Common mistletoe (Viscum album L.) is a semi-parasitic plant that grows on various woody species. Due to its anti-inflammatory, antibacterial and antitumor properties, it is used in the treatment of various diseases and conditions. Particular emphasis is placed on the antioxidant properties of mistletoe, which depend on host type on which it parasitizes, the region and the sampling time. In this paper, the influence of location and sampling time on the biochemical composition and antioxidant activity of freshly harvested mistletoe samples and mistletoe tea preparations was investigated. Higher content of total soluble polyphenols, antioxidant activity, and concentrations of carotenoids and total chlorophyll were observed in samples harvested in October in comparison to those harvested in February. Fresh samples and tea did not show a significant difference in the examined parameters except for the samples in Istria, Petrijevci and Aljmaš. Differences were observed between freshly harvested samples of leaves and twigs and the content of total soluble polyphenols and antioxidant activity among individual tea preparations. The result of the principal components analysis method indicated certain groupings among the samples in accordance with the results of biochemical analyses. The obtained results confirm the influence of location and seasonal variations on the phytochemical composition of mistletoe.