The analysis of the ANOVA of the presented results proved a significant positive influence of N fertilization on the rye grain yield and on the concentration and uptake of microelements (Fe, Mn, Zn and Cu) by its whole-grain dry mass (DM) (see
Table 4 and
Table 5, and
Tables S1 and S2 in the Supplementary Material for the comprehensive dataset analysis). The performed statistical analysis did not show any interaction between the used doses of nitrogen and sulfur. The experimental year was significant for all studied features. The interaction between N fertilization and the year of study was significant only for the yield of grain and Cu uptake by the whole-grain DM of spring rye. The interaction between S fertilization and the year of study was not significant.
3.1. Nitrogen
In our presented research, each applied level of nitrogen fertilization (30, 60 and 90 kg ha
−1) had a significant positive effect on spring rye grain yield. However, the highest grain yields were found after the use of 90 kg N ha
−1 (3.68 t ha
−1). This yield increased by about 28.5% compared to the control. The concentration of microelements in the grain of spring rye increased together with the increase of the nitrogen dose and was the highest after the use of 90 kg N ha
−1 (Fe—33.65, Mn—39.48, Zn—37.50 and Cu—4.65 mg kg
−1 DM). The accumulation of micronutrients in the whole-grain DM also statistically significantly increased at the same time as the increasing N fertilization rate and was the highest after the use of 90 kg N ha
−1 (Fe—124.02, Mn—145.40, Zn—138.17 and Cu—17.14 mg ha
−1) (see
Table 6,
Figure S2A and Tables S1 and S2 in the Supplementary Material for the comprehensive dataset analysis).
Numerous scientific reports show that the amount of macro- and micronutrient accumulation depends on many factors. These include, for example, the plant type, species and variety [
13], climate and soil conditions [
14] as well as other agronomic and production factors. According to Ragaee et al. [
15] rye grain (obtained from the Experimental Farm at UAE University) is ample in Fe and Mn. In turn, Kan [
16] indicates that rye (cultivated on sand–loam soil) has the highest content of Zn among major cereal species. The grain of rye cultivated on a lessive soil of a medium silty loam and fertilized with NPK (203 kg ha
−1) in north-eastern Poland contained fewer micronutrients than in our research, especially Mn [
17]. In another study in Poland, six varieties of winter rye fertilized with nitrogen at doses from 80 to 100 kg N ha
–1 were analyzed [
18]. It was found in the grain of these varieties that there were, in turn, more Fe, Mn and Cu [
18]. In studies with 21 winter rye varieties carried out in Russia, it was found that the content of microelements in the grain varied among rye genotypes: Fe—2.1-fold, Zn—1.6-fold and Mn—2.7-fold [
13].
In the studies of Kadłubiec and Bojarczuk [
19], spring rye was cultivated on a good rye complex of soil and fertilized with nitrogen at a dose of 80 kg N ha
−1; in this condition, it gave a grain yield in the amount of 5.1 t ha
−1. In the study conducted by Stępień et al. [
17] with winter rye fertilized with 90 kg N ha
−1, it was found that the grain yield was about 17.5% higher than at control plots and the contents of Mn, Zn and Cu were a little higher than the control. In the study of Teklić et al. [
20], it was found that fertilization with higher doses of nitrogen influenced the content of Fe, Zn and Cu in grain, but had no effect on the level of Mn accumulation. A similar phenomenon was observed in our own research, where a dose of 30 and 60 kg N ha
−1 significantly affected the Mn content (
Table 6,
Figure S2A and Tables S1 and S2 in the Supplementary Material for the comprehensive datasets analysis). Based on the research of Kutman et al. [
21], we found that the fertilization with a high dose of N has a positive influence on the uptake of iron and zinc by grains of wheat. As can be seen in the literature, the used forms of N application have different influences on the accumulation of Mn in the plant. Therefore, to obtain the best results, nitrogen fertilizers should be used according to the following order: (NH
4)
2SO
4 > NH
4NO
3 > Ca(NO
3)
2 [
22].
3.2. Sulfur
As shown in the presented experiment, the supplementation of sulfur improved the NPK effect, because the rye grain yield increased by 80.0 kg ha
−1 (2.5%) (see
Table 6,
Figure S2A, and Tables S1 and S2 in the Supplementary Material for the comprehensive dataset analysis). As is well known, nitrogen is the most yielding element. Sulfur, in turn, improves nitrogen utilization in the plant and improves the quality characteristics of plants. Klikocka and Cybulska [
11] found that, after using sulfur in wheat grain, the protein, methionine and cysteine content increased, and flour baking properties improved. However, the yield of wheat grain after applying 50 kg S ha
−1 increased by 3.6%. According to Podleśna [
23], the addition of 60 kg S ha
−1 caused an increase in winter wheat grain yield of 11%.
Our own research showed that the concentration of Fe, Zn and Cu in rye grain after sulfur application at a rate of 40 kg ha
−1 increased in comparison with the control, by about 8.8%, 2.8% and 3.6%, respectively. However, the concentration of Mn was decreased by 5.0%. The uptake of Fe, Zn and Cu by grain DM following fertilization of 40 kg S ha
−1 increased by 11.3%, 2.8% and 6.0%, respectively, in comparison with the control. The addition of S to NPK fertilization decreased the uptake of manganese (Mn) by rye grain DM by about 2.8% (see
Table 6,
Figure S2A, Table S1 in the Supplementary Material for the comprehensive dataset analysis).
Many scientific papers show that nitrogen is the most important yield-forming element. However, nitrogen fertilization is effective when the soil is rich in other nutrients, such as sulfur [
11]. Barczak et al. [
24] report that in conditions of sulfur deficit in soil, the yield-generating nitrogen efficiency is reduced, while more intensive nitrogen fertilization deepens sulfur shortage. This in turn inhibits the uptake of nitrogen by plants, limiting their growth and development. In the conditions of sulfur deficiency in soil, N fertilizer does not show optimal efficiency and causes the reduction of yields, as well as changes in the chemical composition of plants and in quality features. Because sulfur decreases soil pH, it indirectly influences the availability of Zn, Fe, Mn, Cu and other micronutrients [
25,
26,
27].
3.3. Interactions and Correlation between Nutrients
The research results showed that the addition of sulfur to each N dose caused a trend of increasing the content and uptake of microelements and the yield of spring rye. This type of interaction between nitrogen and sulfur is known as the additive interaction (of sulfur) [
28], because the assimilation of N and S in plants is closely related [
29]. Therefore, joint fertilization with these two elements significantly influences both the uptake and metabolism of nitrogen and sulfur and other elements in the plant.
Furthermore, we observed statistically significant positive correlations between the grain yield and the content and uptake of all micronutrients (
Table 7). The correlation coefficients between the content of micronutrients and the grain yield decreased in the order Fe > Mn > Cu > Zn. However, in the case of the correlation between the uptake of micronutrients by grain DM and the grain yield, the strength of the relationship was in the order Mn > Zn > Cu > Fe. On the basis of an experiment with the application of increasing doses of sulfur, Klikocka and Marks [
30] found a positive correlation between the sulfur dose and the accumulation of elements in the plant.
In the presented study, a positive correlation between the content and uptake of Fe, Mn and Zn was found. Additionally, a significant correlation was found between the content of Mn and Cu, while no correlation was found between the content and uptake of Zn and Cu and additionally between the Zn uptake and the Cu content (
Table 7). Based on the studied literature, it can be supposed that metallic nutrients can compete with iron and, in consequence, limit iron uptake by the plant. It has been known for a long time that Fe and Mn compete in the process of iron uptake by the plant [
31]. Mn
2+ has similar properties to Ca
2+ and Mg
2+ (alkaline cations) and Fe
2+ and Zn
2+ (heavy metals); therefore, these cations compete with Mn for transport and uptake by the plant [
32]. A negative correlation between Fe and Cu accumulation in durum wheat was also observed. This phenomenon may indicate the mutual antagonism of these two elements and may affect iron deficiency in the plant [
33]. It is well known that a shortage of one chemical element may influence the uptake of one or more other elements [
34]. In the literature, it has been shown that, under the conditions of nutrient shortage in the soil, there is a competition in uptake, especially between Mn
2+ and Cu
2+ or Zn
2+ [
35], and between Fe
2+ and Zn
2+ or Cu
2+ [
36].
3.4. Weather Conditions
In the present experiment, the grain yield of spring rye as well as the accumulation and uptake of microelements were modified by the weather conditions. According to many authors, high yields of cereals are usually dependent on low rainfall during winter and April, while higher rainfall is necessary in the booting (BBCH 41–49) and flowering phase (BBCH 61–69) [
37,
38]. On the other hand, Tarkowski [
39] reports that rye, like other cereals, shows the greatest demand for water in the period from booting (BBCH 41–49) to heading (BBCH 51–59). Chmura et al. [
40] found that the water requirement of spring cereals for their optimal growth is from 230 to 300 mm. In the research of Dopka et al. [
41], the highest yield of spring rye grain was obtained when the sum of atmospheric precipitation during the vegetation period was 254.3 mm. In our own research, the highest yield of spring rye grain (on average 3.315 t ha
−1) was obtained in the vegetation season of 2011, when the sum of precipitation was 414.6 mm (k = 1.3), and the months of April (36.2 mm), May (31.4 mm) and June (52.9 mm) were characterized by an optimal rainfall distribution [
10]. However, high atmospheric precipitation in April, May and June in 2009 and 2010 adversely affected the yield of spring rye grain. It was also found that the air temperature in the vegetation seasons of the presented experiment exceeded the average temperature over several years [
10]. Rymuza et al. [
42] explained that air temperature is the main factor affecting the rate of plant development, while the water factor affects the yielding of cereals, especially in the critical period between the booting (BBCH 41–49) and heading stages (BBCH 51–59).
The content and uptake of Mn and Zn as well as Cu uptake by the grain DM of spring rye were most positive in the weather conditions that occurred in 2011 (rather wet). In the 2009 period (rather dry), the weather conditions also had an important effect on favorable Cu accumulation and on Fe content and uptake by the grain DM of spring rye. The concentration and uptake of microelements, except the Zn content, were smallest in 2010 (period rather wet) (
Table 6). This may be due to the very low precipitation in the stages of grain formation and ripening. In the study of Woźniak and Stępniewska [
14], the lowest content of Cu was also found in the grain harvested in the year with the lowest rainfall and high air temperature. However, the content of Fe, Mn and Zn in the wheat grain in the above-mentioned study was not significantly affected by weather conditions, in contrast to our own results.