## 3 Results and Discussion
### Relations Between Experimental Factors and Parameters Tested  
To gain a deeper understanding of the similarities and differences among the examined barley grain samples regarding their polyphenic and carotenoid content as well as mycotoxin contamination, a principal component analysis (PCA) was conducted (Figure 3a,b, respectively). The PCA of bioactive compounds in barley samples of different varieties (Zoom, Galation, Hobbit, and Sandra) cultivated with different N fertilization levels (60, 90, 120, and 150 kg N ha\({}^{-1}\)) revealed that the first main component explains 39.56% of the total variance, while the second component explains 17.22%. The PCA score plot shows that samples coming from the field plots fertilized with 60 kg and 150 kg N ha\({}^{-1}\) tend to cluster in opposite quadrants, indicating distinct profiles of bioactive compounds at these fertilization levels. No clear grouping of results based on bar  
Figure 3: The plot of principal component analysis (PCA) showing relationships between the chemical composition of grains of the studied barley varieties grown with different doses of N fertilizer: (**a**) carotenoids, phenolic acids, and flavonoids; (**b**) mycotoxins. 60 kg N ha\({}^{-1}\) (N60), 90 kg N ha\({}^{-1}\) (N90), 120 kg N ha\({}^{-1}\) (N120), 150 kg N ha\({}^{-1}\) (N150), 15-acetyldeoxynivalenol (15-AcDON), and deoxynivalenol (DON). ley variety was observed, suggesting that N fertilization level has a more pronounced effect on the studied bioactive compounds than genotype. Lower nitrogen application (60 kg N ha\({}^{-1}\)) was associated with higher contents of phenolic acids, particularly ferulic and caffe acid, as well as flavonoids such as quercetin-3-O-rutinoside and kaempfer, in grains. Conversely, the highest fertilization level (150 kg N ha\({}^{-1}\)) was related to increased levels of gallic acid, quercetin, apigenin, and the kaempfer-3-O-glycoside. Additionally, the highest fertilization dose was linked to increased grain yields, indicating a potential trade-off between bioactive compound profiles and productivity (Figure 3a). The demonstrated relationships are also confirmed by other authors studying cereals [74; 75]. The PCA of 15-AcDON and DON mycotoxins' contents in barley samples revealed that the first principal component explains 83.47% of the total variance, while the second component explains 16.53%. The biplot clearly distinguishes the barley varieties Hobbit and Galation from samples subjected to nitrogen fertilization at 90 and 120 kg N ha\({}^{-1}\), with these groups associated with higher concentrations of 15-AcDON and DON mycotoxins. Specifically, the Hobbit and Galation samples from the highest and intermediate nitrogen treatments tend to exhibit elevated mycotoxin levels, indicating a correlation between increased nitrogen fertilization and mycotoxin contamination. This is confirmed by other studies, which have shown that increasing the nitrogen dose accelerates the development of _Fusarium_ (fungi producing DON and 15-AcDON) and increases the production of mycotoxins, especially at high humidity during flowering [30]. Conversely, the varieties Sandra and Zoom cluster closely together in the PCA space, suggesting similar mycotoxin occurrence profiles across different fertilization levels. Notably, the Sandra samples, despite varying nitrogen applications, remain the most similar within their group (Figure 3b). These findings, similar to another study [76], underscore the influence of both barley variety and nitrogen fertilization on mycotoxin contamination, with certain varieties and treatment levels associated with higher toxin accumulation. The Spearman correlation analysis revealed several significant relationships between phytochemical contents, crop yield, and fertilization levels in barley cultivation (Figure 4). Notably, barley grain yield was strongly and positively correlated with the levels of _p_-coumaric acid and sinapic acid, suggesting that higher concentrations of these phenolic acids may be associated with increased biomass production.