Fast, Direct Dihydrouracil Quantitation in Human Saliva: Method Development, Validation, and Application

Background. Salivary metabolomics is garnering increasing attention in the health field because of easy, minimally invasive saliva sampling. Dihydrouracil (DHU) is a metabolite of pyrimidine metabolism present in urine, plasma, and saliva and of fluoropyrimidines-based chemotherapeutics. Its fast quantification would help in the identification of patients with higher risk of fluoropyrimidine-induced toxicity and inborn errors of pyrimidine metabolism. Few studies consider DHU as the main salivary metabolite, but reports of its concentration levels in saliva are scarce. We propose the direct determination of DHU in saliva by reversed-phase high-performance liquid chromatography (RP-HPLC-UV detector) as a simple, rapid procedure for non-invasive screening. Methods. The method used was validated and applied to 176 saliva samples collected from 21 nominally healthy volunteers and 4 saliva samples from metastatic colorectal cancer patients before and after receiving 5-fluorouracil chemotherapy. Results. DHU levels in all samples analyzed were in the μmol L^-1 range or below proving that DHU is not the main metabolite in saliva and confirming the results found in the literature with LC-MS/MS instrumentation. Any increase of DHU due to metabolism dysfunctions can be suggestive of disease and easily monitored in saliva using common, low-cost instrumentation available also for population screening.