<p>The organic matter is the most important component of soil material, which determines most soil properties. Among humic substances, humin fraction has been the least studied to date, although it usually constitutes over half of their composition. This is probably due to the fact, that humin fraction has highly hydrophobic properties and is insoluble at all pH values, which makes its isolation much more difficult, compared to humic (HA) and fulvic (FA) acid fractions. In addition, humin fraction forms very stable humic-clay complexes with mineral part of the soil (Stevenson 1994), which cannot be destructed during humic substances extraction. According to the literature, the methods of humin fraction isolation can be divided into two main groups: (1) extraction by different organic solvents, and (2) isolation by extraction of HA and FA followed by digestion of mineral soil components. Nevertheless, each of these methods has different limitations.</p><p>We investigated some modifications of the latter method, obtaining humin fraction from eight mollic horizons of Chernozems and Phaeozems, which differed in their physico-chemical properties.</p><p>The first step was to separate HA and FA according to IHSS method described by Swift (1996), however we adopted different shaking procedure. To asses differences, each supernatant obtained was analyzed for the carbon content concentration, which corresponded to HA and FA extracted.</p><p>HA and FA free residue was then digested to reduce the content of mineral components. We used several digestion with 10% HF/HCl , as higher concentrations of HF can result in structural alteration of the organic compounds (Hayes et al. 2017). To find the optimal time of the procedure, the ash content was determined following each digestion stage. After the HF/HCl treatment, the residue was rinsed with 10% HCl to eliminate secondary minerals. The residue was washed with distilled water until the neutral pH, then transferred to dialysis membranes and dialyzed with distilled water until a negative Cl<sup>&#8722;</sup> test with AgNO<sub>3</sub>. Afterwards the humin fraction was freeze dried.&#160;</p><p>Finally, obtained humin fraction contained various ash content, ranged from 6 to 30%, depending on the soil. The conducted test indicated that: (1) the concentration of carbon in supernatant considerably increased as shaking time was extended to 20 hours, and (2) longer than 4 weeks digestion with HF/HCl did not affect the reduction of the ash content of the humin fraction obtained.&#160;&#160; &#160;</p><p>&#160;</p><p>Literature</p><p>Hayes M.H.B., Mylotte R., Swift R.S. 2017. Humin: Its Composition and Importance in Soil Organic Matter. In: Sparks D.L. (ed) Advances in Agronomy, Vol. 143, Academic Press, Burlington, 47&#8211;138.</p><p>Stevenson F.J. 1994. Humus Chemistry; Genesis, Composition, Reaction. 2nd ed. John Wiley & Sons., New York.</p><p>Swift R.S. 1996. Organic matter characterization. In: Sparks, D.L., et al. (Ed.), Methods of Soil Analysis. Part 3. Chemical Methods - Soil Science Society of America, Book Series no 5,&#160; 1011-1069.</p><p>&#160;</p><p>Acknowledgements</p><p>This work was supported by the National Science Center (NCN) Poland (project No 2018/31/B/ST10/00677 &#8220;Chemical and spectroscopic properties of soil humin fraction in relation to their mutual interaction with pesticides").</p>
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