This study evaluated the effect of two cultivation systems (organic and conventional) on the soil properties, agronomic performance, and biochemical quality of strawberries of the Llosana and Fortuna varieties. The soil analysis revealed an improvement in the organic status in a biological system, characterized by a higher organic matter content (1.85%) and a lower C/N ratio (11.0), indicating increased microbial activity. On the other hand, the conventional system showed higher concentrations of trace elements, in particular iron and manganese. From an agronomic point of view, the conventional system has significantly improved growth and yield, with values reaching 883.33 g for Fortuna and 806.33 g for Llosana, against 538.00 g and 480.33 g, respectively, in the organic system. However, the latter favored a better nutritional quality, with an increase in dry matter, soluble sugars, and proteins. Biochemical analyzes have shown that the biological system stimulates the accumulation of phenolic compounds and strengthens antioxidant activity, with a significant increase in total antioxidant capacity and a decrease in IC50 values. Moreover, the volatile profiles revealed a dominance of esters in the conventional system, conferring fruity aromas, while the biological system favors furanones, responsible for more complex and sweet aromas. Finally, the antibacterial activity was more marked in the extracts from the biological system, suggesting an increased accumulation of bioactive compounds. These results highlight a compromise between yield and quality and underline the importance of choosing the cultivation system according to the production objectives.