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Date sugar content considered being one of the most important fruit commercial characteristics. In a previous paper, the author introduced a novel computer-based method for date sugar content estimation in Tamr stage. This method depends on calculating the intensities of the red, green, and blue components of fruit color via image processing algorithm. In this paper, two date varieties were focused on in order to validate the concept. Lolo and Bomaan are among the most famous date varieties in the UAE. Fructose and Glucose content were determined for each variety using the HPLC. Sugar content was estimated in 43 samples of each variety and compared with the chromatography results. The results showed that, for the two varieties, glucose and fructose might be estimated using this technique with a minimum accuracy of about 86%. This paper focuses on the validation of this technique to evaluate its practicability.