Basil (Ocimum basilicum L.) is one of the most widely cultivated herbal plants, valued in the food and pharmaceutical industries for its abundance of bioactive compounds, and also as an ornamental plant. The contents of its bioactive compounds are strongly influenced by both environmental and anthropogenic factors, among which fertilization plays a key role. This study aimed to evaluate the effects of different application doses (5, 10, 15, and 20 g·dm−3 of substrate) of an organic fertilizer (granulated cattle manure) on the fresh biomass yield and quality of two basil varieties: lemon basil and cinnamon basil. The applied manure doses significantly affected the fresh biomass yield of O. basilicum L. Both basil variety and fertilizer dose were found to determine the content of L-ascorbic acid and nitrates in the plants. Increasing manure doses resulted in higher contents of N, P, and K, as well as decreased contents of Ca and Mg in plants of both varieties