An analytical procedure to determine major sugars and organic acids, including vitamin C, in fruits was developed using a C18 Sep-Pak cleanup process and an ion exclusion HPLC column. Dual UV monitoring and refractive index were performed for detection. To attain optimal separation and quantitation, 0.0085N H2SO4 was used as the mobile phase and the column temperature was maintained at 23 °C. This procedure was compared to others for the individual quantitation of sugars, organic acids, and vitamin C. Recovery and reproducibility of this analytical procedure were quite acceptable for strawberry and four other common fruits, allowing the analysis of all the components using a single-injection HPLC analysis in <22 min. Keywords: Simultaneous analysis; high-performance liquid chromatography; sugars; organic acids; vitamin C; strawberry; fruits

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