Cardoon leaves are of great pharmaceutical importance due to their high content of polyphenol compounds. Polyphenolic compounds have attracted much interest due to their health-promoting effects. The content of these compounds in C. cardunculus depends on several factors, such as genotype, crop management, plant tissues, harvest time, and storage time. In this study, the effects of nitrogen (N) fertilisation (rates and forms) on the biomass yield and polyphenol profile of the leaves were determined. Increasing the amount of N up to 180 kg-ha−1 in fertilisation did not significantly increase the air-dried biomass yield of the leaves. On the contrary, it led to lower concentrations of total phenolic compounds (TP), total flavonoids (TF), caffeic acid, cynarin, and luteolin. Improvements in performance were achieved when 120 kg-ha−1 N rate was applied and increases in TP, TF content, and radical scavenging activity were observed. The applied N forms (NO3, NH4 or urea) had different effects on the concentrations of individual compounds and leaf air-dried biomass. Higher concentrations of cynarin, luteolin, and luteolin-7-O-glucoside were found when the N forms NH4 and urea were applied; higher caffeic acid content was found when urea was applied. The application of NO3 and urea in fertilisation reduced the level of luteolin-7-O-rutinoside, while the application of NO3 and NH4 reduced the amount of caffeic acid. The obtained results provide a better understanding of the effects of N rates and forms on cardoon leaves over two growing seasons.