title: Evaluation of uncertainty of measurement from method validation data: an application to the simultaneous determination of retinol and alpha-tocopherol in human serum by HPLC.
abstract: An isocratic RP-HPLC method for the determination of retinol and alpha-tocopherol in serum, with fluorescence and UV/VIS detection, respectively, was developed and validated according to international guidelines. Detection (retinol 0.015 mg/L, alpha-tocopherol 0.29 mg/L) and quantification (retinol 0.05 mg/L, alpha-tocopherol 0.95 mg/L) limits were determined. Repeatability was <3.3% and <2.9% and intermediate precision was <4.6% and <3.2%, for retinol and alpha-tocopherol, respectively. Certified reference materials were utilised to assess bias and guarantee traceability to SI units. Expanded uncertainties (retinol 8.9%; alpha-tocopherol 7.9%), estimated according to the EURACHEM/CITAC guide from method validation data, satisfied fit-for-purpose requirements based on biological variability.
authors.affiliation: Department of Food Safety and Veterinary Public Health, Istituto Superiore di Sanità, Roma, Italy. antonella.semeraro@iss.it
authors.forename: Antonella
authors.initials: A
authors.lastname: Semeraro
authors.forename: Ilaria
authors.initials: I
authors.lastname: Altieri
authors.forename: Marina
authors.initials: M
authors.lastname: Patriarca
authors.forename: Antonio
authors.initials: A
authors.lastname: Menditto