The objective of the study was to demonstrate the applicability of differential pulse polarography (DPP) technique of the ascorbic acid (AA) assay for the analysis of food samples with various matrices. The following validation parameters were determined: selectivity, linearity, precision, accuracy, limit of detection, and limit of quantification. The limits of detection and quantification were 0.17 and 0.5 mg ascorbic acid per 100 g food, respectively. The average recovery of added ascorbic acid from all matrices was 96.3–103.6%. The values of the coefficient of variation calculated on the basis of AA contents for food matrices were in the range 2–9.26% and Horrat values were 0.37–1.10. Ascorbic acid values of the samples obtained from the polarographic method were compared with those obtained from high-performance liquid chromatography with diode-array detection (HPLC-DAD) used as the reference method. The analytical parameters determined showed that the polarographic method was equivalent to the chromatographic method. Validation of the polarographic method of ascorbic acid assay indicates that it can be applied for the analysis of ascorbic acid concentration in food products that do not contain isoascorbic acid. This means that the method can be recommended for routine determinations.