*3.1. Breakthrough Curves and NNO3 Leaching*

The NO3 − breakthrough (Figure 2(top)) and cumulative NNO3 leaching (Figure 2(bottom)) curves show different effects of cover crops on N leaching from the topsoil, depending on tillage. In the NT treatment, there is a clear difference due to plant cover in both leachate NO3 − concentrations and total NNO3 leached over the course of the experiment. Specifically, NO3 − concentrations were lower and there was less NNO3 leached under volunteer plant cover, compared to bare soil. In AuT, the difference between B and V was much more subtle for both NO3 − concentrations and accumulated NNO3 leaching throughout the experiment. Additionally, NT had an overall effect on the elution of nitrate from the intact soil cores, independent of plant cover, where NO3 − peaked earlier and more sharply in NT compared to AuT. This, in turn, indicates increased preferential flow due to better-developed macropore flow pathways [31] in NT.

**Figure 2.** Nitrate concentration breakthrough (**top**) and accumulated leaching of NO3 − N (**bottom**) curves from intact soil cores subjected to ~200 mm of simulated rain in a laboratory lysimiter. The cores belonged to two tillage treatments, no-till (NT) and autumn inversion tillage (AuT), as well as two cover crop treatments, bare fallow (B) and winter rye volunteers (V). Symbols indicate different field experiment blocks.

Statistical analysis of total NNO3 leaching showed significant effects by both plant cover (F = 10.98, df = 8, *p* = 0.010) and tillage (F = 14.72, df = 8, *p* = 0.005), as well as a significant interaction between plant cover and tillage (F = 5.88, df = 8, *p* = 0.041). Pairwise comparisons (Table 1) show that NNO3 leaching under simulated precipitation was significantly lower in V compared to B in NT. The reduction corresponds to 11.3 mg N, i.e., 61% of the NNO3 leached in the bare soil treatment of NT. In contrast, no significant differences were found between cover crop treatments in AuT, with very similar total NNO3 leaching amounts of approximately 20 mg N per core. These results indicate, firstly, that the volunteer treatment in NT significantly and considerably reduced N losses during the leaching experiment. Secondly, this effect was completely lost in the volunteer treatment of autumn-tilled samples. Importantly, there was also no significant difference between NNO3 leaching between the bare fallow treatment in AuT and the bare fallow treatment in NT, indicating that tillage treatment alone did not reduce total N leaching from the intact cores. Finally, all treatments have the same crop history, having been consistently kept under the same crop rotation since the establishment of the long-term trial in 2002. It is therefore highly unlikely that the reduction was caused by factors other than the establishment of volunteers and their interaction with tillage in the year of sampling.

**Table 1.** Nitrogen as NO3 − (NNO3) leached from intact topsoil cores subjected to 200 mm of simulated rain. Mean values and 95% confidence intervals (CI) were obtained from linear mixed effects models with tillage and plant cover as main effects and experimental block as random effect.


<sup>1</sup> Multiple pairwise comparisons significance threshold, *p* = 0.05.

As mentioned earlier, cover crop residence time is known to increase the overall effect of cover crops in reducing N leaching, with significantly lower N leaching with spring incorporation relative to autumn incorporation [10]. However, our results suggest that a difference in autumn cover crop residence of as little as one month can significantly hinder the efficacy of cover crops in reducing N leaching. Indeed, Sieling [17] highlight that N-uptake by cover crops is intrinsically linked to dry matter accumulation and therefore to intercepted photosynthetically active radiation. This means that cover crop residence is of particular importance for N uptake in the autumn when days are still relatively long (and warm). According to Vos and Van Der Putten [32], cover crops in autumn very broadly accumulate dry matter at a rate of 1.12 g per MJ intercepted global radiation. This figure allows us to make a broad calculation of the effect of autumn tillage on cover crops. Based on 65.5 MJ m−<sup>2</sup> total global radiation measured in November at Flakkebjerg station and an aboveground N content of 3.76% measured in plant clippings, the maximum potential N uptake by cover crops in AuT was 87 mg N per core lower than in NT. As discussed before, there are many other factors at play in determining the effect of cover crops in reducing N leaching after harvest, among them the fate of taken-up N once the cover crop is terminated. However, in terms of N uptake alone, prolonged field residence in early and mid-autumn is likely only second in importance to successful establishment of cover crops.
