Characteristics of Enzyme Activities during Phytoremediation of Cd-Contaminated Soil

Abstract

In order to study the effects of exogenous Cd on the soil enzyme activities of three herbs, a pot experiment was conducted to study the changes of soil urease, protease, catalase and phosphatase activities in different growth periods of Solanum nigrum L., Phytolacca acinose Roxb., and Bidens pilosa L. under different concentrations of Cd stress. The results showed that the content and proportion of each form of Cd were different in different periods. Compared with the control, the activities of urease, protease and catalase in the soil of three herbs decreased under 5 mg/kg and 10 mg/kg Cd stress, while the phosphatase activities increased first and then decreased. The activities of urease, protease, catalase and phosphatase were 4.24–6.84, 2.17–5.83, 2.09–2.79 and 34.57–37.25 mg/g, respectively, and the recovery degrees were 50.81–66.41%, 32.10–90.54%, 46.97–69.28% and 54.78–56.69%, respectively. After 60 days of remediation, the activities of urease, protease, catalase and phosphatase were 6.05–8.55, 2.83–9.89, 3.32–4.48 and 37.62–41.15 mg/g, respectively, and the recovery degrees were 70.19–84.57%, 41.86–161.34%, 72.35–140.44% and 58.38–63.20%, respectively. Soil enzyme activities were affected by Cd solution stress, which could be improved to a certain extent by plant self–healing, and different grass species recovered to varying degrees under various Cd solution stresses. Different soil enzymes displayed different responses to Cd stress, the inhibition of urease and phosphatase activities was temporary, and the effect of Cd concentration on soil phosphatase activity was close, and it could stimulate the activities of soil protease and catalase, and the higher the concentration of Cd solution, the greater the degree of stimulation. Principal component analysis shows that, after 60 days of repair, the best repair effect plants were Solanum nigrum L. under 5 mg/kg Cd stress and Phytolacca acinosa Roxb. under 10 mg/kg Cd stress.

1. Introduction

In recent years, soil heavy metal pollution episodes have become more common in China, posing a major threat not only to crop growth and quality, but also to people’s health. Soil heavy metal pollution research has traditionally been a priority of agricultural science or environmental science [1,2]. Cd is a heavy metal with high toxicity and mobility, and it is easily absorbed by plants or crops, it is also refractory. According to the Ministry of Land and Resources’ national soil pollution survey report from 2014, Cd is one of the main constituents of heavy metal contamination, with Cd levels above the norm in 7% of farmed land in China [3]. Cd pollution in China has reached 2105 km², accounting for 16.7% of the country’s cultivated land area [4], affecting not only soil fertility [5,6], but also soil microbial and enzyme activities [1], and a large amount of Cd will accumulate in farmland crops or plants, posing a serious health risk [7]. Liao Jie et al. [2] concluded that urease had the most significant effect on soil heavy metal pollution. The findings revealed that cadmium solution stress reduced soil urease activity, although it increased following phytoremediation, which was considerably less than in the early stages of cadmium stress (p < 0.05). The toxicity of heavy metals depends more on their morphological distribution in the soil than their total amount alone. The bioavailability, movement, transformation, and chemical cycle of heavy metals are all influenced by their speciation distribution, which results in a variety of environmental impacts [8]. Therefore, morphological analysis of elements is helpful to characterize their bioavailability and accurately evaluate ecological risk [9].

The most active component of the soil ecosystem and a major participant in the soil material and energy cycle is the soil enzyme [10,11,12]. The amount of heavy metals in soil influences the activity of soil enzymes. The change in soil enzyme activity might represent significant ecological processes in the soil, and the ecological dosage value can be employed as a soil heavy metals ecological safety threshold [12,13,14,15]. According to relevant research, the degree of inhibition of soil dehydrogenase, catalase, and urease activities varied as Cd stress concentrations increased [16,17,18,19]. Soil urease is a crucial enzyme in the organic carbon cycle and nitrogen transformation in the soil [20], and Liao, J. et al. [2] concluded that urease had the most significant effect on soil heavy metal pollution. Heavy metal toxicity can be efficiently controlled by soil organic matter. The higher its concentration, the better it protects soil enzyme activity [21,22]. Heavy metal passivation technology is currently the most widely used in China for farmland pollution control, but it does not remove heavy metals from soil, only renders them ineffective. Heavy metals are easily reactivated if the soil ecosystem is disturbed [23]. Phytoremediation is a type of chemical or physical remediation process that involves planting a specific plant that decomposes and accumulates heavy metals on heavy metal-polluted soil with the goal of healing the polluted soil by heavy metal absorption and enrichment in the soil [24]. Phytoremediation is now more widely accepted by the general public due to its low cost, ease of usage, and environmental friendliness [25]. Tang, H. et al. [26] and Pan, F. H. et al. [27]’s studies elaborated on the importance of soil extracellular enzyme indicators. As a result, the research findings can be used to not only screen typical remedial plants, but also to establish a solid theoretical foundation for the use of soil enzymes in soil quality and health assessment. Based on this, the enrichment plants Solanum nigrum L., Phytolacca acinosa Roxb., and Bidens pilosa L. were used as the research object, and the pot culture method was used to simulate the occurrence characteristics of Cd in the soil of three herbaceous plants under different Cd solution concentrations and establish the change characteristics. The recovery degree (the percentage of soil enzymes that have returned to their original values) of soil urease, protease, catalase, and phosphatase activities were analyzed in order to provide a scientific basis for phytoremediation. Hence, the major objectives of this research were to study through different cycles of phytoremediation: (1) variation characteristics of heavy metal speciation; (2) changes of soil enzyme activities; and (3) variation characteristics of plant repair rate.

2. Materials and Methods

2.1. The Source and Characteristic of Soil

The experimental soil used in the study was collected from surface layer soil (0–20 cm) in Huludao farmland (China). The physicochemical properties of uncontaminated soil were as follows: clay 19.5%, silt 58.5%, sand 22%, pH 6.98 (1:1 w/v water), organic matter 1.95 g/kg, cation exchange capacity 16.9 cmol/kg; total concentration Cd 0.15 mg/kg, urease 10.3 mg/g, protease 6.76 mg/g, catalase 5.28 mg/g, phosphatase 65.11 mg/g.

2.2. Chemicals

All the chemicals used in the present study were of analytical grade from Sinopharm Chemical Reagent Co., Ltd (Beijing, China). For heavy metal solution preparation, CdCl₂, was dissolved in deionized water to respectively prepare Cd(II). For soil enzyme activities preparation phenol, sodium hypochlorite, toluene, sulphuric acid, sodium sulphate, ninhydrin, potassium permanganate, and disodium benzene phosphate.

2.3. Experimental Design

The determination of Cd concentration in this study is according to the Soil Environmental Quality Risk Control Standard for Soil Contamination of Agricultural Land (Trial). According to the standard, when 6.5 < pH ≤ 7.5, the concentration limit for Cd is 0.3 mg/kg. The pH value of the soil used in the experiment is 6.98, so in order to facilitate the experimental operation, the concentration of Cd was selected as 5 mg/kg and 10 mg/kg. After being air-dried, the collected soil was sieved through a 2 mm sieve to ensure homogeneity. Then, the dried soil was mixed thoroughly with 0, 5, and 10 mg/kg Cd solution, the soil and Cd solution was fully stirred; and the test soil was prepared with varying Cd solution concentrations. The seeds of Solanum nigrum L., Phytolacca acinosa Roxb. and Bidens pilosa L. were collected from the field. After surface sterilized with 2% sodium hypochlorite solution for 30 min, the seeds were washed several times with sterilized distilled water to remove the remaining solvent on the surface. The seeds were then planted into a pot containing 300 g of soil at a 10 cm in bottom radius to 16 cm height. Each treatment was prepared in triplicates. During cultivation, the temperature ranged from 20 to 25 °C. The non-rhizosphere soil in one pot was collected according to 0 d (control), 30 d, and 60 d during the plant growth process.

2.4. Quality Assurance/Quality Control

Analytical data quality was guaranteed through the implementation of laboratory QA/QC protocols, including the use of standard procedures, reagent blanks, percentage of recoveries, and analysis of replicates. All reagents used were of analytical grade (certified purity > 99.9 percent). The entire laboratory was cleaned using an HNO₃ (30 percent, v/v) bath overnight, followed by repeated rinsing with double distilled water [28].

2.5. Fractions of Cd(II) and Enzyme Activity Measurement

The BCR sequential extraction procedure was applied to determine the contents of heavy metal fractions in sediment [29,30]. All enzyme activities were determined in fresh soil samples. Soil urease activity was determined by phenol sodium hypochlorite colorimetry, expressed as mg of NH₃-N in 1 g soil after 24 h of culture at 37 °C; the activity of protease was determined by glass River colorimetry, expressed in milligrams of glycine in 1 g soil after 24 h; catalase activity was determined by potassium permanganate titration, expressed in milliliters of 0.1 N potassium permanganate consumed by titrating 1 g of soil after 20 min; phosphatase activity was determined by sodium phenylene phosphate colorimetry, expressed as mg of P₂O₅ in 1 g soil after 2 h [31,32,33,34] with 3 replicates per indicator.

2.6. Statistical Analysis

Principal component analysis was as follows: the evaluation indexes were selected, including plant species, soil enzyme, and different concentrations of Cd pollution. Standardize relevant data to eliminate differences between different indicators, the dimensional and order of magnitude effects. Kaiser–Meyer–Culkin (KMO) test statistics and Bartlett sphericity test were used to judge the correlation between indicators to determine whether the original variables were suitable for factor analysis. When the KMO value is greater than 0.5, factor analysis can be performed. When the significance of the Bartlett sphericity test is <0.05, there is a correlation between the original variables, and principal component analysis can be performed. Next, the number of principal components was determined. Generally, the principal component with cumulative variance contribution rate > 75% and eigenvalue > 1 is selected. According to the result of the calculation, the expression of principal component is determined. The principal component score was calculated, and the higher the comprehensive score was, the optimal species selection for phytoremediation was indicated.

Membership function method was used to comprehensively evaluate different pollution concentrations, plant species, and soil enzyme activities; the calculated as follow:

$$X_{\left(u\right)}=\frac{X-X_{min}}{X_{max}-X_{min}}$$

(1)

$X$: the measured value of an enzyme activity index of the test value, $X_{max}$ and $X_{min}$ are the maximum and minimum values of the index, respectively.

If an index is negatively correlated with pollution concentration, it can be calculated by using the inverse membership function, the calculated as follow:

$$X_{\left(u\right)}=1-\frac{X-X_{min}}{X_{max}-X_{min}}$$

(2)

Firstly, Formulas (1) or (2) was used to calculate the membership function values of soil enzyme activity under different pollution concentrations, and the membership function values of each index were weighted and averaged. By comparing the size of the weighted average.

The recovery degree of soil enzyme activity was calculated as follows:

Degree of recovery = (enzyme activity in contaminated soil on day N÷enzyme activity in uncontaminated soil at day 0) × 100

(3)

We first used two-way analysis of variation (ANOVA) to test differences in cadmium concentration, plant species and soil enzyme activity, and the least significant difference method (LSD) multiple comparisons were used to compare the significance of differences (p < 0.05). Statistical analyses of the data were carried out using SPSS 21 for Windows (SPSS Inc., Chicago, IL, USA). The data in tables were expressed as average value and the charts were drawn using Origin Pro 2021.

3. Results

3.1. Speciation Characteristics of Cd in Soil

Under several growth cycles, the quantity and distribution of distinct Cd forms are illustrated in Figure 1. The contents of weak acid Cd and oxidized Cd in the soil planted with Solanum nigrum L. at 30 and 60 days were 0, the contents of weak acid Cd and oxidized Cd in the soil planted with Phytolacca acinosa Roxb. and Bidens pilosa L. were low, and the contents of reduced Cd and residual Cd in the soil planted with three kinds of plants were higher in the reduced and residual state than the weak acid state and oxidation state. At 30 days, the contents of reduced Cd and residual Cd in the soil planted with Solanum nigrum L., Phytolacca acinosa Roxb., and Bidens pilosa L. were 1.67, 1.64, 1.77 mg/kg and 1.64, 2.19, 1.63 mg/kg; the proportions were 50.44%, 34.90%, 47.11% and 49.56%, 46.67%, 43.33%; at 60 days; the contents of reduced Cd and residual Cd in the soil planted with Solanum nigrum L., Phytolacca acinosa Roxb. and Bidens pilosa L. were 1.48, 1.82, 1.82 mg/kg and 1.58, 2.09, 1.65 mg/kg; the proportions were 48.43%, 35.36%, 47.48% and 51.57%, 40.56%, 43.13%. When the amount and proportion of four different types of Cd were compared, the content and proportion of reduced Cd and residual Cd were significantly higher than the content and proportion of weak acid Cd and oxidized Cd. The content of Cd in three grass species with 10 mg/kg Cd solution increased the term into day of plant growth compared to the condition of 5 mg/kg Cd solution, the results showed that the content of various forms of Cd increased under Cd pollution stress, indicating that an increase in Cd solution concentration can significantly increase the content of various forms of heavy metals. The proportion of oxidized and weak acid Cd in soil increased, while the proportion of reduced and residual Cd declined. Because, using Cd, it is easy to generate a weak acid extraction state with high activity in soil as the quantity of Cd solution rises, the fraction of weak acid Cd rises.

3.2. Effect of Phytoremediation on Soil Urease Activity in Cd-Contaminated Soil

The changes in soil urease activity and plant recovery degree in different remediation cycles are shown in Figure 2. Urease activity in soil planted with Solanum nigrum L., Phytolacca acinosa Roxb., and Bidens pilosa L. decreased as the Cd pollution concentration in soil increased; before phytoremediation (0 d), urease activity in soil with Cd concentrations of 5 mg/kg and 10 mg/kg decreased by 13.11% and 18.77%, respectively, compared to the control; with the extension of planting time, soil urease activity increased under phytoremediation; after 30 days of repair, 0, 5, 10 mg/kg Cd concentration, the urease activities in the soil planted with Solanum nigrum L., Phytolacca acinosa Roxb., and Bidens pilosa L. were 4.38–6.84, 4.24–6.31, 4.95–6.51 mg/g, respectively, the degree of recovery was 50.81–66.41%, 49.19–64.58%, 56.76–63.20%; after 60 days of repair, 0, 5, 10 mg/kg Cd concentration, the urease activity in the soil planted with Solanum nigrum L., Phytolacca acinosa Roxb. and Bidens pilosa L. were 7.29–7.56, 6.50–7.87, 6.05–8.55 mg/g, respectively, the degree of recovery was 73.40–84.57%, 75.41–83.58%, 70.19–83.01%. The results showed that Cd pollution could inhibit soil enzyme activity, the soil enzyme activity is improved under the action of plant self-repair, and the recovery degree of soil enzyme activity of Phytolacca acinosa Roxb. is high and stable. By comparing the soil urease activities in the restoration process of different herbs, it can be seen that, after 30 days of restoration, three herbs were used, and there were significant differences in urease activity among soils with different Cd solution concentrations (p < 0.05), after 60 days of repair, s the urease activities of contaminated soil were varied with different Cd concentrations; after 30 days of restoration, three herbs were used, and soil urease activities were significantly less than 0 d (p < 0.05), 60 d repair period, in addition to using Solanum nigrum L. to repair the soil with Cd pollution concentration of 10 mg/kg, soil urease activities of other treatments were significantly less than 0 d (p < 0.05). Phytolacca acinosa Roxb. had the best recovery effect on soil urease activity under low concentration Cd stress; Solanum nigrum L. had the best recovery effect on soil urease activity under high concentration Cd stress. Under different Cd solution concentrations, different herb plants have different effects on different soil enzyme activities. This is related to the different remediation abilities and effects of the three herbs on Cd pollution. In the future, different herbs should be selected according to the concentration of Cd solution.

3.3. Effect of Phytoremediation on Soil Protease Activity in Cd-Contaminated Soil

The changes in soil protease activity and plant recovery degree in different remediation cycles are shown in Figure 3. The effect of three plants on protease activity in the remediation of Cd-contaminated soil is consistent with urease activity. When the Cd pollution concentration was 5 mg/kg and 10 mg/kg, the protease activity in soil was 96.75% and 90.68% of 0 mg/kg, respectively. Compared with 0 d, the three plants were in the process of remediation of Cd-contaminated soil with different concentrations, the protease activity in soil decreased first and then increased, after 30 days of repair, under the conditions of 0, 5, and 10 mg/kg Cd concentration. The protease activities in the soil of Solanum nigrum L., Phytolacca acinose Roxb., and Bidens pilosa L. were 2.17–5.83, 2.63–5.51, and 2.55–5.55 mg/g, the degree of recovery was 32.1–89.14%, 38.91–89.89%, 37.72–90.54%, respectively. After 60 days under Cd pollution conditions of 0, 5, and 10 mg/kg, the soil protease activities of Solanum nigrum L., Phytolacca acinosa Roxb. and Bidens pilosa L. were as follows: 4.24–7.62, 4.77–8.74, 2.83–9.89 mg/g, the degree of recovery was 62.72–124.31%, 70.56–142.58%, 41.86–161.34%, respectively. After 60 days of plant self-repair, soil protease activity was improved, and high concentration Cd solution could promote the soil protease activity of the three plants. Comparing the soil protease activity of planting different herbs, over a 30 day repair period, there were significant differences in protease activity among the three herbaceous plants under the concentration of 0 mg/kg and 5 mg/kg Cd solution (p < 0.05). There was no significant difference when the Cd pollution concentration was 10 mg/kg. In the 60 day remediation period, the protease activities of the three herbaceous plants were different under different Cd pollution concentrations; in the 60 day remediation period, the protease activity in soil with 0 mg/kg Cd pollution concentration was significantly less than that in 0 days (p < 0.05). There was no significant difference in soil protease between partial treatment and 0 days (p < 0.05). Under the condition of low concentration Cd stress, Solanum nigrum L. had the best recovery effect on soil protease activity; Bidens pilosa L. had the best recovery effect on soil protease activity under high concentration Cd stress.

3.4. Effect of Phytoremediation on Soil Catalase Activity in Cd-Contaminated Soil

The changes in soil catalase activity and plant recovery degree in different remediation cycles are shown in Figure 4. With the increase in Cd pollution concentration, catalase activity decreased in the soil planted with Solanum nigrum L., Phytolacca acinose Roxb., and Bidens pilosa L., when the Cd concentration is 5 mg/kg and 10 mg/kg, the catalase activity in soil was 80.68% and 60.42% of the control; with the extension of planting time, soil catalase activity increased under phytoremediation. After 30 days of repair, when the Cd concentration is 0, 5, and 10 mg/kg, the catalase activity in the soil planted with Solanum nigrum L., Phytolacca acinose Roxb., and Bidens pilosa L. was 2.09–2.62, 2.21–2.52, 2.14–2.79 mg/g, the degree of recovery was 49.62–65.52%, 46.97–69.28%, 52.84–67.08%, respectively. After 60 days of repair, when the Cd concentration is 0, 5 and 10 mg/kg, the catalase activity in the soil planted with Solanum nigrum L., Phytolacca acinose Roxb., and Bidens pilosa L. was 3.80–4.48, 3.86–4.36, 3.32–4.34 mg/g, the degree of recovery was 72.35–140.44%, 78.60–136.68%, 74.81–104.08%, respectively. According to the change characteristics of soil catalase activity in the process of three kinds of phytoremediation, Cd in soil can promote the catalase activity of three remediation plants in soil, the higher the concentration of Cd solution, the higher the catalase activity. The catalase activity of soil planted with different herbs was compared. At 30 days, there was no significant difference in catalase activity in soils with different Cd pollution concentrations. The differences in catalase activity in soils with different Cd pollution concentrations were different when the three herbs were planted at 60 days, under the condition of low-concentration Cd pollution. The effect of planting Bidens pilosa L. on soil catalase activity was significantly higher than that of the other two plants, however, at high concentration, the inverse was true, which is related to the concentration of Cd solution and stress ability. At 30 days, the catalase activity of the soil planted with the three herbs at different Cd pollution concentrations was significantly less than that at 0 days (p < 0.05). In the 60 day remediation period, the catalase activity in 10 mg/kg Cd-contaminated soil was greater than at 0 days, but did not reach a significant level. Bidens pilosa L. had the best recovery effect on soil catalase activity under low-concentration Cd stress; under the condition of high concentration Cd stress, Solanum nigrum L. had the best recovery effect on soil catalase activity.

3.5. Effect of Phytoremediation on Soil Phosphatase Activity in Cd-Contaminated Soil

The changes in soil phosphatase activity and plant recovery degree in different remediation cycles are shown in Figure 5. With the increase in Cd pollution concentration, the phosphatase activity in the soil planted with Solanum nigrum L., Phytolacca acinose Roxb., and Bidens pilosa L. firs increased and then decreased. The phosphatase activities in 0, 5, and 10 mg/kg Cd-contaminated soil was 65.11, 66.70, 63.11 mg/g, respectively, which shows that a high concentration of Cd will inhibit the activity of phosphatase in soil. The phosphatase activity in 5 mg/kg Cd-contaminated soil increased, indicating that a low concentration of Cd can promote soil phosphatase activity. Compared with 0 days, the phosphatase activity of the three grass species with different Cd solution concentrations first decreased and then increased at 60 days. Under the condition of Cd pollution of 0, 5, and 10 mg/kg, the phosphatase activities in the soil of Solanum nigrum L., Phytolacca acinose Roxb., and Bidens pilosa L. were 38.68–39.32, 38.42–40.12, 37.62–41.15 mg/g, respectively, the degree of recovery was 58.95–61.54%, 59.46–61.62%, 58.38–63.20%, respectively. After 60 days of plant self-repair, soil phosphatase activity increased, however, the effects of different concentrations of Cd on soil phosphatase activity were similar. Compared with the soil phosphatase activity of planting different herbs, at 30 and 60 days, there was no significant difference in phosphatase activity in different Cd-contaminated soils, and significantly less than 0 days (p < 0.05), which shows that although the recovery time is prolonged, the phosphatase activity is still low. Phytolacca acinosa Roxb. had the best recovery effect on soil phosphatase activity under low-concentration Cd stress; under the condition of high-concentration Cd stress, Solanum nigrum L. had the best recovery effect on soil phosphatase activity. This is related to the different repair abilities and effects of the three herbs on Cd pollution.

3.6. Correlations between Cd and Enzyme Activities Correlation Analysis

Table 1. Two-way ANOVA analysis of variance between Cd pollution concentration and plant species on mineralization indexes.

Enzymatic Activity	Variance Source	Sum of Squares	Freedom	Mean Square	F Value	Significance
urease	Cd solution concentration	9.220	2	4.610	57.072	0.000
	plant species	0.298	2	0.149	1.844	0.187
	interaction effect	5.087	4	1.272	15.746	0.000
	error	1.454	18	0.081
	the sum	1463.955	27
protease	Cd solution concentration	105.614	2	52.807	2024.116	0.000
	plant species	1.992	2	0.996	38.182	0.000
	interaction effect	14.204	4	3.551	136.109	0.000
	error	0.470	18	0.026
	the sum	1136.488	27
catalase	Cd solution concentration	0.030	2	0.015	0.607	0.556
	plant species	0.297	2	0.148	6.037	0.010
	interaction effect	2.836	4	0.709	28.836	0.000
	error	0.443	18	0.025
	the sum	437.528	27
phosphatase	Cd solution concentration	13.022	2	6.511	1.512	0.247
	plant species	0.949	2	0.474	0.110	0.896
	interaction effect	11.370	4	2.842	0.660	0.628
	error	77.501	18	4.306
	the sum	41580.363	27

shows that Cd contamination has a substantial impact on the activities of urease and protease in soil (p < 0.05), but no significant effect on catalase and phosphatase activities. Protease and catalase activities were significantly affected by plant species (p < 0.05), whereas urease and phosphatase were not. The interaction exhibited a statistically significant effect on the activities of urease, protease, and catalase (p < 0.05), but not on phosphatase activity.

The amount of each form of Cd is adversely connected with soil enzyme activity, as shown in

Table 2. Correlation coefficients of heavy metal forms and enzyme activities.

Fraction	Urease	Protease	Catalase	Phosphatase
weakly acidic Cd	−0.57 **	−0.85 **	−0.62 **	−0.35
reduced Cd	−0.68 **	−0.89 **	−0.70 **	−0.29
oxidized Cd	−0.47 *	−0.81 **	−0.52 *	−0.32
residual Cd	−0.69 **	−0.92 **	−0.76 **	−0.43

Notes: * p < 0.05, ** p < 0.01

. Urease activity was also significantly or extremely significantly negatively correlated with the content of various forms of Cd. There was a significant or extremely significant negative correlation between protease activity and the content of various forms of Cd, a significant or extremely significant negative correlation between catalase activity and the content of various forms of Cd, and a negative correlation between phosphatase activity and the content of various forms of Cd, but this did not reach a significant level.

3.7. Comprehensive Evaluation of the Effects of Plant Species on Soil Enzyme Activities under Cd Stress

The membership function approach and principal component analysis were used to assess the impact of Cd exposure on soil enzyme activity.

Table 3. Membership degree and restoration comprehensive score of each index under different plant species.

Plant Species	Concentration (mg·kg−1)	Urease	Protease	Catalase	Phosphatase	Comprehensive Score	Sort
Solanum nigrum L.	5	0.107	0.216	0.231	0.059	0.253	2
	10	0.134	0.229	0.138	0.067	0.185	3
Phytolacca acinosa Roxb.	5	0.091	0.393	0.308	0.109	0.312	1
	10	0.207	0.183	0.089	0.144	0.135	5
Bidens pilosa L.	5	0.064	0.541	0.064	0.074	0.131	6
	10	0.132	0.155	0.093	0.064	0.136	4

shows the influence of plant species on the remediation of soil enzyme activity under different Cd concentration stress circumstances. Phytolacca acinosa Roxb. has the best repair effect under low concentration; Solanum nigrum L. has the best repair effect under high concentration. The repair effect of Solanum nigrum L. is relatively good under different Cd solution concentrations, but there was a significant difference in the repair effect of Phytolacca acinosa Roxb. on high- and low-concentration Cd solution, indicating that Phytolacca acinosa Roxb. is sensitive to the repair ability and effect under Cd solution stress, which can easily cause a large degree of fluctuation.

4. Discussion

Cd, a heavy metal that is widely dispersed in soil and pollutes soil and water resources, is a major pollutant. Heavy metal-polluted soil has a negative impact on soil enzyme activity [35]. Plants may absorb and fix heavy metals, and changes in soil enzyme activity during remediation can better represent heavy metal concentrations in soil, which is crucial for soil heavy metal monitoring [36]. Soil urease is a crucial enzyme in the organic carbon cycle and nitrogen transformation in the soil [37]. The major enzyme engaged in the soil nitrogen cycle is soil protease, which is enhanced by microbial activity, plant root secretion, and the degradation of animal and plant residues [38]. The soil protease activity of the three plants declined initially and subsequently increased with the extension of repair time under 5 mg/kg and 10 mg/kg Cd stress, although the soil protease activity was less than at 0 days (p < 0.05), showing that Cd stress inhibits soil protease activity. Soil catalase is a major catalytic enzyme in soil that can transform waste products produced by soil metabolism into innocuous or less hazardous substances [33]. The activity of soil catalase declined first, then increased in this study, which has a similar temporal trend to that of protease, but the rising range is wide. Under 5 mg/kg and 10 mg/kg Cd stress, the catalase activity of the three plants reduced after 30 days of repair, which is attributable to the Cd-induced mortality of certain microorganisms, as reported by Chen Zhixue et al. [39], and Chang Haiwei et al. [40]. However, after 60 days of restoration, soil catalase activity increased, indicating that soil enzyme activity improved after a long period of plant self-healing. Soil phosphatase is crucial for soil phosphorus cycling because it may hydrolyze complex organic phosphorus into inorganic phosphorus, hence alleviating soil phosphorus restriction. Its activity is closely related to the amount of phosphorus in the soil [41]. Cd stress inhibits soil phosphatase activity, and according to the results of this study, Cd pollution has different effects on the soil enzyme activities of three herbaceous plants, and most of them are significantly less than 0 days soil enzyme activities (p < 0.05) [42]. The reason for this phenomenon is that Cd ions combine with the active part of enzyme molecules to produce a more stable complex, while the soil microbial environment polluted by heavy metals is poor, it is not conducive to the reproduction and survival of soil microorganisms, reduces soil enzyme synthesis, and then inhibits the activity of soil enzymes [43]. The change in trends of soil enzyme activities under Cd pollution stress were different in the remediation process of three herbs after 60 days of plant self-repair. Different soil enzyme activities have different responses to Cd pollution stress, which may be related to reactive oxygen free radicals. Different soil enzymes have different effects on pollutants. For example, urease in soil is related to urease transformation [44], acid phosphatase decomposition is related to the transformation of organic phosphorus in soil [45], and invertase is related to the transformation of organic matter in soil. Therefore, based on the findings of the most recent research, if Cd contamination is encountered, we may choose the right plant species based on the pollution concentration, giving phytoremediation of heavy metal-contaminated sites a wider application prospect.

5. Conclusions

• The content of various types of Cd rose as Cd stress concentration increased, with the proportion of weak acid Cd and oxidized Cd increasing and the amount of reduced Cd and residual Cd decreasing.
• The effects of Cd stress on soil enzyme activity differ depending on the kind of enzyme and the level of pollution. During the restoration of three herbs, the activities of soil urease, protease, and catalase reduced as Cd pollution levels increased, while phosphatase activity first increased and then dropped. Cd stress inhibits the activities of urease, protease, and catalase, while a low Cd concentration promotes the activity of phosphatase.
• The enzyme activities of the soil planted with Bidens pilosa L., Phytolacca acinosa Roxb., and Solanum nigrum L. might be greatly boosted by extending the repair period of the three herbs. After a thorough examination, the promoting enzyme activity of Solanum nigrum L. was found to be the best under high-concentration Cd stress, while the promoting enzyme activity of Phytolacca acinosa Roxb. was found to be the best under low-concentration Cd stress.