Enzyme-catalyzed oligomerization of apigenin using laccases and horseradish peroxidase in batch and microreactor systems

Apigenin is a naturally occurring flavonoid widely recognized for its antioxidant, antimicrobial, anti-inflammatory, and anticancer properties. Recent studies have shown that oligomerization and oxidative coupling of flavonoids may significantly improve their physicochemical stability and biological activity compared to the parent compounds. Enzyme-catalyzed oligomerization has emerged as a promising green synthetic approach due to its mild reaction conditions, high selectivity, and reduced environmental impact. In this study, enzymatic dimerization and oligomerization of apigenin were investigated using oxidative enzymes, including laccases from Aspergillus sp. and Trametes versicolor as well as horseradish peroxidase (HRP). Reactions were performed in both a conventional batch reactor and a tubular microreactor to evaluate the influence of reactor configuration on conversion, product formation, and operational enzyme stability. Particular emphasis was placed on comparing the catalytic activity and selectivity of different oxidative enzyme systems towards apigenin coupling reactions under various reaction conditions, including pH, temperature, oxidant concentration, and solvent composition. The obtained results demonstrated successful enzymatic transformation of apigenin in all investigated systems, with noticeable differences in catalytic efficiency and product distribution depending on enzyme origin and reactor type. Microreactor systems enabled improved process control, enhanced mass transfer, shorter residence times, and more stable operating conditions compared to batch processing. Formation of oligomeric apigenin structures was confirmed using Fourier-transform infrared spectroscopy (FT-IR), UV-Vis spectroscopy, high-performance liquid chromatography with diode-array detection (HPLC-DAD), and nuclear magnetic resonance spectroscopy (NMR).