1. Introduction
Contamination of the environment with heavy metals has become one of the biggest health concerns all over the world due to their persistence in the environment and accumulation in the food chain posing significant threats to human health [
1,
2]. The major pathway of human exposure to heavy metals is soil to plant transfer where vegetables take up heavy metals by absorbing them from contaminated soils [
3]. Heavy metal contamination of soil may occur due to anthropogenic activities through irrigation with contaminated water, the addition of fertilizers and metal-based pesticides, emissions from the metallurgical industry, transportation, and harvesting process [
4,
5]. Among the most common heavy metal contaminants, lead (Pb) has been reported as a major concern due to its high stability in soil, and accumulation in plants and animals, and it is considered the second most dangerous hazardous substance on the priority list of the U.S. Environmental Protection Agency [
6]. It is estimated that approximately half of the human Pb intake is through food, with around half originating from plants [
7]. The toxic level of Pb in plants inhibits germination, suppresses growth parameters, reduces the rate of photosynthesis, and alters the levels of photosynthetic pigments, transpiration, gaseous exchange in leaves, and total chlorophyll production [
8,
9]. It is well known that Pb is highly toxic for humans due to its interference with several biochemical processes, contributing to oxidative stress [
10,
11,
12]. Because of its high toxicity, the concentrations of Pb in soil and vegetables are restricted by legislation. The European Union has set standards for Pb at 0.1 mg kg
−1 (f.w.) for fruits and roots and 0.3 mg kg
−1 (f.w.) for leafy greens [
13]. So far, there are no national health-based standards for Pb in vegetables, fruits, or other staple food crops in the United States [
14], although FDA monitors and regulates Pb concentrations in foods and in consumer products.
Previous research has shown that the amount of phytoavailable heavy metal forms in soil and the level of their accumulation in plants depend on soil properties such as: pH, organic matter content, redox potential, cation exchange capacity, and soil texture [
15], as well as plant species and root system, growth stage, type of metal, environmental conditions, and agricultural practices [
16].
Among the remediation solutions for Pb-polluted soils to reduce the mobility and phytoavailability of this metal, the application of amendments has gained much attention in recent years as an environmentally friendly and low-cost agricultural management practice [
17]. Codling [
18] reported that the application of phosphorus (P) and iron (Fe) plus phosphorus as amendments on soils contaminated with Pb arsenate increased water extractable Pb concentration making it less accessible to plants. In another study, Ngole [
19] found that Pb bioavailability for carrot plants in a sludge-amended soil decreased slightly with an increase in the sludge amendment rate. Recently, Guo et al. [
20] reported that dolomite, slaked lime, and limestone applied as amendments to Pb-contaminated soil, significantly reduced Pb content in rice plants. In addition, amendments such as biochar, slag, and ferrous manganese can also successfully reduce the toxicity, leachability, and mobility of Pb in the environment [
21].
Some agricultural soils in Nova Scotia, Canada, have high levels of Pb, due to the historic application of PbHAsO
4 insecticide in apple orchards several decades ago [
22]. Pb arsenate was the most extensively used insecticide as a foliar spray to control codling moth in tree fruit orchards in countries throughout the world, including the USA, Canada, Australia, New Zealand, England, and France [
23]. It remained the preferred insecticide for codling moth control because of its high efficacy and lower phytotoxicity and it was being applied in Nova Scotia as late as 1981 [
24] until it was officially banned in 1988 [
25].
Carrot is a major specialty cash crop in Nova Scotia. Previous studies have shown that carrots, just like other vegetables, take up Pb from contaminated soils by accumulating this metal in their tissues [
26,
27,
28]. Chisolm [
22] reported that Pb concentration in carrots grown in a lead arsenate-contaminated soil exceeded the Canadian tolerance of 2.0 ppm in fresh vegetables. The use of phosphate in Pb immobilization from water or soil is an accepted technique. Furthermore, amendments that contain P can transform the Pb fractions in soil, from highly available to bounded forms, such as pyromorphite Pb5(PO4)3X where X=F, Cl, Br, OH. These Pb compounds are stable under a wide range of pH and Eh. Numerous phosphate materials of natural or synthetic origin have been used to immobilize Pb: apatite and hydroxyapatite, rock phosphate, monoammonium phosphate, diammonium phosphate, biosolids, etc. [
29].
The hypothesis in this study was that the application of different organic and inorganic amendments would bind Pb to soil and make it unavailable to plants cultivated under a lead arsenate-polluted soil from Canning, Nova Scotia.
Therefore, the objective of this study was to evaluate the potential for the immobilization of Pb in the soil by applying organic (sludge, biocompost, yard compost, and peat) and inorganic (bonemeal, zeolite, lime, and wood ash) amendments, in combination with diammonium phosphate (DAP) under greenhouse conditions. The Pb availability was assessed using carrot plants.
2. Materials and Methods
2.1. Greenhouse Experiment
The experiment was conducted in the Cox greenhouse of the Faculty of Agriculture of Dalhousie University (formerly Nova Scotia Agricultural College) under natural daylight with day temperatures of 22 to 25 °C and night temperatures of 18–19 °C. The soil used in this study was a sandy loam with 47% sand, 48.2% silt, and 4.8% clay, pH 6.2, and a cation exchange capacity of 17.9. The total Pb concentration of the soil was 109 mg/kg. The soil was collected from the surface layer (0–20 cm) in Canning, NS, Canada, that has a lead arsenate, PbHAsO4 application history.
Plastic pots (20-cm diameter and 15-cm high, Classic 600; Nursery Supplies, Inc., Fairless Hills, PA, USA) were filled with 2 kg of air-dried soil each.
Certified seeds of carrots (Daucus carota L. Red Core Chantenay) were direct-seeded into the pots. After 10 days, seedlings were thinned to 6 per pot.
2.2. Experimental Design
A factorial experimental design with 3 replications was used in this experiment. Plants were fertilized with potassium (K) as potassium chloride (2 g/kg) and nitrogen (N) as ammonium nitrate (1.25 g/kg) through incorporation in the soil.
Treatments were represented by 3 application rates of diammonium phosphate (DAP): 0 (zero); low (0.25 g/kg), and high (1.25 g/kg), calculated to represent zero, low and high fertilizer application rates under field conditions; and soil amendments (organic and inorganic), also added at zero, low and high application rates (
Table 1). The zero rates of application represents untreated pots in which no treatments (neither amendments nor fertilizers) were added.
The sludge and biocompost were obtained from Fundy Compost (
http://www.fundycompost.com/index.php accessed on 10 June 2021) in Nova Scotia. The sludge originated from the city of Halifax, NS, Canada. The low rate of sludge was selected based on crop N requirements (130 kg ha
−1), and assuming a 25% availability of N in composts. The high rate was set at four times the amount of N required.
The yard compost was supplied by Peter Peill of Minas Seed Ltd. (Canning, NS, Canada). The low rate for each of the composts was decided based on crop N requirements (130 kg ha−1), and assuming a 15% availability of N in composts. The high rate was set at four times the amount of N required.
Sphagnum peat moss (ASB Greenworld) was applied to the soil on a 50% (low) and 100% (high) v/v basis. The calculated volume of soil assumes an incorporating depth of 0.3 m over the plot area.
The bonemeal and zeolite were purchased from Digby O & E farms Ltd. (Drayton, ON, Canada). The rates of bonemeal and zeolite were applied to add 0.5% w/w and 5% w/w, according to the recommendations.
The lime (Easy Spread dolomitic limestone) was supplied by Mosher Limestone Co., Ltd. (Upper Musquodoboit, NS, Canada). The rates of lime were applied to change the soil pH from 6.2 to 6.5 (low) and to 7.0 (high) based on the calculation in [
30].
The wood ash was supplied by the Faculty of Agriculture power plant facilities. Since the effectiveness of wood ash in changing the soil pH was assumed to be half of that of lime, the low and high rates of wood ash were doubled that of the lime rates.
The elemental compositions of the immobilization amendments were determined as described previously [
31,
32] (
Table 2 and
Table 3).
2.3. Determination of Pb in Plant Samples
The plants were harvested 78 days after establishment, when carrots reached a marketable stage. The carrots were washed thoroughly to remove all soil particles, dried in a drying oven at 70 °C for 72 h, until a constant weight, and the dry weight was recorded. The concentration of Pb in tissue and soil samples was determined as described previously [
31,
32]. Briefly, heavy metal concentrations in tissue, soil amendments, and soil samples were determined by an inductively coupled argon plasma spectrometer (ICAP) model 61 (Thermo Jarrell Ash, Franklin, MA, USA) following nitric acid digestion as described previously [
31,
32]. Because of the expected relatively low concentrations of heavy metals in tissue, soil, and soil amendment samples, larger samples of 4 g were digested for 8 h in 250-mL digestion tubes. The available Pb concentration in soil was determined using extraction with 1 M Mg(NO
3)
2 to extract the exchangeable (bioavailable) Pb fraction [
31,
32].
2.4. Determination of Plant Nutrients in Soil Samples
The concentration of plant-available nutrients in soil samples was determined on ICAP by the Nova Scotia Soil Testing Laboratory following the Mehlich 3 extraction, which is a standard procedure in Atlantic Canada.
2.5. Statistical Analysis
The main and interaction effects of Amendment (Bio-compost, Bone Meal, Lime, Peat, Sludge, Wood ash, Yard-compost and Zeolite), Application Rate (Zero, Low and High) and diammonium phosphate (DAP: No DAP [Zero DAP], Low DAP and High DAP) on carrot dry yield, and the concentrations of Pb and P in the tissue (Pb TC, P TC), Pb and P tissue uptake (Pb TU, P TU), and soil available Pb and P (Pb SA, P SA) were determined by conducting Analysis of Variance (ANOVA) of an 8 × 3 × 3 factorial design with 3 replications.
The validity of normal distribution and constant variance assumptions on the error terms were verified by examining the residuals as described in Montgomery [
33,
34]. Some of the response variables required square root transformation; however, the means shown in the tables are back-transformed to the original scale. Independence assumption on the error terms was ensured by the proper randomization performed during the experiment.
All analyses were completed using the Mixed Procedure of SAS 9.4 (SAS Institute Inc., Cary, NC, USA). For significant (
p-value < 0.05) effects, multiple means comparisons were completed by comparing the least squares means of the corresponding treatment combinations. Letter groupings were generated using the Tukey-Kramer method at a 5% level of significance for the main and two-way interaction effects, but for the three-way interaction effect, a 1% level of significance was used to reduce the potential overinflation of Type II experimentwise error rate due to the large number (72) of treatment combinations being compared.
Figure 1,
Figure 2 and
Figure 3 were produced using Minitab 21 software (State College, PA, USA).