DRIFT and UV-VIS spectroscopic characterization of humic substances in grasslands after organic and mineral fertilization

Changes in the chemical composition and stability of humic substances after soil amendment with mineral and organic materials were studied. The study site, a Gleyic Fluvisol (Jaroměřice locality, Pardubice Region, Czech Republic), was under permanent grassland managed intensively (four cuts per year) and fertilised as follows: manure and slurry at a rate of 2 livestock units = 120 kg N per ha. The fertilisation treatments included: C – control (no fertilisation); FYM – farmyard manure (30.0 t ha<sup>-1</sup>); CS – cattle slurry (29.0 t ha<sup>-1</sup>); DIG – digestate (29.0 t ha<sup>-1</sup>); NPK – mineral fertilizer (120-30-60 kg ha<sup>-1</sup>). The study aimed to evaluate changes in the structural composition and stability of humic substances during the period 2022–2024, with the goal of providing a knowledge base for farmers to develop best practices for maintaining soil carbon stocks under intensive agricultural conditions. Humic substances (humic and fulvic acids) were extracted with a mixture of 0.1 M NaOH and 0.1 M Na<sub>4</sub>P<sub>2</sub>O<sub>7</sub> /1:1, w/w. Further, humic acids (HA) were isolated according to the standard IHSS method and characterized using UV-VIS and DRIFT spectroscopy. Calculated indices showed higher HA stability and wettability after FYM, DIG, and CS application in comparison with NPK and control. The differences were found in the content of labile aliphatic hydrophobic groups (at 3000–2800 cm<sup>-1</sup>), aromatic stable and resistant C=C groups (at 1660–1580 cm<sup>-1</sup>), and hydrophilic amido-, carboxylic-, keto-groups (at 1740–1600 cm<sup>-1</sup>). The amount of hydrophilic resistant groups at 1660–1580 cm<sup>-1</sup> was comparable at all sites. DRIFT and UV-VIS spectral methods were proposed as rapid tools for assessing humic substance quality instead of the laborious and time-consuming classical fractionation method.