Phytoremediation Potential of Melia azedarach and Ailanthus altissima for Pb, Zn, and Cu from Aqueous Solution

Abstract

Growing concerns over metal pollution highlight the need for effective remediation strategies. This study evaluates the accumulation capacity and tolerance of Melia azedarach and Ailanthus altissima for trace elements (Pb, Zn, and Cu), aiming to assess their phytoremediation potential. Three-month-old seedlings of both species, grown from seeds collected at the Touissit mine site, were cultivated in perlite and irrigated with Hoagland nutrient solution. Plants were exposed to various concentrations of metal salts—Pb(NO₃)₂ (8, 40, and 80 mg.L⁻¹), ZnSO₄ (8, 40, and 80 mg.L⁻¹), and CuSO₄ (2, 10, and 20 mg.L⁻¹)—over a 90-day period. Growth, biomass, metal accumulation, chlorophyll, and carotenoid contents were measured. Results indicate that M. azedarach exhibited enhanced biomass under Pb exposure, suggesting notable tolerance and potential for phytoremediation. Conversely, A. altissima showed an initial increase in biomass at low Pb levels, followed by a significant reduction at higher concentrations. Both species demonstrated decreased biomass under Zn and Cu treatments, with varying degrees of sensitivity. Notably, A. altissima accumulated significant levels of Pb, Zn, and Cu, particularly in the roots, indicating high phytoremediation potential. While M. azedarach also accumulated metals, levels were comparatively lower. Both species maintained chlorophyll content under metal stress, indicating resilience. Overall, this hydroponic screening highlights the considerable capacities of M. azedarach and A. altissima for Pb, Zn, and Cu tolerance, with A. altissima showing particularly high potential for Pb phytostabilization.

1. Introduction

High levels of trace elements and metalloid elements in the environment pose a significant threat to the health and survival of humans, animals, and plants, contributing to a global environmental issue due to its accumulation in agricultural products and bioamplification through food chains [1,2]. These metals, derived from various anthropogenic sources—including mining activities, urban runoff, wastewater treatment plants, boating operations, industrial effluents and waste, domestic landfill sites, and agricultural fungicide runoff—have progressively impacted ecosystems and environmental quality [3].

Trace elements are defined as metals and metalloids with an atomic density exceeding 4000 kg/m³ [4]. While plants require certain essential metals for growth, development, and production, excessive concentrations of these elements can lead to phytotoxicity, adversely affecting plant productivity, biomass, crop yields, and overall food security [5]. At elevated levels, both essential and non-essential trace elements can cause toxicity and inhibit plant growth [6]. For instance, copper (Cu) and zinc (Zn) are essential for healthy plant growth, as they are integral to various enzymes and proteins. However, exposure to excessive levels of Cu and Zn can harm plants [7]. High soil Cu levels often stem from copper mining, wastewater discharge, and Cu-based agricultural chemicals [8]. Lead (Pb), comprising approximately 0.002% of the Earth’s crust, is highly toxic and persistent, posing significant risks to both plants and animals. Lead ranks as the second most hazardous metal due to its adverse ecological impacts [9,10]. The critical Pb threshold for plant tissue is around 2 mg kg⁻¹; whereas, agricultural soils have a Pb threshold ranging from 50 to 300 mg kg⁻¹ [11].

Improper handling of mine waste can lead to the dispersion and deposition of trace elements on surrounding soils, contaminating stream sediments and potentially impacting the quality of drinking water sources (both surface and groundwater) and agricultural soils located far from the original pollution sites [12,13]. Major sources of soil contamination include mining operations for copper, zinc, lead, manganese, and iron—metals widely used in industrial applications. Given the high concentrations of these metals frequently detected in soil samples from mining areas, a comprehensive evaluation of their environmental impacts is essential [14,15].

After mineral extraction, it is essential to restore productivity to affected areas and prepare them for revegetation. Due to growing environmental concerns, postmining reclamation has become a critical part of the mining process [16]. Phytoremediation stands out among the physical, chemical, and biological approaches to soil remediation, offering a cost-effective, environmentally sustainable, and economically beneficial option [17,18,19]. This emerging technique is widely used for soil remediation through the removal or stabilization of environmental contaminants [20,21,22] and extends its applications beyond soil stabilization and pollution mitigation to include aesthetic enhancement [23]. Phytoremediation is an eco-friendly bioremediation technique that utilizes metal-tolerant species to extract or stabilize trace elements in contaminated soils. These plants can either accumulate metals in their tissues, such as roots, shoots, and leaves, or immobilize them in the soil, preventing further spread. Despite its environmental benefits, phytoremediation has several limitations. It is limited by slow remediation, potential toxicity to plants, and restricted action to upper soil layers (up to 1 m), leaving deeper contamination untreated [24,25]. A practical and effective strategy for managing metal pollution called phytostabilization involves screening and selecting species or genotypes that exhibit a high tolerance to toxic metals, while minimizing their accumulation in the aerial parts [26]. Meanwhile, effective phytoremediation relies on selecting native plant species that are well-adapted to local soil conditions, highly tolerant to contaminants, and with efficient metal accumulation capacities [27]. Studying the concentration-dependent effects of Pb, Zn, and Cu on plant phytotoxicity is essential in this context. Hydroponic screening is often recommended as a cost-effective and efficient method for identifying species suited for trace element removal, reducing the need for expensive field trials [28].

Ailanthus altissima (Mill.) Swingle (A. altissima) is a fast-growing woody tree species regarded as an effective cover crop to prevent soil erosion and provide pulp and wood. It is notably resistant to environmental stressors, including trace elements, and is often identified as a resilient species [29]. Melia azedarach (M. Aedarach) is another fast-growing, drought-resistant species with the ability to thrive in low-fertility soils and exhibits strong tolerance to salinity [30].

This study aims to investigate (i) the accumulation capacity of Pb, Zn, and Cu in A. altissima and M. azedarach and (ii) their resistance to the toxic effects of these metals, thereby identifying candidates for phytoremediation in contaminated soils.

2. Results

2.1. Effect of Metals (Pb, Zn, and Cu) on Biomass

The analysis of dry biomass (shoot and root) after 90 days (Figure 1) of M. azedarach and A. altissima under varying metal salt concentrations of Pb, Zn, and Cu indicates a consistent pattern of metal impact on both shoots and roots.

Biomass analysis revealed a stimulatory effect of Pb on M. azedarach, with shoot biomass increasing significantly at higher concentrations (0.7295 g at 80 mg.L⁻¹ compared to 0.3709 g in the control). This aligns with studies where Pb exposure enhanced dry weight under certain conditions, potentially due to Pb-induced alterations in nutrient uptake or signaling [10,31]. Conversely, A. altissima showed tolerance only at lower Pb concentrations, with biomass declining significantly at 40 mg.L⁻¹ and 80 mg.L⁻¹, which can be explained by the role of Pb that negatively affects plant shoot growth by disrupting photosynthesis, mineral nutrition, water balance, hormonal regulation, and membrane structure and permeability [32].

Both species exhibited pronounced sensitivity to Zn, with significant biomass reductions across all concentrations (p < 0.05). The effects were particularly severe for M. azedarach at 80 mg.L⁻¹, where shoot biomass dropped to 0.1476 g. Similarly, A. altissima showed no significant recovery across treatments, mirroring reports of Zn-induced disruption in nutrient balance and oxidative stress. While Zn is essential in trace amounts, excessive levels can impair photosynthesis and growth [10,32,33].

Both species showed progressive biomass reductions with increasing Cu concentrations (p < 0.05). M. azedarach shoot biomass decreased to 0.1104 g at 20 mg.L⁻¹. Similar findings have been reported for Paulownia fortunei, where Cu exposure significantly reduced growth and biomass [34]. Root biomass was more affected than shoots, consistent with the role of roots as primary sites of metal uptake and damage [35].

2.2. Effect of Metals (Pb, Zn, and Cu) on Photosynthetic Pigments

Chlorophylls and total carotenoids are considered reliable and straightforward indicators of Pb-induced phytotoxicity in higher plants [36]. The effects of various metal treatments on photosynthetic pigments (chlorophyll a, chlorophyll b, and carotenoids) in M. azedarach and A. altissima are summarized in (

Table 1. Chl a, b, and carotenoid content (mg.g⁻¹) in A. altissima and M. azedarach grown for 90 days under different Pb, Zn, and Cu concentrations in the hydroponic medium. Data are mean values of 3 replicates ± standard deviation. Cases indicated by the same letters are not statistically significant.

	M. azedarach								A. altissima
Treatments	Chl a	±	Chl b	±	Chl a + b	±	Carotenoids	±	Chl a	±	Chl b	±	Chl a + b	±	Carotenoids	±
C	3.441 a	0.25	0.458 a	0.12	3.898 a	0.36	0.995 a	0.06	7.513 a	2.74	1.203 ab	0.66	8.716 a	3.39	2.068 a	0.58
Zn 8	3.002 ab	0.24	0.611 a	0.27	3.613 a	0.47	0.996 a	0.11	5.351 ab	0.52	0.235 ab	0.54	5.586 ab	0.73	1.341 ab	0.23
Zn 40	3.243 a	0.64	0.335 a	0.27	3.578 a	0.91	1.058 a	0.11	4.531 ab	0.65	1.300 ab	0.23	5.831 ab	0.42	1.193 ab	0.33
Zn 80	3.072 ab	0.33	−0.095 a	0.22	2.977 a	0.53	1.067 a	0.08	2.966 b	0.42	0.738 ab	0.68	3.704 b	1.11	0.598 b	0.20
Pb 8	1.069 c	0.78	4.537 a	0.21	5.606 a	0.59	−1.352 b	0.28	5.014 ab	0.72	0.497 ab	0.31	5.511 ab	0.47	1.048 b	0.56
Pb40	2.279 abc	0.14	0.476 a	0.16	2.755 a	0.19	0.582 ab	0.08	4.543 ab	0.34	1.236 ab	0.33	5.779 ab	0.65	1.095 b	0.02
Pb 80	2.387 abc	0.80	0.111 a	0.19	2.498 a	0.97	0.823 ab	0.20	3.630 b	0.63	1.605 a	0.70	5.235 ab	1.21	0.698 b	0.14
Cu 2	1.019 c	0.31	2.528 a	0.22	3.548 a	0.49	−0.665 ab	0.05	4.065 b	0.57	0.968 ab	0.13	5.034 ab	0.68	0.996 b	0.18
Cu 10	2.063 abc	0.81	0.033 a	4.67	2.096 a	3.92	0.761 ab	2.13	4.235 b	1.07	1.820 a	0.49	6.055 ab	1.45	0.989 b	0.20
Cu 20	1.632 bc	0.37	0.100 a	2.17	1.733 a	1.81	0.651 ab	0.98	4.881 ab	0.74	−0.175 b	1.07	4.706 b	0.34	1.343 ab	0.34
Pr > F (Model)	0.03		0.61		0.82		0.35		0.30		0.37		0.54		0.20

) The Pb treatments (8, 40, and 80 mg.L⁻¹) significantly influenced the levels of total chlorophyll (Chl a + b) and carotenoids, with a general decline in Chl a + b content observed at higher Pb concentrations, suggesting detrimental effects on photosynthesis. In contrast, carotenoids displayed relatively lower sensitivity, potentially serving a protective role against oxidative damage induced by Pb stress [37]. Although Pb is non-essential to plant metabolism, it exhibits varying degrees of phytotoxicity, particularly impacting photosynthetic processes [38]. When Pb levels exceed critical thresholds, it severely disrupts all morphological, physiological, and biochemical processes [10]. Changes in the rate of photosynthesis in Pb-exposed plants can be attributed to alterations in chloroplast ultrastructure, disruptions in chlorophyll synthesis, disturbances in plastoquinone function, decreased mineral uptake, and the obstruction of electron transport [33].

Zn treatments at various concentrations (8, 40, and 80 mg.L⁻¹) led to significant alterations in chlorophyll a + b (Chl a + b) and carotenoid levels compared to the control (C). An increase in Zn concentrations correlated with a decline in Chl a + b content, which may inhibit the synthesis of photosynthetic pigments, while carotenoid levels remained relatively stable, indicating a potentially protective role against oxidative stress induced by Zn toxicity [39]. Hydroponic experiments have demonstrated that Zn at 50 μM markedly reduces shoot dry weight, total plant biomass, and chlorophyll content in Solanum lycopersicum [40]. Furthermore, Zn at 600 μM has been shown to decrease growth rates, impair photosynthesis, and inactivate photosystem II reaction centers in Triticum durum [39].

Cu treatments at concentrations of 2, 10, and 20 mg.L⁻¹ exhibited variable effects on chlorophyll a + b (Chl a + b) and carotenoid levels. Significant reductions in Chl a + b were observed at higher Cu concentrations, indicating potential toxicity to photosynthetic pigments, while carotenoid levels showed fluctuations, suggesting a protective role in mitigating Cu-induced stress [8]. However, excessive copper concentrations in plants can be toxic and may lead to oxidative damage. Specifically, Cu concentrations of 0.2, 5.0, 25, and 50 μM inhibited the growth of Lolium perenne, resulting in reduced root and shoot biomass, as well as decreased leaf size and area [41].

Overall, the slight decrease in chlorophyll a + b levels under Pb, Zn, and Cu treatments (Table 1) in both species suggests a degree of resilience to stress. The reduction in chlorophyll content may be attributed to direct enzyme inhibition or competitive exclusion of essential nutrients [8]. The stability of carotenoid levels can be interpreted as a strategy to mitigate oxidative stress, supporting the hypothesis that these plants exhibit adaptive responses to their metallic environment [42]. Previous studies highlight that the primary role of carotenoids is to protect plant cells [43].

2.3. Pb, Zn, and Cu Concentrations in Shoots and Roots

The trial conducted in this study revealed statistically significant differences (p < 0.05) in metal accumulation between A. altissima and M. azedarach (Figure 2), underscoring their potential for phytoremediation applications.

In the aerial parts of A. altissima, there was a notable increase in Pb concentrations with rising treatment levels. At 80 mg.L⁻¹, the concentration of Pb reached 38.39 mg.kg⁻¹ in the aerial parts, while at 40 mg.L⁻¹, it was 20.75 mg.kg⁻¹. Similar trends were observed in the roots of this species, indicating that A. altissima is capable of efficiently accumulating Pb in its tissues, positioning it as a potential candidate for the phytoremediation of Pb-contaminated soils. Conversely, M. azedarach exhibited lower levels of Pb accumulation in its aerial parts, with concentrations ranging from 18.66 mg.kg⁻¹ to 24.76 mg.kg⁻¹ across different treatment concentrations. In the roots, Pb levels were also relatively low, peaking at 147.68 mg.kg⁻¹ at the highest concentration. Although M. azedarach does accumulate Pb, the observed concentrations were generally lower compared to those of A. altissima. These findings are consistent with previous studies demonstrating the potential of A. altissima as a Pb accumulator [44,45], while highlighting the moderate Pb accumulation capacity of M. azedarach [46]. Also, the finding of (Mohebzadeh et al., 2021) aligns with our results; they found that A. altissima tends to accumulate at higher concentrations of Pb in both roots (45.33 mg.kg⁻¹) and shoots (24.66 mg.kg⁻¹) compared to M. azedarach, in a pot experiment of metal-contaminated soils with Pb and Ni amended with biochar and compost [47].

Both A. altissima and M. azedarach demonstrate significant Zn accumulation capacity in their aerial parts and roots. For A. altissima, Zn concentrations can reach as high as 283.25 mg.kg⁻¹ in the aerial parts and up to 1348.56 mg.kg⁻¹ in the roots. Similarly, M. azedarach exhibits substantial Zn levels, with concentrations reaching 325.56 mg.kg⁻¹ in the aerial parts and 806.41 mg.kg⁻¹ in the roots. These results affirm the capacity of both species to efficiently accumulate Zn, which may facilitate the remediation of Zn-contaminated soils. These findings are consistent with the reported Zn hyperaccumulation potential of A. altissima [48], while also underscoring the remarkable Zn accumulation capability of M. azedarach [49].

In terms of Cu accumulation, A. altissima shows Cu concentrations reaching up to mg.kg⁻¹ in the aerial parts and up to 49.90 mg.kg⁻¹ in the roots. Đunisijević Bojović et al. (2012) found that A. altissima accumulates up to 75.7 mg.kg⁻¹ in the roots and 9.1 mg.kg⁻¹ of Cu at concentrations of 20 µM.L⁻¹ of Pb(NO₃)₂ [48]. M. azedarach accumulated concentrations reaching up to 24.28 mg.kg⁻¹ in the aerial parts and up to 204.19 mg.kg⁻¹ in the roots. These results indicate that both species are capable of efficiently accumulating Cu in the roots, which can be beneficial for the phytostabilization of Cu-contaminated soils. Previous studies have also shown that certain plant species possess a particular ability to accumulate Cu [50].

In general, the quantity of metals Pb, Zn, and Cu in the root system was greater compared to the shoot. This aligns with earlier studies, which found that most metal ions are primarily retained in the roots before being translocated to the shoots [51]. Plants selected for phytostabilization should exhibit low metal accumulation in aboveground tissues, restricted metal transfer from roots to shoots, robust canopy and root systems, rapid growth, and high resistance to metal pollution and adverse environmental conditions. Ideal candidates create dense vegetative cover, while minimizing metal concentrations in their aboveground biomass [52,53]. Based on their absorption characteristics for Pb, Zn, and Cu, these species can be considered suitable options for the remediation (phytostabilization) of contaminated soils.

2.4. Translocation Factor (TF) and Potential of Phytoremediation

Translocation factors (TF) are presented in

Table 2. Translocation factor of M. azedarach and A. altissima of Pb, Zn, and Cu.

Species	Pb 8	Pb 40	Pb 80	Zn 8	Zn 40	Zn 80	Cu 2	Cu 10	Cu 20
A. altissima	0.73	0.45	0.77	0.24	0.19	0.21	0.54	0.35	0.64
M. azedarach	0.29	0.22	0.17	0.57	0.56	0.34	0.27	0.10	0.12

. The TF values are consistently low (<1) across all treatments for the tested plant species, indicating that trace elements primarily accumulate in the roots. The highest TF values are observed for the Pb treatments (80 mg.L⁻¹ and 8 mg.L⁻¹) in A. altissima, as well as for Cu (20 mg.L⁻¹) and Zn (8 mg.L⁻¹ and 40 mg.L⁻¹) in M. azedarach. According to [53], either of the two plant species can be classified as hyperaccumulators of Pb, Zn, or Cu. These results are consistent with findings from [47], which indicated that A. altissima exhibited the highest values for the transfer factor (TF), bioconcentration factor (BCF), and root-to-shoot factor (RF) for Pb. Our findings further corroborate its superior capability to translocate Pb from the roots to the shoots.

While both species exhibited elevated concentrations of Pb, Zn, and Cu in their roots, the shoot uptake and root accumulation of Zn were higher than those of Cu and Pb in both plant species, indicating their potential for phytostabilization. A. altissima demonstrated an effective ability to limit Zn translocation from roots to shoots, reflected in lower TF values compared to M. azedarach. Conversely, M. azedarach exhibited the strong restriction of translocation from roots to shoots for Pb and Cu, with consistently lower TF values across all treatments, suggesting its suitability for the phytostabilization of these metals.

3. Materials and Methods

3.1. Plant Material

Seeds of A. altissima and M. azedarach were collected from trees growing on polymetallic mine tailings in Touissit, NE Morocco. The seeds were surface-sterilized using 10% (w/v) sodium hypochlorite and germinated in petri dishes within a phytotron at 25 °C. Young seedlings were transplanted into pots, grown in peat, and irrigated with tap water (pH 6.0) for 3 months. Plants aged 3 months, with heights ranging between 5 cm and 10 cm, were then transferred to aerated hydroponic pots filled with perlite and tap water (pH 6.0) and precultured for 9 days with Hoagland nutrient solution. The nutritional solution contained 2.5 mM Ca (NO₃)₂, 2.5 mM KNO₃, 1 mM MgSO₄, 0.2 μM KH₂PO₄, 50 μM NaFeEDTA, 0.2 μM Na₂MoO₄, 10 μM H₃BO₃, 2 μM MnCl₂, 0.5 μM CuSO4, 1.0 μM ZnSO₄, and 0.2 μM NiSO₄, adjusted to pH 6.0 using 0.5 mM MES (2-[N-morpholino] ethane sulfonic acid).

3.2. Experimental Design

A greenhouse study was conducted to assess the accumulation and resistance capabilities of A. altissima and M. azedarach for Pb, Zn, and Cu across varying concentrations. Stock solutions of Pb, Zn, and Cu were prepared using analytical grade reagents (Merck, Darmstadt, Germany) in Hoagland solution containing one of three metal salts (Pb (NO₃)₂, ZnSO₄, CuSO₄). Plants, aged 3 months and approximately 5–10 cm in height, were grown from seeds collected at the Pb-Zn mining site of Touissit in 1 L pots with perlite as the growth medium, one plant per pot. The plants were acclimatized in a hydroponic system for 9 days with Hoagland nutrient solution at 80% field capacity [54]. Subsequently, they were watered every 9 days with Hoagland’s nutrient solution containing one of three tested metal concentrations. Fresh solutions were prepared each time. Control plants received only Hoagland’s nutrient solution. The study took place in a phytotronic chamber at the Department of Biology, University Mohammed First, Oujda, Morocco, with a mean daily temperature of 25 °C and a 12 h photoperiod. The treatments were control (C) with plants only receiving Hoagland nutrient solution, Pb (8, 40, and 80 mg.L⁻¹), Zn (8, 40, and 80 mg.L⁻¹), and Cu (2, 10, and 20 mg.L⁻¹), and the nutrient solutions were 5 to 10 times higher than the maximum phyto-accessible concentrations measured (data not published) in the soils of the Touissit and Zaïda mining sites, as well as the phytotoxic limits for plants growing in these metal conditions [55]. The treatments were labeled with the chemical symbol of the metal followed by a number indicating the metal concentration in mg.L⁻¹.

3.3. Harvesting Procedure

After 90 days of metal treatment, leaf and root samples were collected from control and metal-treated plants for the estimation of various morpho-physiological, biochemical parameters. Leaves in the same position were sampled from control and trace element-treated plants to estimate the metal and photosynthetic pigment content. For each measurement or assay, 3 replicates were used.

3.4. Analytical Determinations

The parameters measured were plant growth (root and shoots dry weight), stress indicators (chlorophylls a, b, and carotenoids), and the concentration and accumulation of metals in plant tissues.

On the 90th day after the start of Pb, Zn, and Cu stress, all plant material was weighed (total dry weight), and the length of the roots and shoots measured.

The third completely grown leaf from the top was taken for chlorophyll and carotenoid analysis. To estimate photosynthetic pigments, fresh leaf blade material (0.1 g) was put in 25 mL glass test tubes, followed by 15 mL of 96% (v/v) ethanol. The plant material tubes were incubated in a water bath at 79.8 °C for three to four hours, or until the samples were completely discolored. The absorbance of chlorophylls a and b was determined at 665 and 649 nm, respectively. Total chlorophyll was determined. Carotenoid concentration was evaluated vis spectrophotometer at 470 nm [56]. Chlorophyll a and b, along with total carotenoid content, were calculated using the following equations and expressed as mg·kg⁻¹ fresh weight (fw). Where V represents the extract volume, W denotes the sample quantity, and D corresponds to the absorbance value at the specified wavelength.

$$\mathrm{Chlorophyll} \mathrm{a} =\left[12.7 \left(\mathrm{D}663\right)-2.69 \left(\mathrm{D}645\right)\right]\times \mathrm{V} ∕1000\times \mathrm{W}$$

(1)

$$\mathrm{C}\mathrm{hlorophyll} \mathrm{b}=\left[22.9 \left(\mathrm{D}645\right)-4.68 \left(\mathrm{D}663\right)\right]\times \mathrm{V} ∕1000\times \mathrm{W}$$

(2)

$$\mathrm{Carotenoids} =\left[4.69 \left(\mathrm{D}440\right)-\left(\mathrm{chlorophyll} \mathrm{a}+\mathrm{chlorophyll} \mathrm{b}\right)\times 0.286\right]\times \mathrm{V}/1000\times \mathrm{W}$$

(3)

The plant samples, including both leaves and roots, were separated and dried in an oven at 60 °C for 72 h to facilitate acid mineralization. Subsequently, the dried samples were subjected to acid digestion in a microwave system, where 0.2 g of plant material was combined with 6 mL of 65% nitric acid (HNO₃⁻) and 3 mL of 37% hydrochloric acid (HCl). The mixture was then heated for 15 min at 180 °C using a pressured vacuum microwave system (Multiwave 3000; Anton Paar GmbH, Ostfildern, Germany). This heating was followed by a 15-min stabilization period at 180 °C and an additional 15 min for cooling. After cooling to room temperature, the samples were diluted in 30 mL of ultrapure water (18 MΩ cm) and filtered through a 0.45 μm nitrocellulose membrane [57]. The concentrations of Pb, Zn, and Cu were analyzed using inductively coupled plasma atomic emission spectroscopy (ICP-AES) (ULTIMA 2, HORIBA, San Francisco, CA, USA), with a detection limit of below 100 ppb. To measure Pb, Zn, and Cu concentrations in the plant organs as described by [58].

3.5. Translocation Factor (TF)

The translocation factor (TF) is utilized to evaluate a plant’s ability to transport trace elements from the roots to the shoots, providing insights into its efficiency in accumulating these metals from the growth medium. A TF value greater than 1 signifies the effective movement of trace elements from the roots to the shoots, indicating the plant’s suitability for phyto-extraction. The TF is calculated using the following formula [59]:

$$\mathrm{TF} =\left[\mathrm{C}\right]\mathrm{metal} \mathrm{plant} \mathrm{shoot}/\left[\mathrm{C}\right]\mathrm{metal} \mathrm{plant} \mathrm{root}$$

(4)

3.6. Statistical Analysis

The experiment was executed in a factorial trial arranged in a completely randomized design (CRD) with two-factor trace elements and concentrations, with three replicates for each parameter. Chlorophyll content, carotenoid trace element accumulation, and plant growth and biomass were analyzed with two-way ANOVA (p < 0.05) using the LSD test with trace elements and concentration as factors.

4. Conclusions

This study evaluates the phytoremediation potential of A. altissima and M. azedarach for Pb, Zn, and Cu under controlled conditions. M. azedarach demonstrates notable lead tolerance, maintaining biomass under increased Pb concentrations, while A. altissima shows reduced resilience at higher Pb levels. Both species exhibit significant sensitivity to Zn and Cu, with decreased biomass across all concentrations compared to control. Analysis of photosynthetic pigments reveals a slight decline in chlorophyll content in response to Pb, Zn, and Cu treatments, suggesting some degree of resilience to metal-induced stress. According to this study, both A. altissima and M. azedarach retain higher metal concentrations in roots than shoots, meeting key criteria for effective phytostabilization. A. altissima exhibits significant Pb accumulation in roots, positioning it as an effective candidate for the phytostabilization of Pb-contaminated soils. Conversely, M. azedarach demonstrates superior Zn and Cu accumulation, highlighting its potential for stabilizing soils contaminated with these metals. This was confirmed by the translocation factors (TF < 1), which indicate a predominant accumulation of trace elements in the roots of both species meeting key criteria for effective phytostabilization. These species, with their robust metal root absorption and translocation patterns, are promising candidates for the phytostabilization of contaminated soils.