Antioxidant studies by hydrodynamic voltammetry and DFT, quantitative analyses by HPLC-DAD of clovamide, a natural phenolic compound found in Theobroma Cacao L. beans

Clovamide (trans-clovamide, (2S)-3-(3,4-dihydroxyphenyl)-2-[[(E)-3-(3,4-dihydroxyphenyl)prop-2-enoyl]amino]propanoic acid) is a naturally occurring caffeoyl conjugate and a potent antioxidant found in the phenolic fraction of Theobroma Cacao L. beans. This work quantified clovamide content in single-origin cocoa beans at different production stages (raw, roasted, and winnowed side and end products) by high-performance liquid chromatography with diode array detector (HPLC-DAD). We analyzed the antioxidant activities of clovamide and these extracts by measuring their superoxide radical scavenging capabilities in a Rotating Ring-Disk Electrode (RRDE) electrochemical system against in-situ generated superoxide radical. Our studies concluded a positive correlation between clovamide concentration and the overall antioxidant activities of beans, with the roasting step showing a reduction effect on both. The subsequent refining steps recover the clovamide concentration. Antioxidant studies on clovamide alone by RRDE and density functional theory (DFT) studies led to the conclusion that it is a powerful oxygen radical scavenger, partially contributed by its molecular catechol moieties.