A spectrophotometric assay for caffeoylquinic acids quantification in coffee using N_α-Boc-l-lysine

A spectrophotometric assay based on the reaction between caffeoylquinic acids (CQAs) and N_α-Boc-l-lysine (LYS) was studied, with the hypothesis that the resulting formation of green trihydroxy benzacridine (TBA) pigments, absorbing at 680 nm, correlates with total CQAs concentration. The reaction was effective on chlorogenic acid (5-CQA) and its isomers neochlorogenic (3-CQA) and cryptochlorogenic acid (4-CQA), and 3,5-di-caffeoylquinic acid (3,5-diCQA). Other caffeoyl derivatives as rosmarinic and chicoric acids, did not form the pigment. A protocol to quantify CQAs in coffee was then developed in 96-well microplate. The performance of the method was evaluated analysing green coffee samples. Exploiting a general calibration curve built with 5-CQA, the total content of CQAs was obtained after 60 min. A comparison with HPLC standard quantification of CQAs was performed. The system was validated on samples of green and roasted coffee, showing very good performance on green coffee, and a matrix effect on roasted coffee.