phloridzin esters

Abstract

Phloridzin is member of chemical class of dihydrochalcones, phenylpropanoids with structure similar to chalcones, immediate flavonoid precursors, hence it is often classified as flavonoid glucoside [1,2]. It is usually extracted from Malus species since it is very abundant in young apple leaves and twigs. Due to its phenolic structure, phloridzin has significant antioxidant activity and anti-UV properties, which makes it interesting for application in food and cosmetics. Major limitation to wider application of phloridzin is its low solubility in hydrophobic environment, which can be circumvented by synthesis of physiologically active compounds derivatives by acylation of phloridzin. Synthesis of acyl esters can be catalyzed by inorganic catalysts or enzymes, but chemical esterification is not regioselective and results with unwanted functionalization of phenolic hydroxyl groups responsible for antioxidative properties. Therefore, in our study enzymatic esterification of phloridzin was performed using immobilized lipase from Candida antarctica (Novozyme® 435). Several organic solvents were tested and acetonitrile was proved to be the most adequate medium for this reaction. Different acyl-donors were used with respect to chain length and saturation level. Potential physiological activity of obtained esters was evaluated by determination of their antioxidant activity using DPPH assay, so acyl donors were compared with respect to both, product yields and antioxidant activity. After comparison of results of preliminary study, phloridzyl oleate was selected as derivative with the best prospects and it used in further experimental series for optimization of key experimental factors. Response surface methodology was applied as statistical tool for optimization of product concentration (in mM) as output and analyzed factors were: reaction time, temperature, enzyme concentration, substrate molar ratio, and phloridzin concentration. After statistical analysis each of examined experimental factors was found significant and second order regression model was obtained. It was established that highest product concentration can be expected at 68 oC, with 0.17 M of phloridzin, 2,5 % (w/v) of enzyme, 19-fold molar excess of oleic acid after 7 days of reaction.