Study Area
Kabwe town is located in the Central Province of Zambia about 136 km north of Lusaka, the capital of Zambia. The town is characterized by Acrisols although Podsols and Vertisols are also found. Vegetation is mainly characterised by Miombo woodlands (Kribek et al., 2010). Kasanda residential area was chosen for soil sample collection because it is near and adjacent to Kabwe Zinc and Lead Mine (Fig. 1). Most studies also reveal that, Kasanda residential area is highly contaminated with Pb from the mining activities.
Soil Collection And Characterisation
Soil samples were collected at 0–20 cm depth using a Trowel at 5 points and thoroughly mixed using plastic buckets. At each point, soil samples from the 5 gardens were collected then soil samples mixed to form one composite sample representative of the whole sampling area.
The collected soil sample was air dried, and visible roots and other debris were removed. The larger soil aggregates were gently broken using fingers sieved through a 2.0 mm sieve and mixed using a Trowel in plastic buckets thoroughly to prepare a composite soil sample. The sieving of the composite soil sample was done to make a homogeneous soil sample. The sieved soil was characterized before setting up the pot experiment for soil pH, total Pb concentration, electrical conductivity, ammonia, and nitrates and phosphorous.
Amendments Collection, Preparations And Characterisation
Chicken manure
Chicken manure was collected from the University of Zambia, School of Agricultural Sciences Field Station poultry house. It was air-dried and sieved using a 2 mm sieve. The chicken manure collected was from broiler chickens and the bedding material was sawdust. The chicken manure was characterised for pH, total Pb concentration, electrical conductivity, organic carbon, cation exchange capacity and some metalloids.
Chicken Manure – Derived Biochar
The chicken manure-derived biochar was made using the pit method under limited oxygen conditions (Fig. 2), and then, the charred materials were ground using a mortar and pestle and sieved through a 2 mm sieve. The chicken manure –derived biochar was equally characterised for pH, total Pb concentration, electrical conductivity, organic carbon, cation exchange capacity and some metalloids.
Experimental Design And Treatments
The complete randomized design (CRD) was employed in this study. A pot experiment was conducted in a greenhouse at the University of Zambia. It was set up with 4 replications. The amendments application rates were calculated as percentage of mass of the soil in the pot. Chicken manure and Chicken manure-derived biochar were applied at rates of 2% and 4% of the soil mass.
For the pot experiment 2500g soil was used and the manure and biochar amendments were set up with a control, meaning, CT- no amendment was applied, 2% of 2500g of soil of which calculated 50g of Chicken manure and Chicken manure-derived biochar was applied (CM2% and CMB2%), 4% of 2500g of soil of which calculated 100g of chicken manure and chicken manure-derived biochar was applied (CM4% and CMB4%) as shown in Table 1 below. The treatments below were repeated with 200g soil for destructive sampling with similar application rates for analysis of soil pH and bioavailable Pb. All treatments were replicated 4 times.
Table 1
Treatment set up showing application rates calculated by percentage of mass of the soil in the pots.
Treatment | Mass of soil (g) | Mass of amendment applied in grams |
Control (CT) | 2500 | 0 |
CM2% | 2500 | 50 |
CM4% | 2500 | 100 |
CMB2% | 2500 | 50 |
CMB4% | 2500 | 100 |
CT is un-amended polluted soil; CM2% and CM4% are chicken manure treatments at 2% and 4% respectively; CMB2% and CMB4% is chicken manure-derived biochar treatments at 2% and 4% respectively.
Planting, Irrigation And Crop Management
Five (5) seeds of each variety of vegetable, that is, rape (Brassica napus) and Chinese cabbage (Brassica rapa) were planted. After germination, two healthy seedlings per pot were maintained after thinning. Tap water was used to irrigate the treatments as needed.
The rape (Brassica napus) was sown on 12th March, 2020 a day after soil-amendments mixture preparations. It was then harvested on 13th May, 2020 (harvested after 9 weeks). Thereafter, the pots were left for five weeks without any plant planted in them. This soil incubation was done to determine the residue effects of the amendments on bioavailable Pb in the soil. Incubation of soil samples is used to measure mineralization (Hannah, 2013). The second crop plant, Chinese cabbage (Brassica rapa), was planted on 21st June, 2020 and was harvested on 11th August, 2020 (8 weeks after). Chinese cabbage was chosen as a second crop, because it is also a Brassica vegetable. It has ability to accumulate Pb in its edible parts. It is equally most grown vegetable in Kabwe town. Management practices were the same for both the first and second crops.
Soil And Amendment Analysis
Soil analysis
During the experiment, soil samples were collected six times for analysis of bioavailable Pb and soil pH. Bioavailable Pb was extracted in DTPA – TEA (Diethylene Triamine Penta Acetic Acid, Triethanolamine) buffered at pH 7.3 according to (Lindsay and Norvell, 1978). Ten grams of soil was weighed and 20 ml of DTPA-TEA solution added. The mixture was shaken for two hours using the mechanical shaker. The suspensions were filtered by gravity through the filter paper. Then filtrates were analyzed for Pb concentration using Flame Atomic absorption spectroscopy (FAAS - Z series 2010). To determine soil pH, 25 ml of CaCl2 was added to each ten grams of soil and shaken for thirty minutes using the mechanical shaker. pH was determined using a soil pH meter. The electrical conductivity for soil was determined by adding 25 ml of deionized water to ten grams of each sample. The mixture shaken for one hour using mechanical shaker. The filtrates analyzed for electrical conductivity using electrical conductivity meter.
Organic carbon was determined by Walkley and Black method. One gram of air dry soil into a 250 cm3 conical flask to which ten centimetre cubic of 1N K2Cr2O7 (with a pipette) was added. Then rapidly 20 cm3 of concentrated H2SO4 was added and was directed into the stream suspension (automatic pipette was used).
Cation exchange capacity was determined using distillation method. Ten millilitres of leached solution (25 ml 1N NaCl solution in a 50mls volumetric flask) was pipetted. Magnesium oxide (MgO) was added as a catalyst. Distillation was done for five minutes while collecting NH4+ in 20 ml boric acid indicator. Thereafter, the indicator was titrated with 0.1 N HCl solution from green to purple and then, meq CEC/100 g soil was calculated.
Ammonium and nitrate was extracted using 2M KCl. Ten grams of soil sample was weighed into conical flask and added to 100 ml 2M KCl solution. Then the mixture had shaken for 30 minutes. Thereafter, filtered the solution and the clear filtrate taken for analysis within 24 hours.
The available phosphorous in the soil was determined using 0.5M NaHCO3 (pH 5.8) as extracting solution. The soil texture of the soil sample was analyzed using Hydrometer method.
Amendments Analysis
The electrical conductivity for amendments (chicken manure and chicken manure – derived biochar) was determined by adding 25ml of deionized water to ten grams of each sample and shaken for one hour. The filtrates analyzed for electrical conductivity using electrical conductivity meter. To determine the pH for amendments, 25 ml of deionized water was added to each ten grams of amendments and shaken for thirty minutes using the mechanical shaker. The pH reading was taken directly from the suspensions of chicken manure and chicken manure-derived biochar. At least 4 replicates were used for each analysis.
For the amendments, the organic carbon, cation exchange capacity and available phosphorous was determined using the same methods outlined in soil analysis. While for the metallic elements in amendments, the samples were weighed 10 g each and put into 250 cm3 Erlenmeyer flask. Fifty centimetre cubic of NH4Oac, pH 7.0 was added. Shaken for 30 minutes on a shaker. The suspension was filtered using number forty two filter paper, and then potassium, calcium, manganese and iron among others were measured in the filtrate.
Lead (Pb) Determination In Soil And Plants
To determine total Pb concentrations in soils, 0.05g of soil was weighed and placed in test tubes to which 5 ml 30% nitric acid (HNO3) and 1 ml hydrogen peroxide (H2O2) were added. The mixture was digested for 45 minutes using the Speedwave digester (Berghof, Germany). Thereafter filled up to 15 ml with deionized water in a 15 ml transparent bottle.
To determine Pb concentrations in the plants, the plants were oven dried for 72 hours at 70oC then ground with pestle and mortar. Pb concentrations in plant samples was determined by weighing 0.05g ground plant sample to which 5 ml 30% nitric acid (HNO3) and 1 ml hydrogen peroxide (H2O2 ) were added. The samples were then digested using the Speedwave digester (Berghof, Germany) digester for 45 minutes. Thereafter filled up to 10 ml with deionized water in a 15 ml transparent bottle.
Post-harvest soil was collected and bioavailable Pb determined in 20 ml of DTPA – TEA as described in soil analysis. In both plant and soil, Pb concentrations were analyzed using Flame Atomic absorption spectroscopy (FAAS-Z series 2010).
Statistical Analysis
All experiments were conducted with four replicates. The data were statistically analyzed using the one-way analysis of variance (ANOVA) test using Graphpad Prism software 9.0.0. Tukey’s multiple comparison tests were conducted at significance level p < 0.05 to separate the statistically significant treatment means. Pearson correlation was used to determine the relationship between soil pH and bioavailable Pb in soil. Other calculations and graphs were prepared using Microsoft Excel 2010.