*2.5. Soil Organic Matter Particle-Size Fractionation*

There are various methods for soil organic carbon partitioning into its granulometric functional pools. The particle-size fractionation of soil organic carbon allows to assess the distribution of SOC according to particle sizes. It gives useful information on the proportion of the different types of soil organic matter, their chemical composition, and potential dynamics in soil which are essential in evaluating the sustainability of various land management options for soil organic carbon rehabilitation. Soil organic carbon fractionation used in this study was adapted from the method develop by Feller [64] for coarse texture and poor humus content soil, used with good overall accuracy by Sainepo et al. [65] in Kenya; by Koussihouèdé et al. [32] in Benin and Gura et al. [66] in South Africa.

A reciprocal shaker was used to mix 50 g of soil with 300 mL of distilled water with 10 mL of Calgon solution (10% sodium hexametaphosphate, 50 g·L −1 ) for 15 h. The solution was passed through a series of nested sieves of sizes 2000 µm, 250 µm and 53 µm in a wet sieving apparatus with deionized water. The particles that passed through the 53 µm was referred as the non-particulate organic matter fraction (NOM). The fraction 53–250 µm was referred as the fine particulate organic matter fraction (fPOM), while the 250–2000 µm fraction was considered as the coarse particulate organic fraction (cPOM) [67]. In the cPOM fraction, plant materials such as plant residues and roots that had partially broken down were carefully separated. The isolated particles for cPOM and fPOM were washed with deionized water until clean and backwashed into an evaporation dish. The fraction that passed through the 53 µm sieve was collected in a volumetric flask, quantified, thoroughly homogenized and a sample of 100 mL was collected in an evaporation dish. The evaporation dishes were dried at 65 ◦C till a constant weight. The oven-dried soil particles were weighed and placed in dry porcelain crucibles and heated in a muffle furnace

at 450 ◦C for 4 h to separate the mineral particles from the organic particles. After cooling, the organic matter contained in each fraction was determined as shown in Equation (2).

$$\text{Fraction} = \frac{\text{Weight at 65 } \text{°C} - \text{Weight at 450 } \text{°C}}{\text{Weight at 65 } \text{°C}} \tag{2}$$

The organic carbon content in the fractions were calculated using the coefficient 1.724 considering that organic matter comprises 58% carbon [68]. Particle-size fractionation of SOC was caried out for two soil layers, 0–10 and 10–20 cm. For each sample the percentage recovery was calculated by the ratio of the sum of the weight of the three fractions by the initial weight of 50 g, multiplied by 100 (Equation (3)) and the total fraction was reported considering 1000 g of soil. The enrichment factor (EF) in each of the fractions was calculated according to Equation (4) [69].

$$\% \text{ Reovery} = \frac{\text{Weight cPM} + \text{WeightfPM} + \text{WeightNOM}}{50} \times 100 \tag{3}$$

$$\text{EF} = \frac{\text{SOC fraction (g/kg)}}{\text{totalSOC (g/kg)}} \times 100\tag{4}$$
