Structures, sensory activity, and dose/response functions of 2,5-diketopiperazines in roasted cocoa nibs (Theobroma...

The taste compounds inducing the blood-like, metallic bitter taste sensation reported recently for a dichloromethane extract prepared from roasted cocoa nibs were identified as a series of 25 diketopiperazines by means of HPLC degustation, LC-MS/MS, and independent synthesis. Among these 25 compounds, 13 cis-configured diketopiperazines, namely, cyclo(L-IIe-L-Phe), cyclo(L-Val-L-Leu), cyclo(L-Pro-L-Pro), cyclo(L-IIe-L-Pro), cyclo(L-Val-L-Tyr), cyclo(L-Ala-L-Tyr), cyclo(L-Phe-L-Ser), cyclo(L-Ala-L-IIe), cyclo(L-Leu-L-Phe), cyclo(L-Pro-L-Val), cyclo(L-Pro-L-Thr), cyclo(L-Pro-L-Tyr), and cyclo(L-Val-L-Val) were identified for the first time in cocoa. In addition, the taste recognition thresholds for the metallic as well as the bitter taste of the diketopiperazines were determined, and after quantitative analysis by using two diastereomeric diketopiperazines as the internal standards, the sensory impact of the diketopiperazines was evaluated on the basis of their dose-over-threshold (DoT) factors calculated as the ratio of the concentration and the threshold concentration of a compound. These data revealed DoT factors above 1.0 exclusively for cis-cyclo(L-Pro-L-Val), cis-cyclo(L-Val-L-Leu), cis-cyclo(L-Ala-L-Ile), cis-cyclo(L-Ala-L-Leu), and cis-cyclo(L-Ile-L-Pro), whereas all of the other diketopiperazines were present below their individual bitter taste threshold concentrations and should therefore not contribute to the cocoa taste. Because the DoT factors do not consider the nonlinear relationship between the concentration and gustatory response of an individual compound, we, for the first time, report on the recording of dose/response functions describing the human bitter taste perception of diketopiperazines more precisely.