*2.3. Source of Primary Materials*

Plant materials for the determination of dry matter and measurement of N concentration were collected from an area of 1.0 m<sup>2</sup> at BBCH 89 (maturity-nearly all pods are ripe, with dark, hard seeds). Plant samples were taken from the same sowing rows across a particular experimental block. The harvested plant sample was partitioned into a sub-sample of seeds, and post-harvest residues (stem, fallen + remaining leaves, and threshed pods). At the stage of BBCH 89, three yield components were analyzed: (i) the number of pods per m<sup>2</sup> (pod density, PD), (ii) the number of seeds per pod, Se/PD; (iii) the thousand-seed weight (TSW, g). The number of seeds per m<sup>2</sup> (seed density, SD) was calculated based on the first two components. Nitrogen concentration (Nc) in the plant samples was determined using a standard macro-Kjeldahl procedure.

The soil content of NH4-N and NO3-N was determined in field-fresh (not air dried) soil samples within 24 h after sampling. Twenty-gram soil samples were shaken for 1 h with 100 mL of 0.01 M CaCl2 solution (soil/solution ratio 1:5 m/v). Concentrations of NH4-N and NO3-N were determined by the colorimetric method using flow injection analyses (FIAstar5000, FOSS). The method of NO3-N concentration analysis consists of two basic steps: reduction from nitrate to nitrite by using a cadmium column, followed by colorimetric determination of nitrite, based on the Griess–Ilosvay reaction with N-(1-Naphtyl) ethylene-diamine dichloride as a diazotizing agent. The measurement was performed at a wavelength of 540 nm. To determine NH4-N, a special FOSS ammonia indicator (mixture of cresol red, bromcresol purple, and bromothymol blue) was used. The measurement was made at a wavelength of 590 nm. The total Nmin was calculated as the sum of NH4-N and NO3-N, and expressed in kg ha−1. The Nmin content was calculated for a given soil layer, using indices which were constructed based on the soil textural class and soil bulk density [28].
