Valorização das procianidinas da semente de açaí (Euterpe oleracea Mart.): fracionamento, caracterização e conversão enzimática por oxidases

Abstract

The significant growth of the açaí (Euterpe oleracea Mart.) production chain has increased its economic and social relevance, but it has also intensified environmental challenges associated with seed generation, an agro-industrial residue that represents approximately 85% of the fruit mass. Considering the potential of this biomass, this thesis aimed to investigate integrated strategies for the valorization of açaí seed extract as a source of procyanidins, with emphasis on chemical characterization, fractionation of procyanidins according to the degree of polymerization, and enzymatic conversion of B-type into A-type procyanidins using commercial oxidases. Initially, the extract was fractionated by medium-pressure liquid chromatography using Sephadex LH-20 resin, resulting in six fractions characterized by HPLC-DAD and HPLC-ESI-FT-ICR-MS. Fractions 1, 2, and 3 were mainly composed of sugars, fatty acids, and lower-mass flavonoids, respectively. Fraction 4, composed of procyanidins and other flavonoids, showed a mean degree of polymerization (mDP) of 2.6 and the highest antioxidant activity among all fractions, indicating that the structural distribution of procyanidins and the presence of other phenolic compounds may influence bioactivity. Fractions 5 and 6, composed exclusively of oligomeric and polymeric procyanidins, showed mDP values of 5.7 and 12.1, respectively, jointly corresponding to more than 85% of the extract mass and more than 70% of total antioxidant activity. Additionally, a selective and linear chromatographic method was adapted for the quantification of procyanidins A1 and B1, enabling direct calculation of conversion rates between these structures. Using this method, a screening was conducted with four commercial oxidases, including two laccases, one peroxidase, and one polyphenol oxidase (PPO). The T. versicolor laccase and PPO showed initial conversion rates of 54% and 68%, respectively, and were selected for subsequent stages. After evaluating pH, temperature, and stability, a 2² central composite rotational design (CCRD) was employed to optimize conditions, with experimental validation showing less than 1% deviation from predicted values. Kinetic studies in a model system allowed identification of optimal conditions of 48 °C, pH 4.3, and 90 minutes for laccase, and 32 °C, pH 6.6, and 60 minutes for PPO, achieving conversions of 71% and 86%, respectively, with by-product formation limited to 5.2% and 2.6% and efficiencies of 93% and 97%. Finally, optimized conditions were applied to real substrates, including crude extract, fraction 4, and fraction 6. Conversion rates of procyanidin B1 to A1 identified for the extract and fraction 4 were 63% and 72% for laccase, and 79% and 85% for PPO, respectively. Mass spectrometry analysis showed increased intensity of ions corresponding to A-type oligomers and polymers, along with a concomitant reduction in B-type procyanidins. Overall, analyses with the extract and fractions demonstrated that conversion efficiency is influenced by matrix complexity and the degree of polymerization of procyanidins. Thus, the results advance the understanding of extract composition, the structure–antioxidant activity relationship of açaí seed procyanidins, and support the use of biocatalytic processes as a potential route for their valorization.