**2. Materials and methods**

#### **2.1 Study area**

This study was carried out on a Kraft pulp and paper mill in Webuye (34–36° E, 0° 03<sup>0</sup> -1°15' N) in the upper catchment of the Nzoia River (**Figure 1**). The Nzoia River basin is approximately 12,696 km<sup>2</sup> [40] and lies within the Lake Victoria basin in Kenya, East Africa. The river flows between 2700 and 1134 m above sea level. The region receives an average annual rainfall of 1350 mm. The average annual air temperature varies between 8°C and 28°C, with minimum temperatures between 8°C and 12°C and maximum temperatures of 24 and 28°C [41]. Food crops such as maize,

**Figure 1.** *Study area and study sites.*

sweet potatoes and cassava, sorghum, millet and vegetables are grown on small-scale farms, usually extending up to the river banks. Livestock farming is practiced and River Nzoia provides water for both domestic and industrial use. The mill consumes about 40,000 m<sup>3</sup> of fresh water and discharges between 35,000 to 40,000 m<sup>3</sup> daily into the river at a dilution rate between 0.3 to 3.2%, depending on the seasonality of the river discharge. The mill's effluent takes 6 weeks to flow through a set of settling tanks (one primary and one secondary), two aerated lagoons, and two stabilization ponds before discharge into River Nzoia. Recent expansion programs within the mill have led to an overloaded wastewater treatment system, initially designed to treat only 25,000 m3 of mill effluent per day.

*Distribution of Potentially Toxic Elements in Water, Sediment and Soils in the Riparian… DOI: http://dx.doi.org/10.5772/intechopen.102440*

## **2.2 Study sites**

There were three sampling sites, namely, the 'Water intake point' (WIP), the 'Effluent deposition/discharge point' (EDP) and the 'Downstream point' (DSP). These sites were located along the profile of River Nzoia (**Figure 1**). The WIP was situated upstream of the factory discharge point and the river width at this location was 6.53 m. The EDP was approximately 3.2 km from the WIP and the river width was 5.92 m. The downstream point was 3.2 km from the EDP with a river width of 6.44 m. All the sampling sites located on the factory side were designated Side AA (**Figure 2**), while those on the opposite side were designated as Side BB.

**Figure 2.** *Trends of heavy metal concentration mg/g in wetland soil at WIP in PanPaper.*

### **2.3 Sampling and analysis of pulp and paper industrial wastes**

Grab samples of 3 kg each of lime mud and boiler ash were collected from the recovery area of the pulp and paper mill in Webuye. These samples were spread on trays; air dried overnight, sieved using a No. 9 mesh sieve (2.00 mm) and stored in plastic bags. The pH of these samples was determined using the ASTM D 4972–01 Standard Test Method for soil pH (Electrometric method). The ASTM D 2216–98 Standard Test Method for Laboratory Determination of moisture was used to measure the moisture content. The concentration of PTEs was determined using an Inductively Coupled Plasma-Optical Emission Spectrometry (ICP-OES). The mill liquid effluent was tested for various parameters at different treatment stages and compared with national effluent discharge standards.

#### **2.4 Sampling and analysis of river water**

Grab river water samples were collected using half liter metal free *Van Dorn* bottle. These samples were collected at about 0.5 m below the water surface. Before collecting the samples, the *Van Dorn* bottle were washed in 2 L tap water, and rinsed three times in distilled water. The collected samples were then transferred to half liter polythene bottles pre-soaked in nitric and sulfuric acids solution at 1:1 volume ratio. The water samples were acidified to a pH 2 using concentrated nitric acid [42], and stored in an ice box before transportation to the laboratory for chemical analysis.

Temperature, pH and electrical conductivity (EC) were measured *in situ* using a calibrated JENWAY 3405 electrochemical analyzer (Barloword Scientific Ltd., Essex, UK), with a specific probe for each variable. The equipment was calibrated using deionized water before measurements. The measurements were conducted in triplicate for each site included in the study.

#### **2.5 Sediments and soil sampling**

Thirty-six sediment samples were collected from each sampling site. The sediments were collected at the bottom of the river using an *Ekmans* Grab Sampler. A polypropylene spatula was used to transfer the sediment sub-samples to acid rinsed polypropylene bottles and placed in an icebox for transportation to the laboratory for chemical analyses. Soil samples were collected using a soil auger along the river banks within the organo-mineral layer of 0–25 cm of the soil. The samples were then kept in black polythene bags, labeled and stored in an icebox before transportation to the laboratory for chemical analyses.

To determine PTEs concentration, soil samples were scooped at five equidistant points from the river bank, i.e. 0.0, 0.5, 1.0, 1.5 and 2.0 kilometers respectively away from the River Nzoia bank on both sides AA and BB. The 0.5 km distance was deemed sufficient to reflect lateral variations of potentially toxic elements concentrations in the soil.

#### **2.6 Determination of leaching capability of PTEs in sediment and soils**

A leachability tests were used to evaluate the possible elution of Pb, Cu and Zn from the sediment under selected treatment conditions. These tests took place because *Distribution of Potentially Toxic Elements in Water, Sediment and Soils in the Riparian… DOI: http://dx.doi.org/10.5772/intechopen.102440*

the paper mill periodically discharges the recovery boiler ash and lime mud into the river. The sediment and soil samples were filtered through a Ø 9.0 mm filter paper, air dried, crushed and then sieved through of 9.5 mm pore size. Distilled water was used as the leaching solution, adjusted to a pH 5 using a mixture of sulfuric and nitric acids (80, 20 by weight). Initially, 50 grams of sediments were added to the leaching solution followed by soil at a solid to liquid ratio of 1:20 (proportional to 1000 mL leaching solution). The mixture was agitated in a rotary agitator for 18 hours at 30 rpm and a constant temperature of 25°C. After agitation, the soil slurry was filtered through a 0.8 mm glass fiber filter. The liquid extract was digested using concentrated HNO3 and analyzed for Cr, Cu, Pb and Zn using an ICP-OES. All the analyses were run in triplicate.

#### **2.7 Analytical determination of PTEs in water**

Water samples were digested using sulfuric and nitric acids before spectrophotometric analysis was conducted to minimize the interference of complex organic matter [43]. The samples were digested and concentrated on a hot plate from 100 mL to 25 mL for 3 hours. After digestion, the samples were allowed to cool to room temperature and then 2 mL of 30% hydrogen peroxide (H2O2) were added to oxidize any residual organic matter. Further cooling followed before the digested samples were filtered through a 0.45 μm nucleopore membrane filter over a vacuum pump. The filtrates were stored in 125 mL polyethylene sample bottles, at 4°C before analysis for Zn, Cu, Pb, and Cd in an Atomic Absorption Spectrophotometer (Model AA 10/20). In this study, at least two calibration standards were prepared for each metal before recording the measurements. The PTEs were determined at various spectrophotometric wavelengths and slit width (Pb = 17.0 nm; Zn = 13.9 nm; Cu = 324.8 nm; Cd = 228.8 nm and slit width for Pb and Zn = 1.0 nm and Cd and Cu = 0.5). The concentrations of PTEs in water were calculated and reported in mg L<sup>1</sup> .

#### **2.8 Determination of PTEs in soil, sediment and industrial wastes**

The study used analytical grade reagents and the stock solutions met the Merck certificate AA standards. In addition, all experiments used milli-Q water. Plastic and glassware were cleaned by soaking them in 14% (v/v) HNO3 for 24 hours and rinsed with nano pure water. The sediments, soils samples and industrial wastes were crushed and homogenized using a Fritsch, Pulverisette 5, planetary mill (Fritsch GmbH Laborgerate, Idar-Oberstein, Germany) for 5 minutes at 400 rpm. About 0.20 g of soil and sediment samples and about 25 mL of water were weighed in Teflon (© poly-tetra-fluoretheen (PTFE), DuPont™) high pressure vessels. Then 4.0 mL concentrated nitric acid (65%), 1.0 mL concentrated hydrochloric acid (37%) and 1.0 mL ultrapure water were added to the samples. Six samples of each item were placed in the carousel of a *Paar* Microwave oven (Anton Paar GmbH – Graz –Austria). The samples were digested in a microwave oven (Anton Paarâ GmbH Kärntner Straße 322 A-8054 Graz/Austria) at a maximum temperature of 220°C and pressure of 75 bars for 15 minutes.

After cooling, the resultant clear solutions were poured into 50 mL volumetric flasks and diluted to the mark with ultra-pure demineralized water (Barnstead NanoPure, Thermo Fisher Scientific Inc., Barnstead International,

Iowa USA). Finally, the diluted solutions of the respective samples were transferred into acid cleaned polyethylene bottles. All elements were determined using an Inductively Coupled Plasma-Optical Emission Spectrometer (Perkin Elmer Optima 3000 XL, ICP-OES) with the PE calibration standards. The concentrations of PTEs in soil and sediments were calculated and reported in mg kg�<sup>1</sup> dry weight. The quality of the analytical process was controlled by the analysis of IAEA MA-A-3/TM certified standard reference material of river sediment. Care was taken to ensure analytical results varied from certified values by less than 10%.
