Validation of analytical techniques for the quantification of NFCs in cocoa and roasted coffee: ensuring precision and reliability

The formation of neoformed compounds (NFCs) during coffee and cocoa roasting, such as acrylamide, furan, hydroxymethylfurfural, and furfuryl alcohol, poses both analytical and public health challenges, as these compounds simultaneously affect sensory attributes and the toxicological profile of foods. Exposure assessment depends not only on their detection but also on the metrological reliability of the analytical methods employed. This chapter describes and validates analytical methodologies based on selective extraction techniques, combined with liquid and gas chromatography, and coupled to UV and mass spectrometric detection. The methods were selected according to the physicochemical properties of the target analytes and the complexity of the food matrix. System suitability, selectivity, linearity, limits of detection and quantification, accuracy, precision, and interlaboratory reproducibility were systematically evaluated. The validated methods exhibited high linearity (R² ≥ 0.99), recoveries ranging from 80 to 120%, precision with %RSD < 16% for acrylamide and < 5% for FFA and HMF, and HorRat values between 0.3 and 1.3, confirming their reproducible performance. Overall, the rigorous validation demonstrates robust analytical performance and fitness for purpose, supporting interlaboratory comparability and evidence-based decision-making in quality control and risk assessment. Nevertheless, analytical reliability remains matrix-dependent and susceptible to extraction bias, matrix interferences, and ionization effects, potentially compromising the accuracy of exposure and risk estimates.