Endocrine-disrupting chemicals (EDCs) are individual compounds or mixtures that can interfere with the normal functioning of hormones, leading to various diseases or even death. Estimates suggest that more than 24% of human diseases and disorders worldwide are attributable to environmental factors, which play a role in approximately 80% of the most fatal diseases, including cancer, cardiovascular diseases, and others. Environmental EDCs can contribute to hormonal imbalances that result in diabetes, neurological disorders, reproductive disorders, inflammation, and impaired immune function [1][2]. Humans may be exposed to EDCs through a wide range of products and exposure pathways, including residues of plasticizers, pesticides, atmospheric pollutants, and industrial waste, among others [3]. Exposure to EDCs can be harmful at any age; however, it is particularly detrimental when it occurs during sensitive developmental periods, such as fetal development, childhood, adolescence, and pregnancy. Phthalates, recognized as ubiquitous EDCs, have been in use for more than 50 years, with an annual global production of approximately 6 million tonnes. They have numerous applications and are found in plastics, cosmetics, medical devices, and many other products. As a result, they are ubiquitous in everyday consumer goods, and their environmental impact is widely monitored [4]-[6]. Although these “emerging” contaminants are less persistent and less toxic than other priority substances (e.g., polychlorinated biphenyls, PCBs), their widespread presence across all environmental compartments underscores the need to reduce water and soil contamination in order to prevent the entry of these EDCs into the food chain [7].

The concept of the exposome encompasses the cumulative effects of environmental exposures on human health throughout the lifespan. Access to clean drinking water is considered a fundamental human right. Previous studies have identified more than 30 types of phthalates and their subtypes originating from various sources of contamination, including tap water, bottled water, water purification systems, and related sources [8]. As a vital component of daily intake, the abundance and diversity of phthalates in drinking water sources therefore raise concerns regarding the long-term safety of their consumption.

The World Health Organization (WHO) has established guideline values for acceptable levels of phthalates in drinking water; for example, limits of 1.3 and 3 μg/L have been set for di-(2-ethylhexyl) phthalate (DEHP) and dibutyl phthalate (DBP), respectively [9]. Studies conducted in various countries indicate that phthalate concentrations in raw water sources—and subsequently in tap water—range from 0.77 to 1438.20 μg/L [10], significantly exceeding the WHO-recommended limits. Such elevated concentrations may result in substantial lifetime exposure and accumulation of these compounds in the human body. In the face of severe water contamination, it is essential to develop effective remediation methods based on low-cost materials. Adsorption is a widely adopted technique for pollutant removal in wastewater treatment due to its relatively high efficiency, low cost, environmental friendliness, and operational simplicity [11]. The production of high-value materials such as biochar derived from agro-industrial waste represents a priority for maintaining ecological balance and is a key objective within the principles of the circular economy [12].

The properties of biochar are to some extent comparable to those of activated carbon, making it a reliable material for the removal of various impurities [13]. The selection of appropriate feedstocks and thermochemical conversion pathways is a decisive factor for achieving feasible biochar performance as an adsorbent [14]. Biochar is considered an efficient, environmentally friendly, and cost-effective material for the adsorption of typical pollutants from aqueous environments, including endocrine-disrupting chemicals [15].

The pollutant removal capacity of biochar can be further enhanced through activation or modification processes, as well as through novel combined treatment strategies aimed at effective pollution reduction [16]. The synthesis of biochar-based nanocomposites enables the combination of the advantages of biochar with those of nanomaterials. The resulting composites typically exhibit significant improvements in surface functional groups, pore structure, active surface sites, catalytic degradation capacity, and ease of separation [17].

The aim of the present study is to evaluate contamination of environmental matrices—namely water, agricultural soils, and grapes—with phthalate residues, as well as to analyze the sorption process of phthalate residues from aqueous solutions onto biochars derived from apple residues and functionalized with metal oxides (Fe₃O₄ and NiO) and plant extracts (ext). This work is intended to support the preliminary application of these materials in environmental remediation and to reduce the risks of food-chain contamination.

An assessment of phthalate levels in environmental samples was conducted based on studies of natural water sources used for industrial water supply, as well as water subjected to various treatment stages at industrial facilities, such as softening. The levels of phthalate contamination in spring and lake waters within the Chișinău municipality, Republic of Moldova, are summarized for the water bodies listed in Table 1.

Table 1. Coordinates of sampling points and content of phthalates in water sources.

N—north; S—south; E—east; W—west; DBP—dibutyl phthalate; DEHP—di-(2-ethylhexyl) phthalate.

Low concentrations of DEHP and DBP were detected in most water samples. With the exception of water from the Byc River, no elevated levels of phthalate residues were observed. Other phthalates were not detected, even in trace amounts. These results suggest that the absence of significant phthalate contamination in the water sources and lakes of Chișinău can likely be attributed to the region’s low industrial development and limited agricultural activity. Soil represents one of the primary environments for phthalate accumulation. The migration, transformation, and retention of these compounds are influenced by multiple factors, including soil properties, the composition and abundance of microorganisms, and the organic matter content. To identify potential sources of phthalate contamination in grapes, samples were collected from vineyards in the Ialoveni district and the Chișinău municipality (Table 2).

Table 2. Coordinates of sampling points and content of phthalates in soil samples.

N—north; S—south; E—east; W—west; DMP—dimethyl phthalate; DEP—diethyl phthalate; DBP—dibutyl phthalate; BBP—benzyl butyl phthalate; DEHP—di-(2-ethylhexyl) phthalate; DnOP—di-n-octylphthalate.

The results of phthalate analysis in soil revealed the presence of two compounds from this group: DBP and DEHP. DBP was detected in all soil samples, with concentrations ranging from 0.080 to 0.279 mg/kg. DEHP was detected only in samples collected from the Mileștii Mici vineyards, where its concentration substantially exceeded that of DBP. The highest total phthalate content was found in the soil from the Mileștii Mici vineyard, amounting to 0.925 mg/kg. These findings are consistent with international data [20] and confirm the presence of phthalate pollution associated with intensive agricultural activity. Previous studies [21] reported that the main phthalic acid contaminants detected in soil were DBP, DEHP, and DMP, accounting for 49.2%, 27.0%, and 12.4% of the total phthalate content, respectively. To evaluate phthalate contamination in plant material, grape samples were analyzed at harvest. The sampling locations and the DBP and DEHP content in destemmed grapes are presented in Table 3.

The experiment demonstrated that the majority of phthalate contamination in fruit occurs on the surface of the berries rather than within the bulk tissue. Ethanol rinsing (~96.4% v/v) of the fruit surface enabled the extraction of 89% - 100% of phthalates, compared with extraction from a homogenized mixture of crushed berries. These findings suggest that phthalate contamination of fruit—and consequently of the resulting must—arises not from migration through the plant’s vascular system from the soil, but rather from vineyard practices such as spraying, irrigation with wastewater, application of contaminated soil, and other anthropogenic processes. DEHP was identified as the predominant contaminant among the o-phthalate esters, while other phthalate homologues were undetectable, even in trace amounts. The results of this study confirm that contamination of plant-derived food products with phthalate esters persists and should be considered in food safety assessments, consistent with the findings of numerous previous studies [22]-[25].

Table 3. The degree of contamination of grape fruits at the harvest period.

N—north; S—south; E—east; W—west; DBP—dibutyl phthalate; BBP—benzyl butyl phthalate; DEHP—di-(2-ethylhexyl) phthalate.

To mitigate environmental contamination, which inevitably contributes to contamination of the food chain, sustainable and cost-effective sorbents were developed based on biochar derived from apple residues and functionalized with metal oxides (Fe₃O₄ and NiO), green-synthesized using plant extracts from Urtica dioicaL. Apple waste is considered biowaste and contributes to environmental pollution, as it is generated in substantial quantities during apple processing. Previous studies have demonstrated high adsorption efficiencies for paracetamol (57.54%), tartrazine (37.48%), and dibutyl phthalate (74.98%) [26].

The sorption performance of 6 experimental sorbent samples was evaluated for two phthalates, namely DBP and DEHP, since low concentrations of DEHP (0.007 - 0.057 mg/dm³) and DBP (0.005 - 0.027 mg/dm³) were detected in most of the tested water samples. The results of the analysis of phthalate residues in soil also revealed the presence of these compounds. DBP was detected in all soil samples, with concentrations ranging from 0.080 to 0.279 mg/kg. DEHP was identified as the predominant contaminant of fruits among o-phthalate esters (0.010 - 0.045 mg/kg), while other phthalate homologues were undetectable, even in trace amounts. According to the experimental results, as well as previous studies [20][21], the main contaminants of the environmental objects were DBP and DEHP.

Figure 1 illustrates examples of the reduction in DBP mass spectral signal intensity in processed water samples compared with the control. For clarity, overlapping peaks have been separated along the time axis. Each DBP and DEHP peak in the chromatogram represents the combined signal of the analyte and its deuterated internal standard. Figure 2 illustrates examples of the reduction in DEHP mass spectral signal intensity in treated water samples relative to the control. For clarity, overlapping peaks have been separated along the time axis.

Figure 1. Visualization of the relative decrease in DBP mass spectrum signals following exposure to the indicated adsorbents: blank, M1, M1-HCl, RM1-HCl, RM1-HCl/Fe₃O₄ (1:1), RM1-HCl/Fe₃O₄ (1:2), RM1-HCl/Fe₃O₄ ext.

Figure 2. Visualization of the relative decrease in DEHP mass spectral signals following exposure to the specified adsorbents: blank, M1, M1-HCl, RM1-HCl, RM1-HCl/Fe₃O₄ (1:1), RM1-HCl/Fe₃O₄ (1:2), RM1-HCl/Fe₃O₄ ext.

The difference in phthalate concentrations between the treated samples and the control was used to calculate the mass of phthalate adsorbed per corresponding mass of sorbent for each measurement. Table 4 shows the relationship between the average sorption capacities of the tested adsorbents for DBP and DEHP.

Table 4. Average sorption capacities of the tested adsorbents.

Values represent means of three replicated trials, ±standard deviation. Different letters (^a-f) designate statistically different results (p < 0.05).

The adsorption mechanism can be either physical or chemical, primarily depending on the aromatic or specific functional groups and the polarity of both the biochar and the organic pollutants. Biochar is known to strongly influence the adsorption of organic contaminants. In physical adsorption, the process is determined by intermolecular attractions and electrostatic forces between the biochar and the pollutants, whereas chemical adsorption relies on specific chemical interactions, such as π-π interactions, hydrogen bonding, and coordination bonds [16]. The sorption phenomena can be further classified according to the following mechanisms [26]: 1) hydrogen bonding, 2) electrostatic interactions, 3) hydrophobic interactions, and 4) non-covalent electron donor-acceptor π interactions. The observed differences in the adsorption behavior of DBP and DEHP are likely attributable to their distinct hydrophobicities, molecular masses, and three-dimensional structures.

Nanometal oxides exhibit high surface energy and a large specific surface area; however, their fine-grained nature often leads to aggregation and passivation. Nanometal oxide-biochar composites combine the distinct advantages of biochar and nanometal oxides, demonstrating considerable potential as adsorbents [26]. All tested sorbents displayed a narrow pore size distribution, with the primary pore size around 4 nm, attributed to the carbon component of the composites. In samples containing Fe₃O₄, a multimodal pore size distribution was observed, with larger pores ranging from 5.8 to 8 nm, likely corresponding to the Fe₃O₄ phase. In contrast, for A-ac-NiO, only 4 nm pores were detected, associated with the carbon matrix, indicating that NiO did not form large grains or a porous structure [18]. Sorption isotherms describe the relationship between the amount of adsorbed substance and its concentration in the equilibrium solution, providing insight into the interactions between the adsorbate and adsorbent (Figure 3).

(a)

(b)

(c)

Figure 3. Sorption isotherms on biochar samples: (a) A-ac; (b) A-ac-Fe₃O₄ (1:1); (c) A-ac-Fe₃O₄ (1:2). Experimental conditions: T = 298 K, pH = 6.2.

Two theoretical models are commonly used to describe sorption processes: Langmuir and Freundlich. The Langmuir model assumes that adsorption reaches a maximum corresponding to a saturated monolayer of molecules on the adsorbent surface, and is frequently applied to phthalate sorption [19][27]. This model further assumes a fixed number of adsorption sites, reversible adsorption, no interaction between adsorbed molecules, and cessation of adsorption once saturation is achieved.

The shape of sorption isotherms provides a first experimental indication of the nature of a specific adsorption phenomenon. According to the classification of isotherms into four main types—L, S, H, and C [28]—the DBP isotherms exhibited an L-shaped curve. This L-shape indicates minimal competition between the solvent and adsorbate for adsorption sites, with the longitudinal axes of adsorbed molecules lying parallel to the adsorbent surface. Experimental data were best described by the Langmuir adsorption model for biochar samples (1 - 3). The monolayer saturation capacity ranged from 35 to 56 μmol/g, and the experimentally obtained adsorption values closely matched the theoretical Langmuir predictions. Adsorption was further supported by the Freundlich model, with empirical 1/n values between 0.52 and 0.68.

For biochar samples 3 - 5, adsorption increased with temperature (Table 5), indicating the endothermic nature of the process. In contrast, samples 1 - 2 exhibited a maximum adsorption at 45˚C (318 K).

Table 5. Experimental DBP adsorption values at different temperatures (pH = 6.2).

Studies have shown that DBP and DEHP are the most frequently detected phthalate residues in environmental samples from the Republic of Moldova. Although water from Chisinau city sources generally does not raise concern—with the exception of the Byc River—significant phthalate concentrations have been detected in some vineyard soils, attributed to agricultural practices and treatments.

Analysis of grapes from various plantations during the harvest period confirmed the presence of these residues. To prevent contamination of the food chain, it is essential to develop efficient sorbents capable of reducing the cumulative effects of exposure to environmental residues on human health. Access to clean drinking water is a fundamental human right, and biochar derived from apple waste and activated with nanooxides represents a promising approach in this context.

The adsorption behavior of DBP and DEHP on the developed biochar differs considerably due to the specific molecular properties of each compound: aqueous solubility (11.2 mg/L for DBP and 0.003 mg/L for DEHP), molecular dimensions (length/width: 1.265/1.229 nm for DBP and 1.388/1.032 nm for DEHP), and octanol-water partition coefficient (4.61 for DBP and 7.5 for DEHP). Previous studies reported stronger adsorption of DEHP, attributed to its higher hydrophobicity and lower solubility in water. Similarly, DEHP was adsorbed more effectively than DBP on magnetic and non-magnetic chitosan/PVA hydrogels, with adsorption governed by hydrogen bonding, complexation, and hydrophobic interactions. In contrast, for the apple biochar examined in this study, DBP was predominantly adsorbed, likely due to steric factors and the compact structure of the DBP molecule. This observation is consistent with the L-shaped adsorption isotherms, which indicate that the molecules are arranged parallel to the biochar surface.

Multilateral project within the European Partnership “Water4All” no. 23.80013.7007.1 M “Developing innovative solution for mitigation of drought effects”, funded by the EU and NARD (Republic of Moldova).