Validation of a chromatographic method to dose ochratoxin A in green coffee

Green coffee and its derivatives, which include beverages, dietary supplements, additional foods, and nutraceuticals, are appreciated for their potent antioxidant properties. However, there is a significant risk of contamination of green coffee and its products by fungi and their mycotoxins, particularly ochratoxin A (OTA). This contamination can happen in traditional and organic green coffee during various stages, such as berry picking, crop storage, and transportation. Ochratoxin A (OTA) is known to have harmful effects on the kidneys (nephrotoxic), liver (hepatotoxic), and nervous system (neurotoxic) and can cause congenital disabilities (teratogenic) and cancer. This work validated a chromatographic method to measure OTA levels in green coffee following the UNI CEI EN ISO/IEC 17025: 2018. The analytical method's validation is essential to guarantee data traceability and avoid measurement errors. The experimental design involved three steps: extraction with polar solvents, elimination of interfering substances using an affinity column, toxins' separation by reversed-phase chromatography, and spectrophotometric identification of toxins. The validation process demonstrated that the method can recover OTA levels (76%) conformed with the performance criteria required for toxins by Regulation (EU) no. 519/2014, the regression coefficient (r-value = 1) and residual analyses proved the linearity of the calibration curve, the LOD, LOQ, and the measuring range confirmed that the test detects the minimum OTA levels permitted by the current legislation in coffee (3 µg/kg - 5 µg/kg). The precision (with a standard deviation of 0.0073) and accuracy (within ± 0.64 µg/kg) were considered statistically satisfactory. The method's results were trustworthy, with a confidence level of 95%, based on the uncertainties determined through the metrological approach. Therefore, the analytical method was considered sensitive, precise, accurate, and suitable for determining the ochratoxin concentrations reported by the regulations in force.