*2.1. Biochar Characteristics*

Biochar was produced from wheat straw (WSBC) at the pyrolysis temperature of 550 ◦C and time remaining in the reactor 60 s. The BET surface area, cation exchange capacity (CEC), pH in deionized water, CNHSO elemental composition, ash and carbonates content (% volume/dry weight), exchangeable cations and anions content (Ca2<sup>+</sup>, Mg2<sup>+</sup>, K+, Na<sup>+</sup>, P, NH4 <sup>+</sup>, NO3 −), and the total contents of trace elements (Co, Mn and Fe) were determined to describe the properties of the material. The total surface area was determined using a BET (Brunauer, Emmett and Teller) specific surface area analyzer Gemini VII 2390 Series (Micrometrics Instruments Corporation, Norcross, GA, USA). The cation exchange capacity and exchangeable cations (Ca2<sup>+</sup>, Mg2<sup>+</sup>, K+, Na<sup>+</sup>, and NH4 +) were determined according to the modified method described by Munera-Echeverri et al. [31] and analyzed on a microwave plasma-atomic emission spectrometer MP-AES 4200 (Agilent Technologies, Santa Clara, CA, USA). Exchangeable P was analyzed on the MP-AES 4200 after sample extraction according to Olsen et al. [32,33], as 0.5 M sodium bicarbonate (NaHCO3) solution at a pH of 8.5 had a similar pH to WSBC. This extractant decreased calcium in solution (through precipitation of calcium carbonate), and this decrease enhanced the dissolution of Ca-phosphates. The nitrate content was analyzed according to the ISO 14256-1:2003 procedure on a UV-Vis Cary 60 (Agilent Technologies, Santa Clara, CA, USA). The pH values were measured at a ratio of 1:5 (w/v) in deionized water after the sample was shaken for 1 h at 130 rpm with a calibration check pH meter (Mettler Toledo, Columbus, OH, USA). The ash content was determined by weight loss after combustion at 750 ◦C for 6 h in a muffle furnace according to ASTM D7348-13 [34]. CaCO3 was determined following the Scheibler method with a calcimeter [35]. The elemental composition (CHNSO) was analyzed on a CHNS analyzer (CE Instruments, Hindley Green, UK), and the O content was calculated from the difference. The total content and exchangeable forms of metals (Co, Cu, Fe, and Mn) were analyzed on a microwave plasma-atomic emission spectrometer MP-AES 4200 (Agilent Technologies, Santa Clara, CA, USA), respectively, after microwave sample digestion in 70% nitric acid (1:10 w/v ratio) in a digestion microwave system StartD (Milestone Srl.Sorisole, Italy) and extraction with deionized water (1:25 w/v ratio).

#### *2.2. Metal Sorption Mechanism Analysis*

Previous studies of Co sorption in soil have showed that Co and Cu as divalent cations can compete for sorption sites, especially in excess of Cu2<sup>+</sup> in multi-contaminated soils, as described by Muyumba et al. [36]. In the metal sorption experiment, both metals were used to simulate natural conditions in contaminated soils and possible interactions of the Co and Cu ions. The FTIR spectroscopy was used to compare potential changes in the functional groups of metal-loaded biochars with the biochar samples before the Co or Cu sorption. The Co and Cu sorption on the wheat straw biochar was determined by the simplified batch equilibrium method according to the OECD 2000/106 protocol [37]. To compare the effect and probable interaction between cobalt and copper that could occur in multi-contaminated soils the following three solution were used in the batch experiment: (1) Co (II) acetate, (2) Cu (II) acetate, and (3) Co (II) + Cu (II) mix of both salts. Briefly, 5 g of each salt was diluted in 500 mL of deionized water and set overnight to reach equilibrium. The pH of each solution was measured before and after the batch experiment to determine if the pH change occurred after biochar BC addition which would affect sorption conditions. One gram of wheat straw biochar and 20 mL of each solution were placed in 50 mL volume polypropylene falcon centrifuge tubes. All samples were prepared in three replicates of each treatment. The sealed samples were placed on the rotary shaker Multi RS-60 (Biosan, Riga, Latvia) at 80 rpm and 22 ± 0.5 ◦C for 24 h. Sorption equilibrium was reached within less than 24 h. Then, samples were centrifuged for 25 min at 10,000 rpm to separate the biochar from the solution according to procedure described by Cwiel ˛ ´ ag-Piasecka et al. [38]. The biochar samples were washed three times with 20 mL of deionized water and prefiltrated on Munktell No. 2 filter papers (Ahlstrom Munksjö, Helsinki, Finland) to rinse off excess metal cations. The biochar samples were dried in an oven drier at 60 ◦C for 6 h to prepare pellets for Fourier transform infrared spectra (FT-IR). The FT-IR analysis of the wheat straw biochar samples were recorded using a Vertex 70 FT-IR spectrometer (Bruker, Billerica, MA, USA) on KBr pellets (about 1 mg sample for 400 mg of KBr) according to the standard method used for sample preparation for FT-IR spectra analysis. The incubation experiment with multi-contaminated soils was a pot experiment with 24, four-liter pots (approximately 3 kg of soil each). Two soil types, sand and loam, were set as 12 control pots, six for each soil type. A similar 12 pots were amended with a dose of BC (5.0% w/v), six for each soil type. The soil mixtures were incubated for two years, keeping the humidity of the pots at 60% of maximum water holding capacity. After the time period, the soil samples were collected from each pot, air dried, and sieved (<2 mm), and sequential extraction of Co was performed. The existence of possible precipitates of Co after metal sorption was checked using a scanning electron microscope (SEM) (Bruker, Billerica, MA, USA) coupled with an energy dispersive X-ray analyzer (EDX) (Bruker, Billerica, MA, USA). The biochar particles for SEM-EDX (scanning electron microscopy with energy dispersive X-ray spectroscopy) analysis were separated from the soil by progressive sieving of the soil-biochar mixture. Biochar fraction <1 mm was used to determine elemental composition of the material surface and the distribution of ions [39]. For the morphological observations, SEM was applied according to Michalak et al. [40].
