Next Article in Journal
Application of In Vitro Techniques for Elimination of Plum Pox Virus (PPV) and Apple Chlorotic Leaf Spot Virus (ACLSV) in Stone Fruits
Previous Article in Journal
Broccoli Cultivation Under Different Sources and Rates of Specialty Phosphorus Fertilizers in the Brazilian Cerrado
Previous Article in Special Issue
Response of Shoot Growth to Ecological Factors Highlights a Synergistic Relationship Between Yield and Catechin Accumulation in Tea Plant (Camellia sinensis L.)
 
 
Search for Articles:
Title / Keyword
Author / Affiliation / Email
Journal
Article Type
 
 
Advanced Search
 
Section
Special Issue
Volume
Issue
Number
Page
 
Journals
Horticulturae
Volume 11
Issue 6
10.3390/horticulturae11060632
horticulturae-logo
Submit to this Journal Review for this Journal Propose a Special Issue
► Article Menu

Article Menu

share Share announcement Help format_quote Cite question_answer Discuss in SciProfiles
first_page
settings
Order Article Reprints
Font Type:
Arial Georgia Verdana
Font Size:
Aa Aa Aa
Line Spacing:
Column Width:
Background:
Article

Phosphoric Acid and Magnesium Chloride Composite-Modified Biochar Improved Pakchoi Growth by Reducing Pb and Cd Accumulation and Altering Soil Properties and Microbial Communities

by
Xuejie Dong
,
Haojie Xu
,
Yanfang Ren
*,
Dongming Lin
,
Ke Li
and
Junyu He
*
School of Environmental Science and Engineering, Changzhou University, Changzhou 213164, China
*
Authors to whom correspondence should be addressed.
Horticulturae 2025, 11(6), 632; https://doi.org/10.3390/horticulturae11060632
Submission received: 22 April 2025 / Revised: 29 May 2025 / Accepted: 3 June 2025 / Published: 4 June 2025
(This article belongs to the Special Issue Biotic and Abiotic Stress Responses of Horticultural Plants)
Browse Figures
Versions Notes

Abstract

:
Soil heavy-metal pollution is one of the most serious environmental issues in the world. There is an urgent need to develop feasible strategies for the remediation of polluted soil. Biochar has great potential to reduce heavy metal phytotoxicity and promote plant growth, but its mechanisms are still unclear. In this study, phosphoric acid and magnesium composite-modified tea branch biochar (PMB) was prepared and characterized. The effects of PMB at 5% addition on pakchoi growth, Cd/Pb accumulation and subcellular distribution in pakchoi, soil physicochemical characteristics and enzyme activities, Cd/Pb bioavailability, bacterial community structure, and diversity in Cd/Pb co-contaminated soils was investigated by a pot experiment. The results showed that PMB significantly alleviated the phytotoxicity of Cd and Pb. The application of PMB effectively increased the plant height and biomass and Cd and Pb proportion in the cell wall, while reducing Cd and Pb accumulation and their distribution in cytoplasm and organelles in pakchoi plants. PMB significantly improved the activities of urease, invertase, and catalase and reduced the available Cd and Pb contents in soil. Moreover, PMB changed the structure and diversity of the soil bacterial community. The relative abundance of several beneficial microbial phyla, including Acidobacteriota, Bacteroidota, Actinobacteriota, and Gemmatimonadota, increased by 13.81%, 19.02%, 68.09%, and 34.79%, respectively. The Shannon and Chao1 index also increased significantly. This study provides an effective strategy for simultaneous Cd and Pb immobilization in soil, promoting plant growth and inhibiting heavy metal accumulation in vegetables, which highlights the application of PMB in sustainable agro-ecosystems.

1. Introduction

With the rapid development of industry and agriculture, heavy metals have entered the environment, causing serious pollution in soils due to metal smelting and processing, chemical wastewater discharge, mining, etc. [1]. Among them, cadmium (Cd) and lead (Pb) are the most common and widespread contaminants [2]. According to the Bulletin of the National Soil Pollution Survey, heavy metals accounted for 82.8% of the total amount of inorganic pollutants, while soil Cd and Pb contents exceeded the standard by 7.0% and 1.5%, respectively. Notably, Cd and Pb combined pollution occurred frequently. Both Cd and Pb have negative impact on plant growth, cell division, and metabolic processes, resulting in declines in crop yield and quality. Long-term exposure to Cd and Pb can cause a variety of health problems in humans, such as cancer, anemia, kidney disorders, neurasthenia, etc. [3,4]. Therefore, it is imperative to explore promising strategies for high-efficiency remediation of soil simultaneously polluted with Pb and Cd.
Cd and Pb in soil are impossible to biodegrade. It is important to change their forms to reduce their ability to harm living organisms and their movement in the environment. Immobilization and stabilization are effective and easy-to-operate methods for remediating soil contaminated with heavy metals and reducing their toxic effects on living organisms [5]. Currently, various substances such as natural minerals, aggregates, and organic fertilizer are used for soil restoration. Due to its alkalinity, rich carbon content, functional groups, porosity, and high cation-exchange capacity, biochar is used in soil remediation, stabilizing heavy metals and improving soil fertility [6]. The alkaline ions of biochar can alter the forms of metals by changing soil pH, thereby decreasing the risk of Cd toxicity in the environment [7]. Biochar can also significantly reduce the mobility of Cd in the soil due to its good physical structure and rich surface functional groups [8]. Xu et al. [9] discovered that the addition of rice straw biochar significantly increased the pH value and cation exchange capacity of paddy soil and reduced the availability of Cd in rhizosphere soil. Coconut shell biochar could elevate soil pH and enzyme activity, reduce Cd bioavailability, and promote the germination of spinach [10]. Dewi et al. [11] reported that the addition of biochar increased the profit of the integrated agricultural system of soybeans and rice in field experiments by 10.1%. The application of biochar and its aging in the field both remarkably increased the immobilization of heavy metals in a field after 3 years of remediation [12]. However, the immobilization effect of pristine biochar on heavy-metal-polluted soil, especially for combined pollution, is far from satisfactory. In order to improve the repair effect, biochar modification has received increasing attention to modify the properties of biochar, which may contribute to its excellent immobilization behavior for heavy metals [13]. Recently, phosphorus (P)-modified biochar was considered as an efficient and low-cost way in soil remediation [14,15]. Phosphoric acid (H3PO4) with low corrosivity and environmental harmfulness is often used to modify biochar with the formation of functional groups, stable phosphorus complexes, and more micropores [16,17], which are suitable for HM immobilization in soil via metal-P precipitation and complexation [18]. For instance, it was reported that H3PO4-functionalized biochar showed excellent immobilization capacity for Cu(II) and Cd(II) due to highly improved surface pore structures and oxygen-containing groups [19]. P-rich biochar has good performance in Cd immobilization by improving soil physicochemical and biological properties [18,20]. Moreover, P-modified biochar can also provide soil with additional P fertilizer necessary for plant growth and promote plant growth [13,21]. Ahmad et al. [22] showed that 3% P-loaded biochar significantly increased the availability of soil P and obviously enhanced plant growth. Biochar modified by magnesium (Mg) ions can also significantly enhance its properties, including functional groups, cation exchange capacity, and pore structures [23,24], and thus can provide more adsorption sites for heavy metals, thereby reducing the available heavy metal content in the soil and increasing crop biomass [25]. Many studies have shown that combined modified biochar has a greater ability to immobilize heavy metals and inhibit their absorption by crops. Wang et al. [26] prepared EDTA-functionalized Mg/Al hydroxide-modified corn stover biochar, which introduced more functional groups and increased the chelating sites and ion exchange with Pb(II) and Cd(II). To date, the effects and mechanisms of P/Mg modified biochar on the sorption and immobilization of heavy metals in soil are still lacking.
Soil microorganisms play a key role in the transformation and translocation of nutrients from soil to plants, and the microbial community structure is often used as an indicator of soil quality. The porous structure of biochar could create a conductive environment for beneficial soil microorganisms, heightening microbial diversity and activity, thereby fortifying heavy metal passivation in soil [21]. It was reported that bone biochar increased microbial biomass by up to 66%, altered the soil microbial community and promoted the abundance of taxa that promote soil repair, including Actinomycetes, Firmicutes, and Proteobacteria [27]. The application of BC-Fe-S treatment also induced an enhancement with most bacterial abundance, including Ellin6067, Blastococcus, Pseudolabrys, and Ramlibacter, etc., and most of these bacterial strains were related to heavy metal resistance, biotransformation, and biosorption [28]. Particularly, Ellin6067 was found to contain genes resistant to cadmium, whereas Blastococcus had stress resistance which could grow vigorously under cadmium stress [29]. Moreover, these beneficial bacteria could not only facilitate heavy metal immobilization but also alter nitrogen and iron biogeochemical transformation in contaminated soil [30].
Tea branches have a large output in China. About 1 million tons of pruned tea branches (TBs) are produced annually. Currently, most of the TB waste is burned or discarded, which not only pollutes the environment but also wastes a large amount of potential biomass resources, presenting both a waste management challenge and an opportunity for value-added remediation materials [31]. Therefore, it is very important to find a suitable method to recycle and reuse tea branch waste. The tea branch is a cost-effective and lignocellulosic-rich material that can be the precursor of biochar and further applied in environmental rehabilitation. Our previous research showed tea branch biochar was porous and had a high specific surface area structure, which could effectively absorb heavy metals [31,32]. However, to the best of our knowledge, there have been few reports about its role and mechanisms in soil properties and heavy metal bioavailability under composite polluted soil. Pakchoi, rich in minerals and vitamins, is the most consumed vegetable in China. However, it is at high risk of heavy metal exposure [33]. Due to the frequent consumption of pakchoi in China, combined Cd and Pb pollution in soil has caused a decline in its yield [1]. Further, the accumulated heavy metals are easily transferred to humans through the food chain. Therefore, it is important to reduce Cd and Pb concentrations in pakchoi. We hypothesized that heavy metal bioavailability and pakchoi growth would probably be dominated by phosphoric acid and magnesium chloride composite-modified tea branch biochar (PMB). Therefore, the current study aimed to (i) examine the impact of PMB on the growth, accumulation, and subcellular distribution of Cd and Pb in pakchoi in Cd- and Pb-contaminated soil; (ii) explore the effect of PMB on soil physicochemical characteristics and enzymatic activity; and (iii) elucidate the response of soil bacterial community structure to PMB application. The application of PMB to heavy-metal-contaminated soil could provide a new way to increase the yield of pakchoi and guarantee food safety.

2. Materials and Methods

2.1. Soil Collection and Biochar Preparation

The soil in this experiment was obtained from the 0–20 cm surface layer of a farm located in Wujin District, Changzhou City, Jiangsu Province, China. The soil was classified as Luvisols with 42.5% clay, 39.2% silt, and 18.3% sand according to the World Reference Base classification. The basic physicochemical properties of the soil are listed in Table 1. After removing plant residues, the collected soil was air-dried at ambient temperature (20–25 °C) for one week and thereafter sieved through a mesh (2 mm) for further utilization. In the preparation of Cd- and Pb-contaminated soil, Cd2+ (CdCl2·2.5H2O) and Pb2+ (Pb(NO3)2) solutions were added to soil sample to prepare simulated contaminated soil (5 mg/kg Cd2+ and 250 mg/kg Pb2+), which was determined based on China’s Soil Environmental Quality Risk Control Standard (GB 15618-2018) [34] and the current pollution situation in farmland surrounding industrial areas in China [35]. Polluted soil was incubated for 35 days, and deionized water was regularly added to the polluted soil during the aging process to ensure that the soil water holding capacity remained at 70% of the maximum field water holding capacity. Then, the soil was allowed to dry naturally and ground through a 2 mm nylon sieve for the pot experiment.
The PMB was prepared according to Xu et al. [35]. Specifically, tea branch (TB) powder was mixed with a H3PO4 solution in the ratio of 1:2 (g:mL) for 12 h at room temperature. The mixture was pyrolyzed at 600 °C for 2 h in a tubular furnace (OTF-1200X, Kejing, Hefei, China) with a nitrogen gas flow rate of 60 mL/min. Then, it was washed with deionized water to neutral and dried at 80 °C. The H3PO4-modified TB biochar was further mixed with magnesium chloride solution (1:12, g:mL) for 24 h. After washing with deionized water, the material (named PMB) was dried, sieved through a 100-mesh sieve, and stored in a sealed container for further use. The characteristics of the PMB were analyzed using standard techniques (Figure S1).

2.2. Pot Experiment

According to the literature, the common amount of biochar added to soil is 1~5%. In the actual soil remediation process, only an amount of biochar less than 5% prevented heavy metals from negatively affecting soil [36]. Therefore, 5% of PMB was added in this study. Four treatments were included as follows: Control (CK), 5% PMB (PMB), 250 mg/kg Pb + 5 mg/kg Cd (Pb + Cd), and 250 mg/kg Pb + 5 mg/kg Cd + 5% PMB (Pb + Cd + PMB). The PMB prepared above was applied to the well-aged, contaminated soil, stirred, mixed with the soil evenly, and then incubated at room temperature for 20 days. During the soil incubation process, the soil maintained 70% of the field water holding capacity. Then, the selected seeds of pakchoi (Brassica rapa subsp. chinensis) were sown and the pots were irrigated with distilled water to maintain about 70% of the soil water holding capacity. The plant was cultured in a control environmental room with 14 h day at 25 °C and 10 h night at 18 °C and a light intensity of 450 μmol/m/s. Hoagland’s nutrient solution replaced distilled water every seven days until harvest. Each treatment consisted of three replications, with each replication including six pots. After 42 days, the pakchoi samples were collected, cleaned, and dried for subsequent analysis. At the same time, the fresh rhizosphere soils were collected and kept in a −80 °C refrigerator, which was divided into two parts: one part was used for the analysis of physical and chemical properties, enzyme activity, and available Pb and Cd contents; while the other part was used to analyze bacterial community structure diversity.

2.3. Growth Parameters and Pb and Cd Contents in Pakchoi

After measuring plant height and root length, the dried weight of both shoots and roots were determined using a milligram balance (INESA, Shanghai, China). Then, the dry samples were ground and approximately 0.05 g of powder was digested using an HNO3-H2O2 system at 150 °C [37]. Cd and Pb contents were determined via AAS (AA-300, PerkinElmer, Waltham, MA, USA). The translocation factor (TF) was determined as the ratio of the Cd/Pb concentration in shoots to that in roots.

2.4. Pb and Cd Subcellular Distribution in Pakchoi

The subcellular distribution of Cd and Pb in pakchoi was determined by differential centrifugation [38]. Fresh roots and leaves (0.5 g) were ground in an extraction buffer (5 mmol/L, pH 7.5) containing 250 mmol/L sucrose, 50 mmol/L trimethylol aminomethane-hydrochloride, 1.00 mmol/L dithithreitol, and 5 mmol/L ascorbic acid for 10 min, and centrifuged at 3000× g for 15 min. The sedimentation was the cell wall component (T1). The supernatant was centrifuged at 15000× g for 30 min. The precipitate was the organelle component (T2), and the supernatant was the cell solute component (T3). The contents of Cd and Pb in each part were determined using AAS (AA-300, PerkinElmer, Waltham, MA, USA).

2.5. Determination of Soil Properties, Enzyme Activities, and Available Content of Heavy Metal

Soil pH was measured with a pH meter at soil-to-deionized-water ratios of 1:2.5 following 30 min of agitation [15]. The content of soil organic matter (SOM) was estimated based on soil organic carbon (SOC) and a constant (1.724). The content of soil organic carbon (SOC) was measured using the potassium dichromate oxidation method, wherein soil samples were digested with a mixture of K2Cr2O7 and concentrated H2SO4, followed by titration with FeSO4 [15]. The available phosphorus (AP) content was determined using the molybdenum–antimony colorimetric method following sodium bicarbonate extraction [39]. The available potassium (AK) content was measured by flame photometry following NH4OAC (pH 7.0) extraction [39]. The alkaline hydrolysis of nitrogen (AHN) content was determined using the alkali-diffusion technique [39]. Soil urease activity (URA) was measured by an indophenol colorimeter at 630 nm [19]. Catalase activity (CAT) was determined via potassium permanganate titration [14]. Invertase activity (INV) was determined by 3,5-dinitrosalicylic acid colorimetry at 540 nm [14]. The contents of available Cd and Pb were determined by the DTPA extraction method. The chemical speciation of Cd and Pb in the soil sample was conducted using the BCR method [40].

2.6. Microbial Community Analysis

Total genetic DNA from soil samples was extracted using the E.Z.N.A™ Mag-Bind Soil DNA Kit (Omega Bio-tek, M5635-02, Norcross, GA, USA). The V3–V4 region of the soil 16S rRNA gene was amplified using PCR and universal primers 341F (CCTACGGGNGGCWGCAG) and 805R (GACTACHVGGGTATCTAATCC). After amplification, Hieff NGS™DNA Selection Beads (Yeasen Biotechnology, 10105ES03, Shanghai, China) were used to purify the amplified products by removing free primers and primer dimers. Purified PCR products of all samples were sequenced in the Illumina MiSeq system (Illumina, San Diego, CA, USA) to analyze bacterial community diversity.

2.7. Data Analysis

Data were expressed as mean ± standard deviation (SD) (n = 3). The differences among treatments were analyzed using a one-way ANOVA and the LSD test (p < 0.05) via the SPSS 20.0 software. The graphs were generated with Origin 2018. Relative abundance (>1%) was used to compare bacterial community composition, and the top 10 phyla were examined using Origin 2018. Alpha diversity indicators, including the Shannon and Chao1 indices, were calculated to estimate the complexity of species variety in a sample using Mothur software (version 1.48.0). Beta diversity was analyzed to estimate the similarity and difference of the microbial community, and a principal coordinate analysis (PCA) was performed on both weighted and unweighted UniFrac distances using Origin 2018. The LEFse based of OTUs was performed using R software (version 4.3.1).

3. Results

3.1. PMB Characterization

The physicochemical properties of the PMB are presented in Table S1. The PMB was alkaline, with a pH of 9.17. The main elements in the PMB were C, O, P, and Mg. Its BET surface area was 442.23 m2/g, and the pore volume was 0.45 cm3/g. The FTIR spectrum of the PMB exhibited several functional groups (Figure S1a), including -OH (3337 cm−1), C-H (2852 and 903 cm−1), -COOH/C=H (1620 cm−1), and C=O/O-C=O (1395 cm−1), which would favor the stabilization of heavy metals [31]. Further, peaks at 1169 cm−1 (P=O/P-O-C/P=OOH), 1068 cm−1 (PO43−), and 620 cm−1 (Mg-O) were observed, indicating that P and Mg compounds existed in the PMB. The XRD analysis showed a strong diffraction peak at 25.91° associated with the typical graphite-like structure of PMB [1]. Moreover, the characteristic peaks at 13.59°, 31.27°, 26.66°, 35.42°, and 43.93° corresponded well with magnesium phosphate, calcium carbonate, calcium phosphate, and calcium carbide (Figure S1b). The SEM image of PMB showed many pore structures on the surface (Figure S1c), which was responsible for the high specific surface area. The EDS analysis confirmed that the elemental composition of PMB was dominated by C, P, O, and Mg (Figure S1d).

3.2. Effect of PMB on the Growth, Contents, and Subcellular Distribution of Cd and Pb of Pakchoi

Compared to CK, the Pb + Cd stress inhibited plant growth, leading to a significant reduction in plant height, root length, leaf dry weight, and root dry weight by 26.31%, 30.61%, 61.57%, and 49.71%, respectively (Table 2). These decreases indicated the toxicity of Pb + Cd to pakchoi growth. However, the addition of PMB reduced the inhibition in pakchoi growth caused by the Pb + Cd stress, leading to a significant increase in those parameters by 28.92%, 33.58%, 81.93%, and 71.59%, respectively, as compared to the Pb + Cd treatment. Furthermore, those growth parameters increased by 8.11%, 11.56%, 13.43%, and 17.71% under the PMB treatment alone, respectively. This indicated that the application of PMB could promote pakchoi plant growth.
As shown in Figure 1, under the Cd + Pb treatment, Cd and Pb contents in leaves and roots were significantly increased (p = 0.037). After the application of PMB, Cd and Pb contents markedly decreased (p = 0.025), which were decreased by 83.84% and 74.29% in leaves and by 51.91% and 47.93% in roots, respectively, compared with the Pb + Cd stress alone. Furthermore, the TFs of Cd and Pb were less than one, indicating a higher accumulation of Cd and Pb in the roots compared to the shoots. The application of PMB reduced the TFs of Cd and Pb by 67.91% and 50.62%, respectively (Figure S2).
In Figure 2, it is evident that Cd and Pb in both the leaves and roots of pakchoi mainly accumulated in the cell wall, followed by the cytoplasm and organelles. Upon the application of PMB, the levels of Cd and Pb in each subcellular component significantly decreased (p = 0.034). For leaves, the use of PMB reduced the contents of Cd and Pb in cell wall, cytoplasm, and organelles by 85.34% and 77.77%, 90.64% and 85.56%, and 84.45% and 92.32%, respectively. For roots, the levels of Cd and Pb were reduced by 55.48% and 44.16%, 72.82% and 63.32%, and 79.69% and 76.43%, respectively.

3.3. Effects of PMB on Soil Properties, Enzyme Activities, Cd and Pb Contents, and Their Chemical Speciation

As depicted in Figure S3, compared with CK, the soil pH, SOM, AP, AK, and AHN value of (Pb + Cd)-polluted soil decreased by 18.65%, 32.17%, 47.65%, 28.57%, and 30.96%, respectively. These values in PMB-added soil significantly increased by 17.92%, 33.14%, 47.82%, 30.33% and 34.36% under Pb + Cd stress, respectively, and they increased by 23.35%, 34.72%, 34.42%, 34.83% and 39.28%, respectively, when PMB was applied alone.
As shown in Figure S4, compared with CK, the urease, sucrase, and catalase activities under Pb + Cd stress decreased by 35.16%, 44.94%, and 48.64%, respectively. When PMB was applied to (Pb + Cd)-contaminated soil, the urease, invertase, and catalase activities increased by 66.30%, 51.71%, and 52.27%, respectively. Meanwhile, the urease, invertase, and catalase in soil treated by PMB alone increased by 35.16%, 25.35%, and 34.83%, respectively.
As depicted in Figure 3, the addition of PMB significantly decreased the contents of available Cd and Pb by 65.05% and 52.25%, respectively. Further, the addition of PMB significantly reduced the acid extractable state of Cd and Pb by 40.62% and 34.01%, respectively (Figure 3). However, the addition of PMB increased the reducible state, oxidizable state, and residual state of Cd and Pb by 5.59% and 0.36%, 10.19% and 5.73%, and 919.47% and 672.2%, respectively.

3.4. Effects of PMB on Soil Bacterial Communities

3.4.1. Soil Bacterial Community Composition

As shown in Figure 4a,c, there were notable differences in the relative abundance of bacteria at the phylum level. The ten dominant soil bacterial communities in all samples were Proteobacteria, Bacteroidota, Acidobacteriota, Actinobacteriota, Gemmatimonadota, Patescibacteria, Verrucomicrobiota, Chloroflexi, Myxococcota, and Planctomycetota, which accounted for 96.59%, 97.23%, 97.33%, and 96.20% of the total number of bacteria in CK, PMB, Pb + Cd, and Pb + Cd + PMB, respectively. Among them, Proteobacteria was the largest bacterial community. Compared with CK, the number of Proteobacteria under Pb + Cd stress increased by 6.01%. However, it decreased in the Pb + Cd + PMB treatment by 22.72% compared with the Pb + Cd treatment. When PMB was applied alone, the number of Proteobacteria decreased slightly by 5.32%.
As depicted in Figure 4b, at the genus level, norank_Caulobacteraceae, Sphingomonas, Gemmatimonas, and norank_Pedosphaeraceae showed a marked decrease in the Pb + Cd treatment compared with CK. The addition of PMB increased these bacteria significantly under the Pb + Cd treatment and CK. Moreover, compared to CK, RB41, norank_Gemmatimonadaceae, and norank_Vicinamibacteraceae showed a significant increase in the Pb + Cd treatment. However, the addition of PMB decreased these bacteria significantly under the Pb + Cd treatment and CK.
In Figure 4d, the Venn diagram illustrated the differences in OTUs between different treatments. There were 2228 core OTUs present in all samples. Additionally, the PMB treatment had the highest number of unique OTUs (859), while the Pb + Cd group had the lowest (516), and the CK and Pb + Cd + PMB groups had 704 and 657 unique OTUs, respectively. Compared to CK, the number of unique OTUs decreased by 26.70% under Pb + Cd stress. Under Pb + Cd stress, the addition of PMB increased the number of unique OTUs by 27.32%. PMB alone also increased it by 22.01% compared to CK.

3.4.2. Alpha and Beta Analyses of Soil Bacterial Community

Table 3 showed the alpha diversity index of soil bacteria based on the 16S rRNA gene. The Chao1 index and Shannon index were used to assess the richness and diversity of soil microbial communities. Compared to CK, the Chao1 and Shannon indices of bacteria in the Pb + Cd treatment group decreased by 14.51% and 7.75%, respectively. Under Pb + Cd stress, the addition of PMB increased the Chao1 and Shannon indices by 15.39% and 7.09%, respectively. The Chao1 and Shannon indices of bacteria treated with PMB alone increased by 10.93% and 11.14% compared to CK, respectively. The findings suggested that the addition of PMB markedly increased the diversity and abundance of soil bacteria.
The principal coordinate analysis illustrated the soil bacterial communities in the four treatments (Figure 5). PC1 and PC2 axes explained 31.7% and 19.4% of the sample composition differentiation, respectively. There were noticeable differences among the four treatments according to their location in Figure 5, indicating that PMB had a significant separation effect on soil microorganisms.

3.4.3. Correlation Analysis and LEFse Analysis

The correlation analysis (Figure 6) demonstrated that soil pH, SOM, AP, AK, AHN, CAT, URA, INV, plant height, root length, leaf dry weight, and root dry weight were significant and positive correlated with the phyla Planctomycetota, Myxococcota, and Verrucomicrobiota but negative correlated with Proteobacteria and Patescibacteria. Soil available Cd, available Pb, leaf Cd, leaf Pb, root Cd, and root Pb were positively correlated with Proteobacteria and Acidobacteriota but significantly and negatively correlated with Bacteroidota.
The taxonomic cladogram line discriminant analysis effect size (LEfSe) indicated the predominant discriminant taxa of the bacterial communities in the different treatments (Figure 7). In CK, Bacteroidota and Chloroflexi (phylum), Chitinophagales and Cytophagales (order), and Chitinophaga and Gemmatimonas (genus) were significantly enriched. For the Pb + Cd treatment, it merely enriched Verrucomicrobiota (Phylum), Pedosphaerales (order), and norank_Pedosphaeraceae c6 (genus). After applying PMB under Pb + Cd, RB41, norank_Subgroup_7, norank_Vicinamibacteraceae, and Cellulomonas at the genus level were significantly enriched. After application of PMB alone, Proteobacteria (phylum), Rickettsiales (order), norank_Bacteroidetes_VC2_1_Bac22 (family), and Ellin6067 (genus) were significantly enriched.

4. Discussion

Many studies have found that biochar has a large specific surface area, rich functional groups, and a stable structure, which can adsorb and fix heavy metals in soil, improve soil physical and chemical properties, and promote plant growth [41]. In this study, the Pb + Cd treatment significantly decreased the plant height, root length, and the leaf and root weight of pakchoi; however, the application of PMB significantly increased these growth parameters of pakchoi under the Pb + Cd treatment. This suggested that PMB could promote pakchoi growth. This was consistent with the findings of Chen et al. [42]. This was because the addition of PMB to the soil reduced the Pb and Cd accumulation in roots and shoots and their translocation from the roots to the leaves. The transport of heavy metals from soil to plants was mainly influenced by the quantity of available metals in the soil [43]. Our findings indicated that the application of PMB in (Pb + Cd)-contaminated soil markedly reduced the content of bioavailable Cd and Pb in the soil. This was consistent with previous research [44], indicating that applying biochar could reduce the contents of Cd and Pb in bioavailable forms. This was because biochar exhibited a wide range of adsorption capabilities for heavy metals [41]. Generally, biochar rich in oxygen-containing functional groups, carbonate, or phosphate is conducive to the passivation of cationic heavy metals. Reduced bioavailability of heavy metals in soil inhibits plant absorption, thereby decreasing heavy metal bioaccumulation in plants. Our results also confirmed that PMB effectively reduced the Cd and Pb contents of pakchoi roots and leaves in (Pb + Cd)-polluted soil.
Compartmentalization is a key approach for detoxifying heavy metals in plant cells. The cell wall, as the primary physical barrier against heavy metals, can provide various negative functional groups, such as carboxyl and hydroxyl, to bind heavy metal ions and inhibit their translocation across the cytoplasmic membrane [3]. Simultaneously, heavy metal ions enter in the vacuole and are sequestered by organic ligands, therefore reducing the interference with cell organelles. In a previous study, Sun et al. [45] showed that most of the Cd2+ ions were immobilized by the hemicellulose 1 of the cell wall in pakchoi roots, and the content of hemicellulose was positively correlated with the Cd immobilization capacity. Pectin was identified as the main polysaccharide in the root cell wall of rice exhibiting significant Cd buildup. In this study, Cd and Pb in leaves and roots of pakchoi mainly accumulated in the cell wall, followed by the cytoplasm and organelles. After PMB was applied, the contents of Cd and Pb in each cell component were significantly decreased. However, the proportion of Cd and Pb content in the cell wall components increased significantly, indicating that more Cd and Pb were bound by these components in the cell wall, thus reducing the toxic effect of heavy metals on pakchoi.
The bioavailability of heavy metals in soils was related to the physical and chemical properties of the soil [28]. Biochar can significantly regulate the physical and chemical properties of the soil. In this study, the addition of PMB in both control and Pb + Cd composite-polluted soils increased the soil pH and SOM value. The increase was attributed to the presence of alkaline substances in biochar, including carbonates, phosphates, and silicates, which may consume protons in the soil solution, leading to an elevation in soil pH [46]. The increased pH value could deprotonate soil colloids, facilitating the electrostatic adsorption of Pb2+ and Cd2+ [9]. Hence, the addition of PMB was favorable for the stabilization of heavy metals in the soil. The increase in soil SOM caused by the addition of PMB was attributed to the carbon enrichment of biochar, as higher organic matter in the soil can facilitate the complexation of Cd and Pb in the soil [47]. Previous research also indicated that the improved capacity of biochar to mitigate heavy metal damage was primarily attributable to the alterations in soil physicochemical properties, which were beneficial for immobilizing heavy metals and reduced their mobility and effectiveness in the soil [10,48]. In addition, applying PMB greatly reduced the acid extractable state of Cd and Pb, transforming them into a residual state with low bioavailability in this study. The residual state has the weakest migration ability in soil and is the form with the lowest bioavailability [18]. Both reduced bioavailability and reduced migration ability can be used as indicators to evaluate the effectiveness of the original passivation. Gondek et al. [49] applied biochar and zeolite to Cd, Pb, and Zn-contaminated soils to reduce the exchangeable states of Cd, Pb, and Zn and increase the parts with low bioavailability. The content of the residual state increased, and heavy metal stabilization was realized. The observed changes in soil properties in this investigation indicated that the application of PMB was a viable approach to decrease the bioavailability of Cd and Pb.
Biochar is primarily composed of carbon, nitrogen, phosphorus, and potassium, which can enhance the growth of microorganisms [41]. Soil enzymes are essential for soil physical and chemical reactions and highly sensitive to heavy metal contamination. Numerous research studies have indicated that the moderate amount of biochar could boost the activities of urease, sucrase, and catalase in the soil [10,49]. A similar role was confirmed in this study. This could be attributed to the layered porous structure of PMB, which created a better environment for microbial growth and enhanced enzyme activity in the soil. It may be attributed to heightened substrate utilization, which in turn stimulated the reaction of metals with organic substances, leading to the formation of metal–organic complexes and a reduction in the toxicity of Cd and Pb [50].
Soil microorganisms are highly sensitive indicators of changes in soil properties and play a crucial role in maintaining ecosystem health and nutrient cycling [51]. The use of PMB had a significant impact on the composition of the bacterial community at the phylum level and increased the number of operational taxonomic units (OTUs), bacterial diversity, and bacterial abundance under Pb + Cd stress, possibly due to the introduction of substantial quantities of phosphorus, oxygen-containing functional groups, and carbonate. Alpha diversity is an important indicator of bacterial richness and diversity. It was observed that the addition of PMB enhanced the abundance and diversity of bacteria and increased the Shannon and Chao1 indices. This could be attributed to the appropriate increase in soil pH, SOM, AP, AK, AHN, INV, URA, and CAT, due to the application of biochar, promoting the growth of certain bacteria [47], as evidenced by the increase in Acidobacteriota in the soil.
Previous research has demonstrated exposure to abiotic stresses such as heavy metals, wastewater, and pesticides can impact the beneficial microorganisms present in the soil, including Proteobacteria, Acidobacteriota, Gemmatimonadota, Bacteroidota, Chloroflexi, Actinobacteriota, and Myxococcota [52,53]. This research found that Proteobacteria, Bacteroidota, Acidobacteriota, and Actinobacteriota were the most predominant bacterial phyla in soil, similar to previous studies [54,55]. Proteobacteria is known for its high resistance to heavy metals due to its ability to adapt to extreme environments such as soil polluted with heavy metals [13]. Actinobacteria is predominant in carbon-rich environments and plays a crucial role in the carbon cycle. They are usually associated with the decomposition of organic materials such as chitin and cellulose and are thought to play a major role in converting soil organic matter in the ecosystem. In this study, the number of Actinobacteriota in PMB treatment increased significantly when compared with CK and the Cd + Pb treatment. This suggested that the PMB enhanced microbial activity involved in the carbon cycle. In addition, our study found that adding PMB in contaminated soil increased the abundance of beneficial microorganisms, including Bacteroidota, Acidobacteriota, Gemmatimonadota, and Patescibacteria. Previous research has showed that these beneficial microorganisms played a crucial role in immobilizing Cd and Pb in the soil [56], leading to the passivation of these metals and an increase in the proportion of fixed Cd and Pb in the soil. These bacteria could reduce the available Cd and Pb in soil via their complexation and absorption abilities [57]. In the present study, the correlation heatmap demonstrated that available Cd and Pb were positively correlated with Proteobacteria and Acidobacteriota but significantly and negatively correlation with Bacteroidota. Bacteroidota affects the cycling and persistence of soil carbon. This may relate to an alteration in soil pH, SOM, AP, AK, AHN, INV, URA, and CAT. Our research findings confirmed that PMB could potentially regulate the structure of soil microorganisms, altering the forms and availability of Cd and Pb, reduce absorption of these metals by plants, and enhance plant growth. Future research should focus on identifying its heavy metal passivation role in the different environmental factors, such as temperature and moisture. Additionally, field trials are essential to validate the broad applicability of our findings across environmental conditions and for long-term stability.

5. Conclusions

The present study demonstrated that PMB significantly promoted pakchoi growth in soil contaminated with Cd and Pb by reducing Cd and Pb accumulation in the plant and their migration from the roots to the shoots of pakchoi. PMB significantly reduced the available contents of Cd and Zn and converted bioavailable Cd and Pb into immobilized forms, while increasing soil pH, organic matter, and available nutrient content. PMB application also improved the activities of urease, invertase, and catalase. Additionally, PMB increased the relative abundance of beneficial bacteria in the soil, including Bacteroidota, Acidobacteriota, Actinobacteriota, and Gemmatimonadota. In summary, this study provided a new strategy for reducing heavy metal pollution in vegetables and fostering sustainable agro-ecosystem development. However, further research is essential to ascertain the effective application of PMB on a large scale under real-field conditions.

Supplementary Materials

The following supporting information can be downloaded at: https://www.mdpi.com/article/10.3390/horticulturae11060632/s1, Table S1: Physicochemical properties of PMB; Figure S1: FTIR (a), XRD (b), SEM (c) and EDS (d) of PMB; Figure S2: The translocation factor of Cd and Pb; Figure S3: Effects of PMB on soil pH (a), soil organic matter (b), available phosphorus (c), available potassium (d) and alkali hydrolyzed nitrogen (e); Figure S4: Effects of PMB on soil enzyme activity. (a) urease, (b) invertase, (c) catalase.

Author Contributions

X.D.: writing—original draft, visualization, methodology, investigation, formal analysis, data curation. H.X.: experimental operation, methodology, formal analysis, data curation. Y.R.: writing—review and editing, supervision. D.L.: formal analysis, data curation. K.L.: methodology, formal analysis. J.H.: writing—review and editing, supervision. All authors have read and agreed to the published version of the manuscript.

Funding

This work was funded by the National Natural Science Foundation of China [31460100].

Data Availability Statement

The original contributions presented in this study are included in the article. Further inquiries can be directed to the corresponding author.

Conflicts of Interest

The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

References

  1. Zhang, L.; Ren, Y.; Xue, Y.; Cui, Z.; Wei, Q.; Han, C.; He, J. Preparation of biochar by mango peel and its adsorption characteristics of Cd(II) in solution. RSC Adv. 2020, 10, 35878–35888. [Google Scholar] [CrossRef] [PubMed]
  2. Gu, J.; Hu, C.; Jia, X.; Ren, Y.; Su, D.; He, J. Physiological and biochemical bases of spermidine-induced alleviation of cadmium and lead combined stress in rice. Plant Physiol. Biochem. 2022, 189, 104–114. [Google Scholar] [CrossRef] [PubMed]
  3. Ji, Y.; Ren, Y.F.; Han, C.; Zhu, W.J.; Gu, J.Y.; He, J.Y. Application of exogenous glycinebetaine alleviates lead toxicity in pakchoi (Brassica chinensis L.) by promoting antioxidant enzymes and suppressing Pb accumulation. Environ. Sci. Pollut. Res. 2022, 29, 25568–25580. [Google Scholar] [CrossRef] [PubMed]
  4. Li, K.; Ren, Y.; Lin, D.; Tong, M.; Yang, B.; Xiao, F.; Hou, Y.; Lu, Y.; He, J. Green preparation of bimetallic CuO/ZnO nanoparticles by using Cinnamomum camphora leaf and its potential for antifungal and cadmium removal applications. Inorg. Chem. Commun. 2025, 178, 114577. [Google Scholar] [CrossRef]
  5. Lan, J.; Zhang, S.S.; Dong, Y.Q.; Li, J.H.; Li, S.Y.; Feng, L.; Hou, H.B. Stabilization and passivation of multiple heavy metals in soil facilitating by pinecone-based biochar: Mechanisms and microbial community evolution. J. Hazard. Mater. 2021, 420, 126588. [Google Scholar] [CrossRef]
  6. Algethami, J.S.; Irshad, M.K.; Javed, W.; Alhamami, M.A.M.; Ibrahim, M. Iron-modified biochar improves plant physiology, soil nutritional status and mitigates Pb and Cd-hazard in wheat (Triticum aestivum L.). Front. Plant Sci. 2023, 14, 1221434. [Google Scholar] [CrossRef]
  7. Bandara, T.; Franks, A.; Xu, J.; Bolan, N.; Wang, H.; Tang, C. Chemical and biological immobilization mechanisms of potentially toxic elements in biochar-amended soils. Crit. Rev. Environ. Sci. Technol. 2020, 50, 903–978. [Google Scholar] [CrossRef]
  8. Haider, F.U.; Ain, N.; Khan, I.; Farooq, M.; Habiba Cai, L.; Li, Y. Co-application of biochar and plant growth regulators improves maize growth and decreases Cd accumulation in cadmium-contaminated soil. J. Clean. Prod. 2024, 440, 140515. [Google Scholar] [CrossRef]
  9. Xu, M.L.; Dai, W.J.; Zhao, Z.L.; Zheng, J.T.; Huang, F.; Mei, C.; Huang, S.T.; Liu, C.F.; Wang, P.; Xiao, R.B. Effect of rice straw biochar on three different levels of Cd-contaminated soils: Cd availability, soil properties, and microbial communities. Chemosphere 2022, 301, 134551. [Google Scholar] [CrossRef]
  10. Noronha, F.R.; Manikandan, S.K.; Nair, V. Role of coconut shell biochar and earthworm (Eudrilus euginea) in bioremediation and palak spinach (Spinacia oleracea L.) growth in cadmium-contaminated soil. J. Environ. Manag. 2022, 302, 114057. [Google Scholar] [CrossRef]
  11. Dewi, R.K.; Huang, Q.; Hashimi, R.; Komatsuzaki, M. Enhancing agroecosystem sustainability: Integrative soil health strategies in regenerative organic soybean production on Andosol in Japan. Geoderma Reg. 2025, 40, e00910. [Google Scholar] [CrossRef]
  12. Chen, X.; Jiang, S.; Wu, J.; Yi, X.; Dai, G.; Shu, Y. Three-year field experiments revealed the immobilization effect of natural aging biochar on typical heavy metals (Pb, Cu, Cd). Sci. Total Environ. 2024, 912, 169384. [Google Scholar] [CrossRef] [PubMed]
  13. Mandal, S.; Pu, S.; He, L.; Ma, H.; Hou, D. Biochar induced modification of graphene oxide & nZVI and its impact on immobilization of toxic copper in soil. Environ. Pollut. 2020, 259, 113851. [Google Scholar] [CrossRef] [PubMed]
  14. Huang, K.; Sun, X.; Sun, J.; Guo, Y.; Hu, X.; Hu, C.; Tan, Q. The role of phosphorus speciation of biochar in reducing available Cd and phytoavailability in mining area soil: Effect and mechanism. Sci. Total Environ. 2023, 894, 164868. [Google Scholar] [CrossRef]
  15. Zhang, H.; Shao, J.; Zhang, S.; Zhang, X.; Chen, H. Effect of phosphorus-modified biochars on immobilization of Cu(II), Cd(II), and As(V) in paddy soil. J. Hazard. Mater. 2020, 390, 121349. [Google Scholar] [CrossRef]
  16. Chu, G.; Zhao, J.; Huang, Y.; Zhou, D.D.; Liu, Y.; Wu, M.; Peng, H.B.; Zhao, Q.; Pan, B.; Steinberg, C.E.W. Phosphoric acid pretreatment enhances the specific surface areas of biochars by generation of micropores. Environ. Pollut. 2018, 240, 1–9. [Google Scholar] [CrossRef]
  17. Dechapanya, W.; Khamwichit, A. Biosorption of aqueous Pb(II) by H3PO4-activated biochar prepared from palm kernel shells (PKS). Heliyon 2023, 9, e17250. [Google Scholar] [CrossRef]
  18. Netherway, P.; Reichman, S.M.; Laidlaw, M.; Scheckel, K.; Pingitore, N.; Gasćo, G.; Méndez, A.; Surapaneni, A.; Paz-Ferreiro, J. Phosphorus-rich biochars can transform lead in an urban contaminated soil. J. Environ. Qual. 2019, 48, 1091–1099. [Google Scholar] [CrossRef]
  19. Zhang, H.; Ke, S.; Xia, M.; Bi, X.; Shao, J.; Zhang, S.; Chen, H. Effects of phosphorous precursors and speciation on reducing bioavailability of heavy metal in paddy soil by engineered biochars. Environ. Pollut. 2021, 285, 117459. [Google Scholar] [CrossRef]
  20. Ge, Q.; Tian, Q.; Wang, S.; Zhu, F. Synergistic effects of phosphoric acid modified hydrochar and coal gangue-based zeolite on bioavailability and accumulation of cadmium and lead in contaminated soil. Chinese J. Chem. Eng. 2022, 46, 150–160. [Google Scholar] [CrossRef]
  21. Sahin, O.; Yagcioglu, K.D.; Kadioglu, Y.K.; Gunes, A. The importance of acidified biochar as a sustainable phosphorus source and its role in balanced nutrition for spinach (Spinacia oleracea L.). J. Soil Sci. Plant Nutri. 2024, 24, 8035–8045. [Google Scholar] [CrossRef]
  22. Ahmad Munir Usman, A.R.A.; Al-Faraj, A.S.; Ahmad Mahtab Sallam, A.; Al-Wabel, M.I. Phosphorus-loaded biochar changes soil heavy metals availability and uptake potential of maize (Zea mays L.) plants. Chemosphere 2018, 194, 327–339. [Google Scholar] [CrossRef] [PubMed]
  23. Behrooz, B.A.; Oustan, S.; Hosseini, H.M.; Etesami, H.; Padoan, E.; Magnacca, G.; Marsan, F.A. The importance of presoaking to improve the efficiency of MgCl2-modified and non-modified biochar in the adsorption of cadmium. Ecotoxicol. Environ. Saf. 2023, 257, 114932. [Google Scholar] [CrossRef] [PubMed]
  24. Deng, Y.; Li, X.; Ni, F.; Liu, Q.; Yang, Y.; Wang, M.; Ao, T.; Chen, W. Synthesis of magnesium modified biochar for removing copper, lead and cadmium in single and binary systems from aqueous solutions: Adsorption mechanism. Water 2021, 13, 599. [Google Scholar] [CrossRef]
  25. Shan, R.; Li, W.; Chen, Y.; Sun, X. Effects of Mg-modified biochar on the bioavailability of cadmium in soil. BioResources 2020, 15, 8008–8025. [Google Scholar] [CrossRef]
  26. Wang, Y.; Li, J.; Xu, L.; Wu, D.; Li, Q.; Ai, Y.; Liu, W.; Li, D.; Zhou, Y.; Zhang, B.; et al. EDTA functionalized Mg/Al hydroxides modified biochar for Pb(II) and Cd(II) removal: Adsorption performance and mechanism. Sep. Purif. Technol. 2024, 335, 126199. [Google Scholar] [CrossRef]
  27. Azeem, M.; Ali, A.; Jeyasundar, P.G.S.A.; Li, Y.M.; Abdelrahman, H.; Latif, A.; Li, R.H.; Basta, N.; Li, G.; Shaheen, S.M.; et al. Bone-derived biochar improved soil quality and reduced Cd and Zn phytoavailability in a multi-metal contaminated mining soil. Environ. Pollut. 2021, 277, 116800. [Google Scholar] [CrossRef]
  28. Qu, J.H.; Yuan, Y.H.; Zhang, X.M.; Wang, L.; Tao, Y.; Jiang, Z.; Yu, H.; Dong, M.; Zhang, Y. Stabilization of lead and cadmium in soil by sulfur-iron functionalized biochar: Performance, mechanisms and microbial community evolution. J. Hazard. Mater. 2022, 425, 127876. [Google Scholar] [CrossRef]
  29. Wang, Y.; Ren, Q.; Li, T.; Zhan, W.; Zheng, K.; Li, Y.; Chen, R. Influences of modified biochar on metal bioavailability, metal uptake by wheat seedlings (Triticum aestivum L.) and the soil bacterial community. Ecotox. Environ. Saf. 2021, 220, 112370. [Google Scholar] [CrossRef]
  30. Zhang, J.; Jiang, Y.; Ding, C.; Wnag, S.; Zhao, C.; Yin, W.; Wang, B.; Yang, R.; Wang, X. Remediation of lead and cadmium co-contaminated mining soil by phosphate-functionalized biochar: Performance, mechanism, and microbial response. Chemosphere 2023, 334, 138938. [Google Scholar] [CrossRef]
  31. Han, C.; Wang, M.; Ren, Y.F.; Zhang, L.; Ji, Y.; Zhu, W.; Song, Y.; He, J.Y. Characterization of pruned tea branch biochar and the mechanisms underlying its adsorption for cadmium in aqueous solution. RSC Adv. 2021, 11, 26832–26843. [Google Scholar] [CrossRef] [PubMed]
  32. Xu, H.J.; Zhou, Q.; Yan, T.Y.; Jia, X.W.; Lu, D.D.; Ren, Y.F.; He, J.Y. Enhanced removal efficiency of Cd2+ and Pb2+ from aqueous solution by H3PO4-modified tea branch biochar: Characterization, adsorption performance and mechanism. J. Environ. Chem. Eng. 2024, 12, 112183. [Google Scholar] [CrossRef]
  33. Ren, Y.F.; Wang, W.; He, J.Y.; Zhang, L.; Wei, Y.; Yang, M. Nitric oxide alleviates salt stress in seed germination and early seedling growth of pakchoi (Brassica chinensis L.) by enhancing physiological and biochemical parameters. Ecotox. Environ. Saf. 2020, 187, 109785. [Google Scholar] [CrossRef] [PubMed]
  34. GB 15618-2018; China: National Standard Soil Environmental Quality Risk Control Standard for Soil Contamination of Agricultural Land. USDA: Beijing, China, 2018.
  35. Yang, Q.; Li, Z.; Lu, X.; Duan, Q.; Huang, L.; Bi, J. A review of soil heavy metal pollution from industrial and agricultural regions in China: Pollution and risk assessment. Sci. Total Environ. 2018, 642, 690–700. [Google Scholar] [CrossRef]
  36. Bian, R.; Li, L.; Bao, D.; Zheng, J.; Zhang, X.; Zheng, J.; Liu, X.; Cheng, K.; Pan, G. Cd immobilization in a contaminated rice paddy by inorganic stabilizers of calcium hydroxide and silicon slag and by organic stabilizer of biochar. Environ. Sci. Pollut. Res. 2016, 23, 10028–10036. [Google Scholar] [CrossRef]
  37. Su, D.M.; Wang, M.F.; Xu, H.J.; Zhang, Y.Z.; Hu, Q.F.; Ren, Y.F.; He, J.Y. Performance and mechanism of Ficus carica branch waste based biochar in removing Cd2+ from aqueous solution. Biomass Convers. Bior. 2024, 14, 24137–24150. [Google Scholar] [CrossRef]
  38. Weigal, H.J. Jager HJ. Subcellular distribution and chemical form of cadmium in bean. Plant Physiol. 1980, 65, 480–482. [Google Scholar] [CrossRef]
  39. Wang, W.; Ren, Y.F.; He, J.; Zhang, L.; Wang, X.; Cui, Z. Impact of copper oxide nanoparticles on the germination, seedling growth, and physiological responses in Brassica pekinensis L. Environ. Sci. Pollut. Res. 2020, 27, 31505–31515. [Google Scholar] [CrossRef]
  40. Hamidpour, M.; Sadeghi, R.; Abbaszadeh-Dahaji, P.; Alaei, H.; Shafigh, M.; Omidvari, M.; Kariman, K. The effects of EDTA and Trichoderma species on growth and Cu uptake of maize (Zea mays) plants grown in a Cu-contaminated soil. Environ. Geochem. Health 2024, 46, 372. [Google Scholar] [CrossRef]
  41. Lu, X.; Sun, J.; Pan, G.; Qi, W.; Zhang, Z.; Xing, J.; Gao, Y. Ball-milling-modified biochar with additives enhances soil Cd passivation, increases plant growth and restrains Cd uptake by chinese cabbage. Horticulturae 2025, 11, 168. [Google Scholar] [CrossRef]
  42. Chen, X.L.; Ran, Z.F.; Li, R.; Duan, W.Y.; Song, Z.J.; Fang, L.; Guo, L.P.; Zhou, J. Biochar reduces the cadmium content of Panax quinquefolium L. by improving rhizosphere microecology. Sci. Total Environ. 2024, 915, 170005. [Google Scholar] [CrossRef] [PubMed]
  43. Gu, T.Y.; Lu, Y.Q.; Li, F.; Zeng, W.M.; Shen, L.; Yu, R.L.; Li, J.K. Microbial extracellular polymeric substances alleviate cadmium toxicity in rice (Oryza sativa L.) by regulating cadmium uptake, subcellular distribution and triggering the expression of stress-related genes. Ecotox. Environ. Saf. 2023, 257, 114958. [Google Scholar] [CrossRef]
  44. Awad, M.; El-Sayed, M.M.; Li, X.; Liu, Z.; Mustafa, S.K.; Ditta, A.; Hessini, K. Diminishing heavy metal hazards of contaminated soil via biochar supplementation. Sustainability 2021, 13, 12742. [Google Scholar] [CrossRef]
  45. Sun, Q.H.; Zhang, Y.X.; Ming, C.S.; Wang, J.M.; Zhang, Y. Amended compost alleviated the stress of heavy metals to pakchoi plants and affected the distribution of heavy metals in soil-plant system. J. Environ. Manag. 2023, 336, 117674. [Google Scholar] [CrossRef] [PubMed]
  46. Qi, X.; Gou, J.L.; Chen, X.M.; Xiao, S.Q.; Ali, I.; Shang, R.; Wang, D.; Wu, Y.W.; Han, M.W.; Luo, X.G. Application of mixed bacteria-loaded biochar to enhance uranium and cadmium immobilization in a co-contaminated soil. J. Hazard. Mater. 2021, 401, 123823. [Google Scholar] [CrossRef]
  47. Liu, Q.Q.; Sheng, Y.Q.; Wang, W.J.; Liu, X.Z. Efficacy and microbial responses of biochar- nanoscale zero-valent during in-situ remediation of Cd-contaminated sediment. J. Clean. Prod. 2021, 287, 125076. [Google Scholar] [CrossRef]
  48. Bashir, S.; Hussain, Q.; Zhu, J.; Fu, Q.L.; Houben, D.; Hu, H.Q. Efficiency of KOH- modified rice straw-derived biochar for reducing cadmium mobility, bioaccessibility and bioavailability risk index in red soil. Pedosphere 2020, 30, 874–882. [Google Scholar] [CrossRef]
  49. Gondek, K.; Mierzwa-Hersztek, M.; Jarose, R. Effect of willow biochar and fly ash-derived zeolite in immobilizing heavy metals and promoting enzymatic activity in a contaminated sandy soil. Catena 2023, 232, 107429. [Google Scholar] [CrossRef]
  50. Yang, X.; Wen, E.; Ge, C.; El-Naggar, A.; Yu, H.; Wang, S.S.; Kwon, E.E.; Song, H.; Shaheen, S.M.; Wang, H.L.; et al. Iron-modified phosphorus- and silicon-based biochars exhibited various influences on arsenic, cadmium, and lead accumulation in rice and enzyme activities in a paddy soil. J. Hazard. Mater. 2023, 443, 130203. [Google Scholar] [CrossRef]
  51. Lentini, M.; Ciriello, M.; Rouphael, Y.; Campana, E.; Vaccari, F.P.; De Pascale, S. Harnessing biochar for sustainable horticulture: Strategies to cope with abiotic stress. Horticulturae 2025, 11, 73. [Google Scholar] [CrossRef]
  52. Pradhan, S.K.; Singh, N.S.; Kumar, U.; Mishra, S.; Perumal, R.C.; Benny, J.; Thatoi, H. Illumina MiSeq based assessment of bacterial community structure and diversity along the heavy metal concentration gradient in Sukinda chromite mine area soils. India Ecol. Genet. Genom. 2020, 15, 100054. [Google Scholar] [CrossRef]
  53. Narendrula-Kotha, R.; Nkongolo, K. Bacterial and fungal community structure and diversity in a mining region under long-term metal exposure revealed by metagenomics sequencing. Ecol. Genet. Genom. 2017, 2, 13–24. [Google Scholar] [CrossRef]
  54. Ibrahim, M.M.; Tong, C.X.; Hu, K.; Zhou, B.Q.; Xing, S.H.; Mao, Y.L. Biochar-fertilizer interaction modifies N-sorption, enzyme activities and microbial functional abundance regulating nitrogen retention in rhizosphere soil. Sci. Total Environ. 2020, 739, 140065. [Google Scholar] [CrossRef] [PubMed]
  55. Yang, W.H.; Li, C.J.; Wang, S.S.; Zhou, B.Q.; Mao, Y.L.; Rensing, C.; Xing, S.H. Influence of biochar and biochar-based fertilizer on yield, quality of tea and microbial community in an acid tea orchard soil. Appl. Soil Ecol. 2021, 166, 104005. [Google Scholar] [CrossRef]
  56. Wu, S.J.; Zhou, Z.J.; Zhu, L.; Zhong, L.M.; Dong, Y.X.; Wang, G.J.; Shi, K.X. Cd immobilization mechanisms in a Pseudomonas strain and its application in soil cd remediation. J. Hazard. Mater. 2022, 425, 127919. [Google Scholar] [CrossRef]
  57. Wang, P.; Cao, J.; Mao, L.; Zhu, L.; Zhang, Y.; Zhang, L.; Jiang, H.; Zheng, Y.; Liu, X. Effect of H3PO4-modified biochar on the fate of atrazine and remediation of bacterial community in atrazine-contaminated soil. Sci. Total Environ. 2022, 851, 158278. [Google Scholar] [CrossRef]
Horticulturae 11 00632 g001
Figure 1. Effects of PMB on Cd (a) and Pb (b) contents in pakchoi. Different lowercase letters on the top of column indicate a significant difference at the 0.05 level.
Figure 1. Effects of PMB on Cd (a) and Pb (b) contents in pakchoi. Different lowercase letters on the top of column indicate a significant difference at the 0.05 level.
Horticulturae 11 00632 g001
Horticulturae 11 00632 g002
Figure 2. Effects of PMB on subcellular distribution of Cd (a,c) and Pb (b,d) in leaves and roots parts of pakchoi under Pb and Cd stress. Different lowercase letters on the top of column indicate a significant difference at the 0.05 level.
Figure 2. Effects of PMB on subcellular distribution of Cd (a,c) and Pb (b,d) in leaves and roots parts of pakchoi under Pb and Cd stress. Different lowercase letters on the top of column indicate a significant difference at the 0.05 level.
Horticulturae 11 00632 g002
Horticulturae 11 00632 g003
Figure 3. The available content and proportion of different forms of Cd (a,c) and Pb (b,d). Different lowercase letters on the top of column indicate a significant difference at the 0.05 level.
Figure 3. The available content and proportion of different forms of Cd (a,c) and Pb (b,d). Different lowercase letters on the top of column indicate a significant difference at the 0.05 level.
Horticulturae 11 00632 g003
Horticulturae 11 00632 g004
Figure 4. The changes in the rhizosphere soil microbial community under different treatments. (a) Relative abundance of bacteria, (b) relative abundance heat map, (c) collinearity diagram, (d) Venn diagram of OTU distribution of soil bacteria. D1 refers to CK, D2 refers to PMB, D3 refers to Pb + Cd, and D4 refers to Pb + Cd + PMB.
Figure 4. The changes in the rhizosphere soil microbial community under different treatments. (a) Relative abundance of bacteria, (b) relative abundance heat map, (c) collinearity diagram, (d) Venn diagram of OTU distribution of soil bacteria. D1 refers to CK, D2 refers to PMB, D3 refers to Pb + Cd, and D4 refers to Pb + Cd + PMB.
Horticulturae 11 00632 g004
Horticulturae 11 00632 g005
Figure 5. PCA principal coordinate analysis. D1 refers to CK, D2 refers to PMB, D3 refers to Pb + Cd, and D4 refers to Pb + Cd + PMB.
Figure 5. PCA principal coordinate analysis. D1 refers to CK, D2 refers to PMB, D3 refers to Pb + Cd, and D4 refers to Pb + Cd + PMB.
Horticulturae 11 00632 g005
Horticulturae 11 00632 g006
Figure 6. Correlation heatmap of bacterial community at the phylum level with soil and plant variables.
Figure 6. Correlation heatmap of bacterial community at the phylum level with soil and plant variables.
Horticulturae 11 00632 g006
Horticulturae 11 00632 g007
Figure 7. A taxonomic cladogram of line discriminant analysis effect size (LEfSe) based on OTUs. The cladogram shows the biomarker microbes of the microbial lineages from phylum to genus among four different treatments. Yellow dots represent microbes with no statistical differences among the five treatments. D1 refers to CK, D2 refers to PMB, D3 refers to Pb + Cd, and D4 refers to Pb + Cd + PMB.
Figure 7. A taxonomic cladogram of line discriminant analysis effect size (LEfSe) based on OTUs. The cladogram shows the biomarker microbes of the microbial lineages from phylum to genus among four different treatments. Yellow dots represent microbes with no statistical differences among the five treatments. D1 refers to CK, D2 refers to PMB, D3 refers to Pb + Cd, and D4 refers to Pb + Cd + PMB.
Horticulturae 11 00632 g007
Table 1. Basic physicochemical properties of the soil.
Table 1. Basic physicochemical properties of the soil.
Soil TypepHCation Exchange Capacity (cmol/L)Organic Matter (g/kg)Total Nitrogen (g/kg)Available
Phosphorus (mg/kg)		Available
Potassium (mg/kg)
Yellow-brown soil6.21.2520.321.7127.84157.14
Table 2. Plant height, root length, and biomass of pakchoi under Pb and Cd stress.
Table 2. Plant height, root length, and biomass of pakchoi under Pb and Cd stress.
TreatmentsPlant Height
(cm)		Root Length
(cm)		Leaf Dry Weight (g/plant)Root Dry Weight (g/plant)
CK20.60 ± 0.49 c5.88 ± 0.23 ab2.16 ± 0.14 c0.175 ± 0.014 b
PMB22.27 ± 0.76 a6.56 ± 0.51 a2.45 ± 0.16 a0.206 ± 0.021 a
Cd + Pb15.18 ± 0.42 d4.08 ± 0.45 c0.83 ± 0.054 d0.088 ± 0.007 c
Cd + Pb + PMB19.57 ± 0.35 b5.45 ± 0.42 b1.51 ± 0.12 b0.151 ± 0.013 b
Note: Different lowercase letters after data in the same column indicate a significant difference at the 0.05 level.
Table 3. Richness and diversity indices of soil bacterial communities under different treatments.
Table 3. Richness and diversity indices of soil bacterial communities under different treatments.
TreatmentsChao1Shannon
CK2040.32 ± 73.48 b4.13 ± 0.04 b
PMB2263.10 ± 65.31 a4.59 ± 0.04 a
Pb + Cd1744.18 ± 81.64 c3.81 ± 0.08 c
Pb + Cd + PMB2012.62 ± 85.73 b4.08 ± 0.11 b
Note: Different lowercase letters after data in the same column indicate a significant difference at the 0.05 level.
Disclaimer/Publisher’s Note: The statements, opinions and data contained in all publications are solely those of the individual author(s) and contributor(s) and not of MDPI and/or the editor(s). MDPI and/or the editor(s) disclaim responsibility for any injury to people or property resulting from any ideas, methods, instructions or products referred to in the content.

Share and Cite

MDPI and ACS Style

Dong, X.; Xu, H.; Ren, Y.; Lin, D.; Li, K.; He, J. Phosphoric Acid and Magnesium Chloride Composite-Modified Biochar Improved Pakchoi Growth by Reducing Pb and Cd Accumulation and Altering Soil Properties and Microbial Communities. Horticulturae 2025, 11, 632. https://doi.org/10.3390/horticulturae11060632

AMA Style

Dong X, Xu H, Ren Y, Lin D, Li K, He J. Phosphoric Acid and Magnesium Chloride Composite-Modified Biochar Improved Pakchoi Growth by Reducing Pb and Cd Accumulation and Altering Soil Properties and Microbial Communities. Horticulturae. 2025; 11(6):632. https://doi.org/10.3390/horticulturae11060632

Chicago/Turabian Style

Dong, Xuejie, Haojie Xu, Yanfang Ren, Dongming Lin, Ke Li, and Junyu He. 2025. "Phosphoric Acid and Magnesium Chloride Composite-Modified Biochar Improved Pakchoi Growth by Reducing Pb and Cd Accumulation and Altering Soil Properties and Microbial Communities" Horticulturae 11, no. 6: 632. https://doi.org/10.3390/horticulturae11060632

APA Style

Dong, X., Xu, H., Ren, Y., Lin, D., Li, K., & He, J. (2025). Phosphoric Acid and Magnesium Chloride Composite-Modified Biochar Improved Pakchoi Growth by Reducing Pb and Cd Accumulation and Altering Soil Properties and Microbial Communities. Horticulturae, 11(6), 632. https://doi.org/10.3390/horticulturae11060632

Note that from the first issue of 2016, this journal uses article numbers instead of page numbers. See further details here.

Article Metrics

Back to TopTop