The aim of this study was to examine the impact of cultivar and spear color on the composition of polyphenols in asparagus spears (Asparagus officinalis). The five genotypes (Schwetzinger Meisterschuss, Huchel’s Alpha, Gijnlim, Grolim and Eposs) and three growing conditions of asparagus spears (Asparagus officinalis) were investigated. The polyphenols were determined by applying the HPLC-DAD system. The obtained results were subjected to the principal component analysis. Among the analyzed asparagus samples cv. Grolim contained the highest amounts of phenolic acids and flavonols. The varied quantitative and qualitative composition of polyphenolics resulted most probably from changes occurring during vegetation, such as a lack of access to light in the case of white asparagus and limited access to light in purple asparagus. The scavenging activity on DPPH radicals by asparagus extract is dependent on the variety and color and was the greatest for green asparagus samples. Similar green extracts scavenged ABTS radicals to the highest degree. Results of this study suggested that asparagus may constitute a good source of natural antioxidants to be used in our diet as well as by industries for functional food formulations.