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. 2026 Aug 18;16(45):56885–56898. doi: 10.1039/d6ra05133b

Polystyrene microplastics reduce the food chain transfer and toxicity of TiO2 nanoparticles in crustacean Artemia salina through dietary exposure of microalga Chlorella sp.: impact of visible light and ultraviolet-A radiation

Camil Rex M a, Manshi Kumari Gupta b, Chinnappan Sudandiradoss b, Amitava Mukherjee a,✉
PMCID: PMC13484639  PMID: 42614804

Abstract

Microplastics and nanoparticles are emerging contaminants in the marine system. Although UV-A radiation is environmentally relevant, most ecotoxicity studies overlook its role in modulating contaminant toxicity. Therefore, this study aimed to investigate the trophic transfer potential of titanium dioxide nanoparticles (TiO2-NPs) in the absence and presence of amine (NH2 MPs) and carboxyl (COOH MPs) functionalized polystyrene microplastics (PS-MPs) under both visible-light and UV-A radiation. The experiments were performed using marine microalgae Chlorella sp. and marine crustacean Artemia salina, representing the producer-consumer food chain. Bio-uptake of Ti in Chlorella sp. was higher under UV-A treatment than under visible-light illumination. Bio-uptake of Ti in Chlorella sp. was higher in the absence of PS-MPs. A comparable pattern was observed in the bio-uptake of Ti in A. salina following dietary exposure. Consequently, A. salina fed with microalgae pre-treated with TiO2-NPs reduces survival, enhances oxidative stress, and alters neurotransmitter activity. In contrast, in the presence of PS-MPs, these detrimental effects exhibited by TiO2-NPs were reduced. In silico studies with acetylcholinesterase provided complementary mechanistic insights into the observed neurotoxicity. Under both light treatments, the biomagnification factor for TiO2-NPs and TiO2-NPs + PS-MPs was < 1, indicating no biomagnification from Chlorella sp. to A. salina. Risk quotient analysis revealed the higher ecological risk posed by the TiO2-NPs. Pearson correlation and the cluster heatmap revealed that oxidative stress and altered neurotransmitter activity may have impaired A. salina survival. Overall, this study shows how light and microplastics modulate the trophic transfer potential and toxicity of TiO2-NPs in marine organisms.

Illustration of dietary uptake of Ti in Artemia salina under visible light and UV-A light.graphic file with name d6ra05133b-ga.webp

1. Introduction

The extensive usage of plastic products, along with their improper waste management, has significantly contributed to marine plastic pollution.1 Rivers constitute the primary pathway through which plastic waste enters the ocean, and reports indicate that around 5.25 trillion plastic particles are present in global oceans.2,3 These polymers undergo physical, chemical, and biological degradation in the marine system and lead to the formation of smaller plastics (<5 mm) known as microplastics (MPs).4 Among the various polymer types, polystyrene is one of the most prevalent and extensively used in disposable food packaging, cosmetics, medical devices, and construction materials.5 Amine-functionalized (NH2 MPs) and carboxyl-functionalized microplastics (COOH MPs) are among the most commonly used surface-functionalized MPs and are widely utilized in biosensing, drug delivery, and scientific research.6 In the marine environment, MPs possess the ability to interact and alter the fate and toxicity of their co-contaminants.7 These interactions can substantially alter the bioavailability and ecotoxicological impact of their co-contaminants, such as nanoparticles, towards marine organisms.8

Engineered nanoparticles exhibit a steady rise in worldwide production due to their extensive usage in a wide range of applications.9 Globally, titanium dioxide nanoparticles (TiO2-NPs) are among the most widely produced engineered nanoparticles, with annual production projected to reach 2.5 × 106 metric tons by 2050.10,11 TiO2-NPs possess exceptional photo-reactivity along with various other distinctive physicochemical properties.12 These remarkable properties enable TiO2-NPs to be employed in a wide range of applications, including sunscreens, paints, food processing, household products, textiles, photocatalytic technologies, and biomedical research.11,13 However, the unregulated release of TiO2-NPs into the marine system, along with environmental factors such as ultraviolet-A (UV-A) radiation, can trigger their photocatalytic activity.14 Consequently, excessive reactive oxygen species (ROS) are produced, which ultimately induce a range of toxic effects in marine organisms.15

As MPs and TiO2-NPs co-exist in aquatic systems, there is a significant risk of their transfer across trophic levels from lower to higher organisms.16,17 Marine microalgae, as primary producers, form the foundation of the marine food chain and are highly sensitive to environmental contaminants.18 Contaminants accumulated by microalgae can subsequently be transferred to primary consumers through dietary exposure.19 Therefore, in this study, the marine microalga Chlorella sp. was selected as the primary producer (prey), and the microcrustacean A. salina was chosen as the primary consumer (predator) to assess the trophic transfer and associated toxicity.

Light plays a crucial role in ecotoxicity assessment, especially when determining the toxicity of photocatalytic materials.20 Most scientific investigations utilized visible-light as the light source for trophic transfer studies, often overlooking the impact of environmentally relevant light treatments like UV-A radiation. A few studies have evaluated the effect of UV radiation in altering the waterborne toxicity of nanoparticles and MPs in aquatic organisms.20–23 No prior research has evaluated the role of UV radiation in influencing the trophic transfer of contaminants in aquatic biota. Notably, a previous study investigated the trophic transfer of P25 TiO2-NPs from marine microalgae Dunaliella salina to marine microcrustacean A. salina under visible-light treatments.24 Similarly, another study evaluated the trophic transfer potential of CuO nanoparticles from freshwater microalgae Chlorella vulgaris to freshwater crustacean Daphnia magna under visible-light treatments.25 However, investigations addressing the trophic transfer of potential nanoparticles in combination with MPs remain scarce. A prior work evaluated the dietary transfer of ZnO nanoparticles together with polystyrene MPs from freshwater microalgae Chlorella vulgaris to freshwater crustacean Daphnia magna under visible-light exposure.26

To the best of our knowledge, no previous work has studied the comparative effects of UV-A illumination and visible-light exposure on the food chain transfer potential of nanoparticles. Notably, evidence concerning the effects of UV-A radiation and visible-light on the food chain transfer potential of nanoparticles in combination with MPs remains scarce. Furthermore, most of the trophic transfer studies evaluate the bioaccumulation and biomagnification potential, often overlooking the underlying mechanism(s) of toxicity assessed through biochemical parameters. Therefore, this investigation aimed to evaluate the effect of UV-A radiation and visible-light on the trophic transfer potential of TiO2-NPs alone and their binary mixture with surface functionalized polystyrene MPs (PS-MPs). Our previous study revealed that, due to the comparable size of microalgae Chlorella sp. and the PS-MPs, PS-MPs cannot be internalized into the algal cells.23 However, PS-MPs can reduce the bioaccumulation of TiO2-NPs under both visible and UV-A light, which was also evident from our previous study.22 The present study hypothesizes that the elevated photocatalytic potential of TiO2-NPs upon UV-A radiation may induce enhanced cellular damage, thereby promoting their increased uptake in microalgal cells relative to visible-light treatment. Furthermore, this uptake of TiO2-NPs can be modulated in the presence of functionalized PS-MPs. Consequently, the trophic transfer potential of TiO2-NPs towards microcrustaceans is expected to be modulated under UV-A radiation compared to visible-light exposure. The specific objectives of this work were (i) to determine the uptake of TiO2-NPs alone and in presence of both NH2 MPs and COOH MPs in microalgae Chlorella sp. under both visible-light and UV-A radiation; (ii) to assess the trophic transfer potential of TiO2-NPs alone and in presence of both NH2 MPs and COOH MPs from microalgae to microcrustacean A. salina under both visible-light and UV-A radiation; and (iii) to elucidate the mechanism(s) of toxicity in A. salina following dietary exposed to microalgae through various biological endpoints. The endpoints include estimation of survival, ROS, malondialdehyde (MDA) levels, neurotransmitter activity, and biomagnification factor (BMF). In silico molecular docking and molecular dynamics (MD) simulation were performed for the neurotransmitter acetylcholinesterase (AChE) to understand the molecular basis for the observed neurotoxicity. Risk quotient (RQ) analysis was performed to determine the ecological risk posed by the TiO2-NPs. Pearson correlation and cluster heatmap analysis were performed to determine the association among the biological endpoints.

2. Materials and methods

2.1. Chemicals and characterization of test materials

Commercial TiO2 nanoparticles (average diameter: 21 nm) supplied by Sigma-Aldrich (USA) were used. The NH2 and COOH PS-MPs (1 µm) were procured from Corpuscular, Inc., USA. In this study, all experiments were performed in artificial seawater (ASW), and its composition is detailed in the SI (SM), Table S1. All other chemical utilized in this study is reported in the SM (Method, S1). The stock solution for TiO2-NPs (100 mg L−1) was prepared using ultrapure water and subjected to probe sonication for 30 min (Sonic, USA) to obtain a homogeneous dispersion.22 Similarly, a 100 mg L−1 stock solution was prepared for both NH2 and COOH PS-MPs, which were prepared using ultrapure water and subjected to bath sonication (Crest ultrasonics) for 15 min.22

The shape and size of the TiO2-NPs were examined using a field emission scanning electron microscope (FE-SEM). For the analysis, TiO2-NPs powder was placed on the aluminium foil-coated glass slide, and these samples were sputter-coated before imaging. The morphological characteristics and particle size of PS-MPs were determined using FE-SEM in our previous study.22 The surface charge and hydrodynamic diameter of TiO2-NPs and PS-MPs in the ASW were analysed using a particle size and zeta potential analyser (Brookhaven Instruments, USA).

2.2. Test organisms

The present study utilized marine microalgae Chlorella sp. and microcrustacean A. salina as model organisms. The Chlorella sp. was procured from the Central Marine Fisheries Research Institute (CMFRI), Tamil Nadu, India. The microalgae cultures were maintained in Conway medium under fluorescent light with a 16 h light and 8 h dark cycle. To facilitate the hatching of cysts of A. salina, filtered and autoclaved natural seawater (NSW) was utilized with a pH of 8.1 ± 0.05. To facilitate hatching, 1 g of dry cysts was introduced into 1 L filtered and autoclaved NSW, with continuous aeration and illumination using fluorescent light. After 24 h, the hatched nauplii were transferred to fresh NSW medium. For the dietary exposure experiments, the 48 h nauplii were used.

2.3. Experimental design

The trophic transfer experiments were carried out under two distinct continuous lighting (48 h) conditions at 24 ± 2 °C: (i) visible-light (Philips fluorescent lamp – 400–700 nm; 0.4 mW cm−2; exposure dose: 69.12 J cm−2; lamp distance 28 cm) and (ii) UV-A exposure (Philips UVA lamp – 320–400 nm; 0.2 mW cm−2; exposure dose: 34.56 J cm−2; lamp distance 28 cm). Initially, the microalgae Chlorella sp. (0.1 optical density) was exposed to both individual and binary mixtures of contaminants under (i) visible-light, and (ii) UV-A treatments. For individual contaminant exposure, the microalgae Chlorella sp. was exposed to two concentrations (2.5 and 10 mg L−1) of PS-MPs (NH2 MPs and COOH MPs), and one concentration of TiO2-NPs (1 mg L−1). Although internalization of PS-MPs is not possible in Chlorella sp. due to their comparable size, it was still exposed to Chlorella sp. to evaluate whether any biochemical alterations could occur in A. salina following dietary exposure. Furthermore, for the binary mixture exposure, each concentration of NH2 MPs and COOH MPs was separately combined with 1 mg L−1 TiO2-NPs. The projected environmental concentrations of TiO2-NPs have been reported up to mg L−1.27 Reports indicate that MPs levels in marine environments may reach concentrations of up to 0.1 to 100 mg L−1.28 In addition, environmentally realistic scenarios such as accidental spills may lead to a drastic increase in TiO2-NPs and MPs concentrations in the aquatic system.29,30 Therefore, the exposure concentrations selected in the present study were intended to represent worst-case yet environmentally plausible exposure scenarios. Following the exposure period (48 h), the microalgal cells that were exposed to contaminants under visible-light treatments were washed several times with fresh ASW by centrifugation (7000 rpm, 4 °C). Subsequently, only the washed cell pellets were fed to A. salina under visible-light illumination for 48 h to evaluate the trophic transfer potential of TiO2-NPs both in the absence and presence of PS-MPs. Similarly, microalgal cells exposed that were exposed to contaminants under UV-A were washed several times with fresh ASW by centrifugation (7000 rpm, 4 °C). Subsequently, only the washed cell pellets were fed to A. salina under UV-A conditions for 48 h to assess the trophic transfer potential of TiO2-NPs and their combination with PS-MPs.

2.3.1. Uptake of Ti in Chlorella sp

Ti uptake in Chlorella sp. exposed to TiO2-NPs alone and co-exposure with PS-MPs was quantified after 48 h of exposure under visible-light and UV-A irradiation. Following the exposure period, the experimental samples were subjected to centrifugation (7000 rpm; 10 min), and the pellets were washed with EDTA (0.02 M) to eliminate the surface-bound TiO2-NPs. After washing, the experimental samples were subjected to acid digestion using conc.HNO3, and the digested samples were analyzed to determine the Ti concentration using atomic absorption spectroscopy (AAS).

2.3.2. Dietary uptake, accumulation, and determination of the biomagnification factor

To determine Ti uptake in A. salina following dietary exposure, the test groups were fed with microalgae pre-treated with TiO2-NPs alone and TiO2-NPs + PS-MPs under both visible-light and UV-A treatment. After a 48 h period, the samples were washed and subjected to acid digestion with conc.HNO3, and the digested samples were analyzed using AAS to determine the Ti concentration. For the accumulation experiments, following the dietary exposure period, A. salina was transferred to a fresh ASW medium for 24 h to facilitate the depuration of TiO2-NPs. Following depuration, the nauplii were collected and subjected to acid digestion with conc.HNO3 and subjected to AAS analysis for quantification of Ti concentrations. To examine the dietary transfer of TiO2-NPs in the absence and presence of PS-MPs from microalgae to brine shrimp were determined by computing BMF using the following formula,31 BMF = concentration of Ti in consumer/concentration of Ti in producer.

2.3.3. Survival estimation of Artemia salina following dietary exposure

The survival of A. salina nauplii (n = 10) fed with microalgae pre-treated with TiO2-NPs alone, PS-MPs alone, and TiO2-NPs + PS-MPs was evaluated after a 48 h exposure period. Following the exposure period, the nauplii were subjected to microscopic analysis, and the nauplii exhibiting movements were considered surviving/alive organisms.32

2.3.4. Oxidative stress estimation in Artemia salina following dietary exposure

The total ROS levels in A. salina (n = 200), followed by dietary exposure to contaminated microalgae, were assessed using non-fluorescent dye 2′,7′-dichlorodihydrofluorescein diacetate. After the dietary exposure period (48 h), the A. salina nauplii were homogenized, centrifuged (7000 rpm; 10 min; 4 °C), and the subsequent supernatant was used for ROS measurements.33 After the dietary exposure period, the MDA levels in A. salina (n = 200) were evaluated using the TBA-TCA method.32 The detailed protocol for ROS and MDA measurements is incorporated in the SM (Method S2).

2.3.5. In vitro AChE activity and in silico molecular docking and molecular dynamics simulation of TiO2 with AChE

The AChE activity of A. salina nauplii (n = 200) fed with microalgae pre-treated with TiO2-NPs alone, PS-MPs alone, and TiO2-NPs + PS-MPs was evaluated after a 48 h exposure period. Subsequent to the exposure period, homogenization of the experimental samples were performed, and the supernatant was utilized to evaluate the AChE activity following the procedure detailed in SM (Method, S3).34

For the in silico analysis, structural models were obtained from publicly available databases.35–37 Since no experimentally resolved crystal structure of Artemia salina AChE is currently available in the Protein Data Bank, the crystal structure of AChE (PDB ID: 7RB7; resolution: 2.60 Å) was selected as a representative model for investigating the potential interaction of TiO2 with AChE at the molecular level.12 The inorganic ligand, titanium dioxide (PubChem CID: 26042), was obtained in a two-dimensional (2D) format and converted into a three-dimensional (3D) structure using Open Babel.10,11 Owing to the limitations of conventional molecular docking in representing an entire nanoparticle surface, TiO2 was modelled as a simplified inorganic ligand to provide qualitative insights into its potential interactions with the AChE binding site. The detailed molecular docking and molecular dynamics simulation procedures are provided in the SM (Method, S4).

2.4. Environmental risk quotient

Risk quotient analysis is an approach for determining the risk posed by contaminants in the aquatic system. In this study, the ecological risk posed by the TiO2-NPs was estimated using risk quotient analysis. To determine the risk quotient of TiO2-NPs, the following formula was used,

Predicted no-effect concentration = EC50/assessment factor 1
RQ = Measured environmental concentration/predicted no-effect concentration 2

Additional details of the risk quotient analysis were provided in the SM (Method, S5).

2.5. Pearson correlation, cluster heatmap, and data analysis

To evaluate the association between various biological endpoints, the Pearson correlation and cluster heatmap analyses were performed using OriginPro. The statistical analysis was performed using GraphPad Prism 8.0, and the statistical difference between the pristine and the mixture group was analyzed using a two-way ANOVA with a Bonferroni post-test. All tests were conducted in triplicate (n = 3), and the findings were reported as mean ± standard deviation.

3. Results

3.1. Characterization

The FE-SEM analysis (SM, Fig. S1) revealed that TiO2-NPs were 21 nm and spherical in morphology. Similarly, our previous study revealed that the PS-MPs were spherical in morphology and 1.9 µm.22 Fig. S2A shows the hydrodynamic size of TiO2-NPs, PS-MPs, and their co-exposure groups in ASW. PS-MPs exhibited a larger hydrodynamic size than pristine TiO2-NPs. Moreover, PS-MPs (10 mg L−1) exhibited a larger hydrodynamic size than PS-MPs (2.5 mg L−1). In contrast to TiO2-NPs, TiO2-NPs co-exposed with NH2 MPs showed a substantial increase in hydrodynamic size. A comparable pattern was observed for the TiO2-NPs co-exposed with COOH MPs. The initial and final zeta potential of TiO2-NPs, PS-MPs, and their co-exposure groups in ASW under both visible-light and UV-A exposure are provided in Fig. S2B-D. The initial zeta potential results revealed that TiO2-NPs and NH2 MPs exhibited a positive surface charge. In contrast, COOH MPs showed a negative surface charge. The co-exposure of TiO2-NPs with NH2 MPs showed less positive charge compared to TiO2-NPs. Similarly, TiO2-NPs co-exposed with COOH MPs exhibited more decrease in negative charge compared to COOH MPs. A similar trend was observed under both light conditions after 48 h, with much more reduction under UV-A treatment than under visible-light conditions.

3.2. Bio-uptake of Ti in microalgae Chlorella sp.

Fig. 1A and B show the uptake of Ti in Chlorella sp. following treatment with TiO2-NPs alone and TiO2-NPs + PS-MPs under visible-light and UV-A treatment, respectively. Exposure of TiO2-NPs alone exhibited a significant Ti uptake (p < 0.001) relative to TiO2-NPs + PS-MPs in Chlorella sp. under both visible and UV-A treatment. Under UV-A treatment, the Ti uptake in microalgae was higher with respect to visible-light following treatment with TiO2-NPs alone. Notably, Ti uptake upon exposure to TiO2-NPs + PS-MPs was higher under UV-A treatment with respect to visible-light treatment. Furthermore, the binary mixture of TiO2-NPs + 2.5 mg L−1 PS-MPs showed a significant increase in Ti uptake than the binary mixture of TiO2-NPs + 10 mg L−1 PS-MPs under both light treatments (p < 0.001). The joint exposure to TiO2-NPs + NH2 MPs showed more Ti uptake in the microalgal cells relative to the joint exposure to TiO2-NPs + COOH MPs, irrespective of the light treatments.

Fig. 1. The uptake of Ti by Chlorella sp. exposed to TiO2-NPs alone and combined with functionalized PS-MPs under (A) visible-light and (B) UV-A treatments. Statistical differences between the pristine TiO2-NPs and their binary exposure groups are represented in asterisks (***p < 0.001). The statistical differences among the co-exposure sets are indicated by α (p < 0.001).

Fig. 1

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3.3. Bio-uptake and accumulation of Ti in Artemia salina

Fig. 2A and B show the Ti uptake and accumulation in A. Salina following dietary exposure to microalgae pre-treated with TiO2-NPs alone, TiO2-NPs + PS-MPs under visible-light and UV-A exposure, respectively. The dietary exposure of pristine TiO2-NPs pre-treated microalgae exhibited more Ti uptake in A. salina under UV-A exposure than under visible-light treatments. The microalgae pre-treated with the combination of TiO2-NPs and PS-MPs showed significantly (p < 0.001) reduced Ti uptake in A. salina than those exposed to TiO2-NPs alone under both light treatments. Furthermore, dietary exposure of microalgae treated with TiO2-NPs in the presence of PS-MPs showed more Ti uptake in A. Salina under UV-A exposure than in visible-light treatments. The A. salina fed with microalgae pre-treated with TiO2-NPs + NH2 MPs showed increased Ti uptake than those fed with microalgae pre-treated with TiO2-NPs + COOH MPs. The dietary exposed organisms exhibited the following trend in the Ti uptake under both UV-A and visible-light exposure: TiO2-NPs > TiO2-NPs with PS-MPs 2.5 mg L−1 > TiO2-NPs with 10 mg L−1 PS-MPs. Regardless of light condition, the accumulation results exhibited a similar trend of Ti uptake, with a significant reduction (p < 0.001) in Ti concentrations compared to uptake.

Fig. 2. The uptake (U) and accumulation (A) of Ti in A. salina nauplii upon dietary exposure to TiO2-NPs alone and their joint exposure with PS-MPs, under (A) visible-light; (B) UV-A irradiation. The statistical differences between uptake and accumulation among test sets are represented by α (p < 0.001).

Fig. 2

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3.4. Effects on the survival of Artemia salina

Fig. 3A and B show the survival of A. salina fed with Chlorella sp. pre-treated with TiO2-NPs alone, PS-MPs alone, and their co-exposure under visible-light and UV-A light treatments, respectively. Under both visible and UV-A light treatments, dietary exposure of microalgae pre-treated with TiO2-NPs exhibited a substantial diminution in survival of A. salina with respect to the control (p < 0.001). Notably, dietary-exposed TiO2-NPs showed a more pronounced decline in the survival percentage of A. salina under UV-A light than those exposed under visible-light treatments. Nevertheless, no significant decline in the survival of the organisms was observed following dietary exposure to microalgae pre-treated with PS-MPs under both UV-A and visible-light treatments, with respect to the control (p > 0.05). The same trend was evident in organisms fed with microalgae pre-treated with TiO2-NPs + PS-MPs, under both light treatments.

Fig. 3. Survival percentage of A. salina upon dietary exposure to TiO2-NPs alone, functionalized PS-MPs alone, and their co-exposure under (A) visible-light; (B) UV-A irradiation. Statistical changes relative to control and exposure sets are represented as asterisks (***p < 0.001); the statistical differences among the individual and co-exposure sets are indicated by α: (p < 0.001) and β: (p < 0.01).

Fig. 3

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3.5. Effects on ROS generation in Artemia salina

Fig. 4A and B depict the ROS generation in A. salina following dietary exposure to Chlorella sp. pre-treated with TiO2-NPs alone, PS-MPs alone, and their co-exposure under visible-light and UV-A illumination, respectively. In contrast to the control, dietary exposure to TiO2-NPs pre-treated microalgae revealed a significant increment in ROS generation in A. salina under both light treatments (p < 0.001). Furthermore, ROS generation was more pronounced in organisms exposed to TiO2-NPs under UV-A radiation than under visible-light treatments. Nevertheless, ROS production did not differ significantly in the test organism fed with microalgae pre-treated with PS-MPs alone, and co-exposed to TiO2-NPs with 10 mg L−1 COOH MPs (p > 0.05). Compared to the test groups fed with TiO2-NPs alone-pretreated microalgae, the test groups fed with microalgae pre-treated with TiO2-NPs + PS-MPs exhibited a significant decrease in ROS generation. The dietary exposure of binary mixtures pre-treated microalgae to A. salina under UV-A treatments showed elevated ROS production than those under visible-light treatments.

Fig. 4. Changes in the ROS production in A. salina upon dietary exposure to individual TiO2-NPs alone, functionalized PS-MPs alone, and their co-exposure under (A) visible-light; (B) UV-A irradiation. Statistical changes relative to control and exposure sets are represented by asterisks (***p < 0.001 and * < 0.05); the statistical differences among the individual and co-exposure sets are represented as α: (p < 0.001), β: (p < 0.01), and γ: (p < 0.05).

Fig. 4

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3.6. Effects on the MDA levels in Artemia salina

Fig. 5A and B show the MDA content in A. salina following dietary exposure to Chlorella sp. pre-treated with TiO2-NPs alone, PS-MPs alone, and their co-exposure under visible-light and UV-A illumination, respectively. Regardless of the light treatments, all test groups showed a significant enhancement in the MDA content relative to the control sets, except for A. salina fed with microalgae pre-treated with PS-MPs alone and TiO2-NPs + 10 mg L−1 COOH MPs. The organism fed with microalgae pre-treated with TiO2-NPs under UV-A exposure exhibited higher MDA levels than the experimental groups under visible-light. A. salina fed with microalgae pre-treated with binary mixtures exhibited a significant reduction in MDA levels compared to A. salina fed with TiO2-NPs alone pre-treated microalgae (p < 0.001). The A. salina fed with microalgae pre-treated with TiO2-NPs + PS-MPs under UV-A treatments showed an increased MDA content in A. salina relative to the groups maintained under visible-light illumination.

Fig. 5. MDA level of A. salina upon dietary exposure to TiO2-NPs alone, functionalized PS-MPs alone, and their co-exposure under (A) visible-light; (B) UV-A irradiation. Statistical changes relative to control and exposure sets are represented by asterisks (***p < 0.001, **p < 0.01, and * p < 0.05); the statistical differences among the individual and co-exposure sets are represented as α (<0.001).

Fig. 5

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3.7. Effects on the AChE activity in Artemia salina

3.7.1. In vitro

Fig. 6A and B depict the AChE activity of A. salina fed Chlorella sp. that had been pre-treated with TiO2-NPs alone, PS-MPs alone, and their joint exposure under visible-light and UV-A light treatments, respectively. In contrast to the control, dietary exposure to TiO2-NPs alone revealed a significant increment in AChE activity under visible as well as UV-A light treatments (p < 0.001). Notably, the observed effects were more prominent under UV-A irradiation. In both light treatments, no significant alterations in AChE activity were observed in A. salina fed with microalgae pre-treated with PS-MPs alone and TiO2-NPs + PS-MPs, in contrast to control sets (p > 0.05).

Fig. 6. The AChE enzyme activity of A. salina upon dietary exposure to TiO2-NPs alone, functionalized PS-MPs alone, and their co-exposure under (A) visible-light; (B) UV-A irradiation. Statistical changes relative to control and exposure sets are represented by asterisks (***p < 0.001); the statistical differences among the individual and co-exposure groups are represented as α: (p < 0.001).

Fig. 6

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3.7.2. In Silico

To investigate the interaction potential of TiO2 with AChE, the active site architecture of the enzyme was initially characterized. The analysis revealed a large and well-defined binding pocket comprising 27 amino acid residues, including ALA117, GLY118, SER121, GLY122, ALA123, and ALA130. Notably, this pocket exhibited a high binding probability (0.964) along with a moderate conservation score (0.729), indicating that it is both structurally accessible and functionally relevant. Taken together, these features suggest that AChE may provide a favorable microenvironment for potential interactions with TiO2. Building on this structural insight, molecular docking was performed to evaluate the binding of TiO2 within the predicted active site. The AChE-TiO2 complex demonstrated a binding affinity of −3.43 kcal mol−1 (Fig. 7A), suggesting a potential interaction. A closer examination of the binding interface revealed that key residues, including ALA117, GLY118, SER121, GLY122, PHE295, and TYR341, were predicted to participate in the interaction with the complex through hydrogen bonding and van der Waals interactions. Importantly, these residues largely overlap with those identified in the active site prediction, thereby suggesting that TiO2 may interact within the functional core of the enzyme. This observation suggests that the predicted interaction may influence the structural dynamics of AChE.

Fig. 7. Structural and dynamic analysis of AChE in the absence and presence of TiO2 (A) Docked structure of the AChE-TiO2 complex showing the TiO2 positioned within the predicted active site pocket; (B) backbone RMSD profiles over a 100 ns molecular dynamics simulation for AChE (black) and AChE-TiO2 complex (green), indicating reduced structural deviation upon nanoparticle binding; (C) radius of gyration (Rg) as a function of simulation time, demonstrating maintained or slightly enhanced compactness of AChE in the presence of TiO2; (D) residue-wise RMSF comparison highlighting decreased atomic fluctuations in the AChE-TiO2 complex relative to the wild type, suggesting increased structural stability upon interaction with TiO2.

Fig. 7

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To further probe this possibility, molecular dynamics simulations (100 ns) were performed to evaluate the structural stability of the predicted TiO2-AChE complex. Structural deviations and flexibility were quantified using RMSD, RMSF, and radius of gyration (Rg) (Fig. 7B–D). In its native state, AChE displayed an average RMSD of ∼0.20 nm, RMSF of ∼0.18 nm, and Rg of ∼2.33 nm, reflecting a moderately flexible yet stable structure. The observed fluctuations are primarily associated with loop regions and peripheral domains, which are known to contribute to the intrinsic dynamic behavior required for enzymatic function. In contrast, upon the predicted interaction with TiO2, a consistent reduction in structural fluctuations was observed across all parameters. Specifically, the AChE-TiO2 complex exhibited a lower RMSD (∼0.15 nm) and RMSF (∼0.10 nm), indicating enhanced structural stability and reduced residue-level mobility (Fig. 7B and D). Concurrently, the Rg profile remained relatively stable with a slight tendency toward reduced values (∼2.31–2.33 nm), suggesting that the overall compactness of the protein is preserved or marginally improved (Fig. 7C). These coordinated changes collectively suggest a potential stabilization effect associated with the predicted TiO2-AChE interaction.

3.8. Biomagnification factor of TiO2-NPs in the presence and absence of PS-MPs

Table 1 shows the BMF of TiO2-NPs in the presence and absence of PS-MPs from Chlorella sp. to A. salina under both UV-A and visible-light. The highest BMF was observed for the dietary exposure of TiO2-NPs under UV-A treatments. However, there was no significant difference (p > 0.05) in the BMF of TiO2-NPs, both in the presence and absence of PS-MPs.

Table 1. Biomagnification factor of TiO2-NPs in the presence and absence of PS-MPs from Chlorella sp. to A. salina under both UV-A and visible-light illumination.

Contaminant UV-A Visible-light
TiO2-NPs 1 mg L−1 0.1 ± 0.002 0.069 ± 0.0008
2.5 mg L−1 NH2 MPs + TiO2-NPs 0.062 ± 0.0009 0.054 ± 0.0009
10 mg L−1 NH2 MPs + TiO2-NPs 0.041 ± 0.002 0.033 ± 0.003
2.5 mg L−1 COOH MPs + TiO2-NPs 0.054 ± 0.002 0.045 ± 0.001
10 mg L−1 COOH MPs + TiO2-NPs 0.029 ± 0.001 0.014 ± 0.001
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4. Discussion

TiO2-NPs and PS-MPs are widely distributed in marine environments.38,39 Although the selected exposure concentrations are higher than the commonly reported concentrations, these concentrations can be plausible exposure scenarios in localized contamination hotspots and during accidental release.27–30 The bioaccumulation of contaminants can be influenced by various factors, including light, contaminant size, concentration, co-pollutants, and various other physicochemical properties of the contaminants.40–42 TiO2-NPs are highly photoreactive under UV-A exposure due to their wide band gap energy, which enables excessive electron–hole pair generation and consequently augments ROS generation.43 In the present study, the bio-uptake of Ti was higher in microalgal cells under UV-A illumination relative to visible-light. This increment could be attributed to the excess generation of free radicals and associated cell membrane damage, which may have facilitated higher Ti bio-uptake in microalgal cells under UV-A treatments. A previous study has also reported that under UV-A treatments, TiO2-NPs induced substantial cell damage to marine microalgae Phaeodactylum tricornutum.21 Our previous study revealed that the microalgae used in this study, Chlorella sp., cannot internalize PS-MPs because of their comparable size.23 However, plastics can influence the bioavailability of their co-contaminants due to their exceptional sorption properties.44 The current study revealed that the bio-uptake of Ti was higher in algal cells on exposure to TiO2-NPs alone than in combination with PS-MPs. This could be attributed to the reduced bioavailability of TiO2-NPs in the presence of PS-MPs, which was also evident from our previous study.22 Similarly, a previous investigation revealed that MPs reduced the bioavailability of ZnO nanoparticles towards marine microalgae Tetraselmis helgolandica.44 Furthermore, this study exhibited a more reduction in the Ti bio-uptake into the algal cells on exposure to COOH MPs + TiO2-NPs than NH2 MPs + TiO2-NPs. This comparable reduction in the bio-uptake can be associated with the hetero-aggregation of the pollutant mixture, attributed to the strong electrostatic attraction between oppositely charged COOH MPs and TiO2-NPs.22

In the marine system, microalgae are exposed to a wide range of contaminants.45 Consequently, these contaminants can be transferred to various higher trophic level organisms through the dietary uptake of contaminated microalgae. The present study revealed the trophic transfer of TiO2-NPs from marine microalgae Chlorella sp. to marine crustacean A. salina. In accordance with our findings, a prior work revealed the trophic transfer potential of TiO2-NPs from freshwater microalgae Chlorella pyrenoidosa to freshwater crustacean Daphnia magna.46 As expected, the dietary exposure of pristine TiO2-NPs pre-treated algal cells showed higher Ti bio-uptake in A. salina when compared to the dietary exposure of binary mixture pre-treated algal cells. Furthermore, pristine and binary mixture pre-treated algal cells under UV-A exposure showed higher Ti bio-uptake in A. salina than those under visible-light treatments. These trends explicitly revealed that Ti bio-uptake patterns in Chlorella sp. are strongly in alignment with dietary bio-uptake of Ti in Artemia salina. Consequently, A. salina fed with algal cells pre-treated with TiO2-NPs alone exhibited a significant reduction in survival under both light treatments, with a more pronounced decrease observed under UV-A exposure. Though the binary mixture pre-treated algal cells showed uptake in A. salina, there was no significant alteration in their survival. This indicates that the uptake levels of Ti may not be sufficient to induce mortality but may induce sublethal toxicity.

ROS and MDA are important markers for the oxidative stress, especially for contaminants with photocatalytic activity.47 The current study showed enhanced ROS production in A. salina followed by their dietary exposure to microalgal cells pretreated with TiO2-NPs alone under both light treatments. Notably, the pronounced ROS production in A. salina fed with microalgal cells pre-treated with TiO2-NPs under UV-A exposure may be associated with more uptake of Ti in algal cells than under visible-light treatments. This excessive ROS production can lead to membrane damage in A. salina, which was evident from MDA level estimation. A prior study reported that MDA production was increased in the fish Clarias gariepinus fed with TiO2-NPs pre-treated with cyclopoid copepod sp.48 Interestingly, though the survival of A. salina fed with the binary mixture pre-treated microalgae was not affected, oxidative stress production was observed. This indicated the sub-lethal toxicity of dietary-exposed TiO2-NPs + PS-MPs. Furthermore, the insignificant oxidative stress exhibited by the dietary-exposed TiO2-NPs + 10 mg L−1 COOH MPs could be attributed to the low levels of Ti uptake, which may not have been sufficient to induce oxidative stress. Overall, the findings of ROS generation were strongly consistent with those of MDA production.

Elevated oxidative stress may be associated with alterations in neurotransmitter activity in A. salina. AChE is a widely used biomarker for neurotoxicity, as it is associated with cholinergic neurotransmission.49 It is mainly responsible for behavioral changes, locomotory functions, and can ultimately impact the survival of the organism.50,51 In this study, the findings of AChE activity revealed that dietary exposure to TiO2-NPs alone affected AChE activity, which also aligned with survival results. Furthermore, in silico docking and MD simulations were performed to provide complementary mechanistic insights into the observed neurotoxicity of TiO2-NPs in A. salina. Molecular docking suggests that TiO2 may interact with the predicted active site of AChE, with a binding affinity of −3.43 kcal mol−1. This interaction involves critical residues such as PHE295 and TYR341, which are located within the acyl pocket and the peripheral anionic site. These sites are essential for regulating the entry of acetylcholine into the catalytic gorge, and their predicted interaction with TiO2 suggests a potential influence on natural enzymatic turnover.52–55

The 100 ns MD-simulation further explains this mechanism. In its native state, AChE maintains a dynamic, flexible structure (RMSD ∼0.20 nm) necessary for its high catalytic rate. However, the predicted interaction with TiO2 was associated with a “stabilization” effect, reducing fluctuations to ∼0.15 nm and decreasing the Radius of Gyration. This reduction in conformational flexibility may represent a structural “locking” of the enzyme. While increased stability might seem beneficial for an enzyme, excessive rigidity often alters substrate-binding kinetics. Taken together, the combined docking and simulation results provide a coherent picture in which TiO2 interaction stabilizes AChE by restricting its conformational flexibility. Mechanistically, the simulations suggest that this stabilization may arise from sustained interactions within the active site that limit local motions and dampen backbone fluctuations. Consequently, the protein may adopt a more rigid and structurally cohesive conformation, which may have direct implications for its catalytic activity. Thus, results suggest that TiO2 may modulate the structural and functional behaviour of AChE. Overall, these findings provide complementary and plausible mechanistic insights into the observed neurotoxicity.

Fig. 8A and B show the cluster heatmap of the biochemical parameters under visible-light and UV-A treatment, respectively. The cluster heatmap revealed that oxidative stress parameters, such as ROS and MDA levels, were closely related to each other under both visible and UV-A treatment. Furthermore, these oxidative stress parameters were closely related to neurotransmitter (AChE) activity under both light treatments. Notably, the analysis revealed that the oxidative stress parameters and neurotransmitter activity were clustered with the survival of A. salina.Fig. 8C and D show the Pearson correlation results of the biochemical parameters under visible-light and UV-A illumination, respectively. The Pearson correlation results revealed that survival of the organism was negatively correlated with ROS and MDA content in organisms exposed to either visible-light or UV-A illumination. Similarly, AChE activity displayed a negative correlation with survival under both light treatments. Furthermore, under both light treatments, the ROS generated positively correlated with MDA levels in A. salina. Likewise, ROS and MDA production were positively correlated with the AChE activity of the organism under both light treatments. All these results revealed that ROS generation is one of the pivotal modes of action for the detrimental effects in A. salina, irrespective of the light treatments. This enhanced ROS generation triggered lipid peroxidation, which was evident from MDA production. These heightened oxidative stress levels may be associated with altered neurotransmitter activity in A. salina. Ultimately, oxidative damage and alterations in AChE activity may affect the survival of A. salina.

Fig. 8. (A) Heatmap analysis of the biological endpoints under visible-light conditions; (B) heatmap analysis of the biological endpoints under UV-A irradiation; (C) Pearson correlation of the biological endpoints under visible-light conditions; (D) Pearson correlation of the biological endpoints under UV-A irradiation.

Fig. 8

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The depuration experiment revealed lower bioaccumulated Ti levels compared to dietary Ti uptake. This could be attributed to the ability of the organism to eliminate Ti, and it may influence the biomagnification potential. BMF is a quantitative index to evaluate the degree of transfer of a contaminant from a prey to a predator in a food chain.56 Notably, both TiO2-NPs alone and their co-exposure with PS-MPs exhibited lower BMF (<1), indicating no biomagnification for TiO2-NPs alone or TiO2-NPs + PS-MPs from Chlorella sp. to A. salina under the present experimental conditions. This could be attributed to the lower concentration of TiO2-NPs used in this study.

Table 2 shows the estimated RQ of TiO2-NPs in the marine crustacean A. salina. The calculated RQ is a screening-level estimate and is subject to uncertainty associated with extrapolating laboratory-derived toxicity data to environmental conditions using literature-reported environmental concentrations. RQ analysis provides a critical evaluation of the ecological risk posed by the contaminants towards aquatic organisms in the natural environment.57 The contaminants exhibiting an RQ value <0.1 are regarded as low ecological risk, <1 is moderate ecological risk, and ≥1 is regarded as high ecological risk. Our findings from the RQ analysis revealed that TiO2-NPs pose a high ecological threat to A. salina under environmentally relevant concentrations in the marine environment.

Table 2. Estimated risk quotient of TiO2-NPs in marine crustacean Artemia salina.

Sl. No Area MEC (µg L−1) RQ References
1 Laizhou Bay 200 34.5 58
2 Jiaozhou Bay 308 53.1 58
3 Mediterranean shores 50 8.6 59
4 Coastal waters of Mallorca 37.6 6.5 60
5 French Mediterranean coast 20 (lower level) 3.4 61
900 (higher level) 155.1
6 Mediterranean coast 100 (lower level) 17.2 58
900 (higher level) 155.1
7 Jiaozhou Bay sediments 0.697 (lower level) 0.12 62
2.44 (higher level) 0.42
8 Mediterranean waters near Marseille 20 (lower level) 3.44 60
900 (higher level) 155.1
9 Coastal waters in Majorca beaches 12.1 (lower level) 2.06 63
37.6 (higher level) 6.48
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5. Conclusion

The findings of this study revealed that exposure to TiO2-NPs alone resulted in the highest Ti uptake in microalgae Chlorella sp. compared to TiO2-NPs + PS-MPs. Furthermore, the bio-uptake of Ti was more in Chlorella sp. under UV-A than under visible-light treatments. These trends were well aligned with Ti bio-uptake in A. salina following dietary exposure to contaminated microalgae. A. salina fed with microalgae pre-treated with TiO2-NPs alone showed a significant decline in survival, with a more pronounced decline under UV-A treatments. The microalgae exposed to TiO2-NPs alone and in combination with PS-MPs, showed elevated ROS and MDA levels in A. salina, with a more pronounced elevation observed in the TiO2-NPs alone dietary exposed group. This enhanced oxidative stress may be associated with impaired AChE activity, and the concurrent impact of these two parameters may adversely impact the survival of dietary-exposed A. salina. Under both light treatments, the BMF was less (<1) for TiO2-NPs in both the presence and absence of PS-MPs, indicating reduced risk of biomagnification. RQ analysis revealed a high risk potential of TiO2-NPs in the marine system. Overall, this study elucidates the significance of light treatments and the presence of co-contaminants (PS-MPs) in modulating the trophic transfer potential of TiO2-NPs from microalgae to brine shrimp. Future studies should investigate how environmental factors such as ocean acidification, ocean warming, and natural organic matter influence contaminant behavior and toxicity. Furthermore, future studies can extend this investigation to higher trophic level organisms like fish to better understand their potential risk in the marine food chain.

Author contributions

Camil Rex M: investigation, methodology, visualization, conceptualization, formal analysis, and writing – original draft; Manshi Kumari Gupta: methodology and writing – original draft; Chinnappan Sudandiradoss: methodology, validation, formal analysis, and writing – review and editing; Amitava Mukherjee: conceptualization, methodology, supervision, validation, formal analysis, project administration, writing – review and editing.

Conflicts of interest

There are no conflicts to declare.

Supplementary Material

RA-016-D6RA05133B-s001

Acknowledgments

The authors would like to acknowledge Vellore Institute of Technology (VIT) for funding and providing the Field Emission Scanning Electron Microscopy (FE-SEM) facility.

Data availability

The data supporting this article have been included as part of the supplementary information (SI). Supplementary information is available. See DOI: https://doi.org/10.1039/d6ra05133b.

References

  1. Ru J.-C. Zhao X.-L. Cao Z.-H. Chen C.-Z. Li P. Li Z.-H. Chronic toxic effects of polystyrene micro-plastics, DCOIT and their combination on marine Chlorella sp. Comp. Biochem. Physiol., Part C:Toxicol. Pharmacol. 2022;261:109426. [Google Scholar]
  2. Pal D. Prabhakar R. Barua V. B. Zekker I. Burlakovs J. Krauklis A. Hogland W. Vincevica-Gaile Z. Microplastics in aquatic systems: A comprehensive review of its distribution, environmental interactions, and health risks. Environ. Sci. Pollut. Res. 2025;32:56–88. doi: 10.1007/s11356-024-35741-1. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Jalaudin Basha N. N. Adzuan Hafiz N. B. Osman M. S. Abu Bakar N. F. Unveiling the noxious effect of polystyrene microplastics in aquatic ecosystems and their toxicological behavior on fishes and microalgae. Front. Toxicol. 2023;5:1135081. doi: 10.3389/ftox.2023.1135081. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Siddiqui S. A. Singh S. Bahmid N. A. Shyu D. J. Domínguez R. Lorenzo J. M. Pereira J. A. Câmara J. S. Polystyrene microplastic particles in the food chain: Characteristics and toxicity-A review. Sci. Total Environ. 2023;892:164531. doi: 10.1016/j.scitotenv.2023.164531. [DOI] [PubMed] [Google Scholar]
  5. Singh A. Chauhan A. Gaur R. A comprehensive review on the synthesis, properties, environmental impacts, and chemiluminescence applications of polystyrene (PS) Discov. chem. 2025;2:47. doi: 10.1007/s44371-025-00125-y. [DOI] [Google Scholar]
  6. Bergami E. Pugnalini S. Vannuccini M. L. Manfra L. Faleri C. Savorelli F. Dawson K. A. Corsi I. Long-term toxicity of surface-charged polystyrene nanoplastics to marine planktonic species Dunaliella tertiolecta and Artemia franciscana. Aquat. Toxicol. 2017;189:159–169. doi: 10.1016/j.aquatox.2017.06.008. [DOI] [PubMed] [Google Scholar]
  7. Bhaumik S. Chakraborty P. Interactions between microplastics (MPs) and trace/toxic metals in marine environments: Implications and insights—A comprehensive review. Environ. Sci. Pollut. Res. 2024;31:59681–59699. doi: 10.1007/s11356-024-34960-w. [DOI] [PubMed] [Google Scholar]
  8. Kalčíková G. Roy R. Klun B. Rozman U. Marolt G. Skalar T. Feckler A. Bundschuh M. Interactions between titanium dioxide nanoparticles and polyethylene microplastics: Adsorption kinetics, photocatalytic properties, and ecotoxicity. Chemosphere. 2023;329:138628. doi: 10.1016/j.chemosphere.2023.138628. [DOI] [PubMed] [Google Scholar]
  9. Khan W. S., Asmatulu E. and Asmatulu R., in Nanotechnology Safety, Elsevier, 2025, pp. 1–21 [Google Scholar]
  10. Ishitani Y. Ciacci C. Ujiié Y. Tame A. Tiboni M. Tanifuji G. Inagaki Y. Frontalini F. Fascinating strategies of marine benthic organisms to cope with emerging pollutant: Titanium dioxide nanoparticles. Environ. Pollut. 2023;330:121538. doi: 10.1016/j.envpol.2023.121538. [DOI] [PubMed] [Google Scholar]
  11. Javed R. Ain N. U. Gul A. Arslan Ahmad M. Guo W. Ao Q. Tian S. Diverse biotechnological applications of multifunctional titanium dioxide nanoparticles: An up-to-date review. IET Nanobiotechnol. 2022;16:171–189. doi: 10.1049/nbt2.12085. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Gonçalves S. G. Milanez P. R. de Souza Pereira M. Benetti R. M. Bellettini G. C. Elyseu F. Dal-Bó A. G. Bernardin A. M. Sensitization of TiO2 NPs with curcumin and chlorella and photocatalytic performance for degradation of phenol and organic dyes. Chem. Eng. Sci. 2024;285:119598. doi: 10.1016/j.ces.2023.119598. [DOI] [Google Scholar]
  13. Bevacqua E. Occhiuzzi M. A. Grande F. Tucci P. TiO2-NPs toxicity and safety: an update of the findings published over the last six years. Mini Rev. Med. Chem. 2023;23:1050–1057. doi: 10.2174/1389557522666220929152403. [DOI] [PubMed] [Google Scholar]
  14. Sanchez-Campos Y. Cárcamo-Fincheira P. González-Villagra J. Jorquera-Fontena E. Acevedo P. Soto-Cerda B. Nunes-Nesi A. Inostroza-Blancheteau C. Tighe-Neira R. Physiological and molecular effects of TiO2 nanoparticle application on UV-A radiation stress responses in Solanum lycopersicum L. Physiological and molecular effects of TiO2 nanoparticle application on UV-A radiation stress responses in Solanum lycopersicum L. Protoplasma. 2023;260:1527–1537. doi: 10.1007/s00709-023-01868-0. [DOI] [PubMed] [Google Scholar]
  15. Yang L. Xu J. Gao H. Dai S. Liu L. Xi Y. Zhang G. Wen X. Toxicity enhancement of nano titanium dioxide to Brachionus calyciflorus (Rotifera) under simulated sunlight and the underlying mechanisms. Ecotoxicol. Environ. Saf. 2023;251:114556. doi: 10.1016/j.ecoenv.2023.114556. [DOI] [PubMed] [Google Scholar]
  16. Timerbaev A. R. Kuznetsova O. V. Keppler B. K. Current trends and challenges in analysis and characterization of engineered nanoparticles in seawater. Talanta. 2021;226:122201. doi: 10.1016/j.talanta.2021.122201. [DOI] [PubMed] [Google Scholar]
  17. Wang Z. Kong Y. Cao X. Liu N. Wang C. Li X. Xing B. Co-photoaging inhibited the heteroaggregation between polystyrene nanoplastics and different titanium dioxide nanoparticles. Water Res. 2024;259:121831. doi: 10.1016/j.watres.2024.121831. [DOI] [PubMed] [Google Scholar]
  18. Yu L. Xia W. Du H. The toxic effects of petroleum pollutants to microalgae in marine environment. Mar. Pollut. Bull. 2024;201:116235. doi: 10.1016/j.marpolbul.2024.116235. [DOI] [PubMed] [Google Scholar]
  19. Mehner T. Attermeyer K. Brauns M. Brothers S. Hilt S. Scharnweber K. van Dorst R. M. Vanni M. J. Gaedke U. Trophic transfer efficiency in lakes. Ecosystems. 2022;25:1628–1652. doi: 10.1007/s10021-022-00776-3. [DOI] [Google Scholar]
  20. Zhu L. Booth A. M. Feng S. Shang C. Xiao H. Tang X. Sun X. Zhao X. Chen B. Qu K. UV-B radiation enhances the toxicity of TiO 2 nanoparticles to the marine microalga Chlorella pyrenoidosa by disrupting the protection function of extracellular polymeric substances. Environ. Sci.: Nano. 2022;9:1591–1604. doi: 10.1039/D1EN01198G. [DOI] [Google Scholar]
  21. Sendra M. Moreno-Garrido I. Yeste M. P. Gatica J. M. Blasco J. Toxicity of TiO2, in nanoparticle or bulk form to freshwater and marine microalgae under visible light and UV-A radiation. Environ. Pollut. 2017;227:39–48. doi: 10.1016/j.envpol.2017.04.053. [DOI] [PubMed] [Google Scholar]
  22. Rex M. C. Mukherjee A. Impact of UVA and visible light conditions in modulating the toxicity of binary mixture of polystyrene micro plastics and TiO2 nanoparticles in Brine Shrimp (Artemia salina) J. Hazard. Mater. Adv. 2025:100848. [Google Scholar]
  23. Rex M C. Mukherjee A. The comparative effects of visible light and UV-A radiation on the combined toxicity of P25 TiO2 nanoparticles and polystyrene microplastics on Chlorella sp. Environ. Sci. Pollut. Res. 2023;30:122700–122716. doi: 10.1007/s11356-023-30910-0. [DOI] [PubMed] [Google Scholar]
  24. Bhuvaneshwari M. Thiagarajan V. Nemade P. Chandrasekaran N. Mukherjee A. Toxicity and trophic transfer of P25 TiO2 NPs from Dunaliella salina to Artemia salina: effect of dietary and waterborne exposure. Environ. Res. 2018;160:39–46. doi: 10.1016/j.envres.2017.09.022. [DOI] [PubMed] [Google Scholar]
  25. Wu F. Bortvedt A. Harper B. J. Crandon L. E. Harper S. L. Uptake and toxicity of CuO nanoparticles to Daphnia magna varies between indirect dietary and direct waterborne exposures. Aquat. Toxicol. 2017;190:78–86. doi: 10.1016/j.aquatox.2017.06.021. [DOI] [PubMed] [Google Scholar]
  26. Guo J. Liu N. Xie Q. Zhu L. Ge F. Polystyrene microplastics facilitate the biotoxicity and biomagnification of ZnO nanoparticles in the food chain from algae to daphnia. Environ. Pollut. 2023;324:121181. doi: 10.1016/j.envpol.2023.121181. [DOI] [PubMed] [Google Scholar]
  27. Sun B. Hu M. Bock C. Shao Y. Chen H. Waiho K. Liu W. Khadka K. Xu C. Wang Y. Effects of perfluorooctanoic acid and nano titanium dioxide on the immune response and energy allocation in Mytilus coruscus. Chemosphere. 2025;370:143958. doi: 10.1016/j.chemosphere.2024.143958. [DOI] [PubMed] [Google Scholar]
  28. Pencik O. Durdakova M. Molnarova K. Kucsera A. Klofac D. Kolackova M. Adam V. Huska D. Microplastics and nanoplastics toxicity assays: A revision towards to environmental-relevance in water environment. J. Hazard. Mater. 2023;454:131476. doi: 10.1016/j.jhazmat.2023.131476. [DOI] [PubMed] [Google Scholar]
  29. Loosli F. Wang J. Rothenberg S. Bizimis M. Winkler C. Borovinskaya O. Flamigni L. Baalousha M. Sewage spills are a major source of titanium dioxide engineered (nano)-particle release into the environment. Environ. Sci.: Nano. 2019;6:763–777. doi: 10.1039/C8EN01376D. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Li W. C. Tse H. F. Fok L. Plastic waste in the marine environment: A review of sources, occurrence and effects. Sci. Total Environ. 2016;566:333–349. doi: 10.1016/j.scitotenv.2016.05.084. [DOI] [PubMed] [Google Scholar]
  31. Wang Z. Yin L. Zhao J. Xing B. Trophic transfer and accumulation of TiO2 nanoparticles from clamworm (Perinereis aibuhitensis) to juvenile turbot (Scophthalmus maximus) along a marine benthic food chain. Water Res. 2016;95:250–259. doi: 10.1016/j.watres.2016.03.027. [DOI] [PubMed] [Google Scholar]
  32. Rex M C. Bairoliya A. Gairola S. Sureshkumar P. Niranjan V. Nag A. Mukherjee A. Engineered biocorona on microplastics as a toxicity mitigation strategy in marine environment: Experiments with a marine crustacean Artemia salina. J. Hazard. Mater. Adv. 2025;17:100558. [Google Scholar]
  33. Athulya P. A. Sunil Z. Manzo S. Chandrasekaran N. Prepared microplastics interaction with Artemia salina under low pH conditions representing ocean acidification; a simulated environmental exposure. J. Environ. Manage. 2023;348:119367. doi: 10.1016/j.jenvman.2023.119367. [DOI] [PubMed] [Google Scholar]
  34. Cavion F. Fusco L. Sosa S. Manfrin C. Alonso B. Zurutuza A. Della Loggia R. Tubaro A. Prato M. Pelin M. Ecotoxicological impact of graphene oxide: Toxic effects on the model organism Artemia franciscana. Environ. Sci.: Nano. 2020;7:3605–3615. doi: 10.1039/D0EN00747A. [DOI] [Google Scholar]
  35. Vallat B. Rose Y. Piehl D. W. Duarte J. M. Bittrich S. Bi C. Segura J. Zalevsky A. Sekharan M. R. Webb B. M. RCSB Protein Data Bank: Delivering integrative structures alongside experimental structures and computed structure models. Nucleic Acids Res. 2026;54:D489–D498. doi: 10.1093/nar/gkaf1187. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Kim S. Thiessen P. A. Bolton E. E. Chen J. Fu G. Gindulyte A. Han L. He J. He S. Shoemaker B. A. PubChem substance and compound databases. Nucleic Acids Res. 2016;44:D1202–D1213. doi: 10.1093/nar/gkv951. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. O'Boyle N. M. Banck M. James C. A. Morley C. Vandermeersch T. Hutchison G. R. Open Babel: An open chemical toolbox. J. Cheminf. 2011;3:33. [Google Scholar]
  38. Wang Y. Zhu X. Lao Y. Lv X. Tao Y. Huang B. Wang J. Zhou J. Cai Z. TiO2 nanoparticles in the marine environment: physical effects responsible for the toxicity on algae Phaeodactylum tricornutum. Sci. Total Environ. 2016;565:818–826. doi: 10.1016/j.scitotenv.2016.03.164. [DOI] [PubMed] [Google Scholar]
  39. De-la-Torre G. E. Dioses-Salinas D. C. Pizarro-Ortega C. I. Saldaña-Serrano M. Global distribution of two polystyrene-derived contaminants in the marine environment: a review. Mar. Pollut. Bull. 2020;161:111729. doi: 10.1016/j.marpolbul.2020.111729. [DOI] [PubMed] [Google Scholar]
  40. Long S. Hamilton P. B. Wang C. Li C. Xue X. Zhao Z. Wu P. Gu E. Uddin M. M. Li B. Bioadsorption, bioaccumulation and biodegradation of antibiotics by algae and their association with algal physiological state and antibiotic physicochemical properties. J. Hazard. Mater. 2024;468:133787. doi: 10.1016/j.jhazmat.2024.133787. [DOI] [Google Scholar]
  41. Gomes M. Juneau P. Temperature and light modulation of herbicide toxicity on algal and cyanobacterial physiology. Front. Environ. Sci. 2017;5 [Google Scholar]
  42. Chen Y. Ling Y. Li X. Hu J. Cao C. He D. Size-dependent cellular internalization and effects of polystyrene microplastics in microalgae P. helgolandica var. tsingtaoensis and S. quadricauda. J. Hazard. Mater. 2020;399:123092. doi: 10.1016/j.jhazmat.2020.123092. [DOI] [PubMed] [Google Scholar]
  43. Kumar A. Choudhary P. Kumar A. Camargo P. H. Krishnan V. Recent advances in plasmonic photocatalysis based on TiO2 and noble metal nanoparticles for energy conversion, environmental remediation, and organic synthesis. Small. 2022;18:2101638. doi: 10.1002/smll.202101638. [DOI] [PubMed] [Google Scholar]
  44. Li J. Mao S. Ye Y. Lü J. Jing F. Guo Y. Liu H. Wang P. Ma W. Qi P. Micro-polyethylene particles reduce the toxicity of nano zinc oxide in marine microalgae by adsorption. Environ. Pollut. 2021;290:118042. doi: 10.1016/j.envpol.2021.118042. [DOI] [PubMed] [Google Scholar]
  45. Le V.-G. Nguyen M.-K. Nguyen H.-L. Thai V.-A. Le V.-R. Vu Q. M. Asaithambi P. Chang S. W. Nguyen D. D. Ecotoxicological response of algae to contaminants in aquatic environments: a review. Environ. Chem. Lett. 2024;22:919–939. doi: 10.1007/s10311-023-01680-5. [DOI] [Google Scholar]
  46. Chen X. Zhu Y. Yang K. Zhu L. Lin D. Nanoparticle TiO2 size and rutile content impact bioconcentration and biomagnification from algae to daphnia. Environ. Pollut. 2019;247:421–430. doi: 10.1016/j.envpol.2019.01.022. [DOI] [PubMed] [Google Scholar]
  47. Takhar V. Singh S. Nanomaterials ROS: a comprehensive review for environmental applications. Environ. Sci.: Nano. 2025;12:2516–2550. doi: 10.1039/D5EN00049A. [DOI] [Google Scholar]
  48. Perbiche-Neves G. Pomari J. Serafim-Junior M. Nogueira M. G. Cyclopoid copepods as indicators of trophic level in South American reservoirs: A new perspective at species level based on a wide spatial-temporal scale. Ecol. Indic. 2021;127:107744. doi: 10.1016/j.ecolind.2021.107744. [DOI] [Google Scholar]
  49. Kim L. Kim H. Song Y. An Y.-J. Chronic effects of irregular and fibril microplastics on Artemia franciscana in a benthic environment: Size and shape-dependent toxicity. Mar. Pollut. Bull. 2024;209:117270. doi: 10.1016/j.marpolbul.2024.117270. [DOI] [PubMed] [Google Scholar]
  50. Eom H.-J. Nam S.-E. Rhee J.-S. Polystyrene microplastics induce mortality through acute cell stress and inhibition of cholinergic activity in a brine shrimp. Mol. Cell. Toxicol. 2020;16:233–243. doi: 10.1007/s13273-020-00088-4. [DOI] [Google Scholar]
  51. Kukavica B. Davidović-Plavšić B. Savić A. Dmitrović D. Šukalo G. Đurić-Savić S. Vučić G. Oxidative stress and neurotoxicity of cadmium and zinc on Artemia franciscana. Biol. Trace Elem. Res. 2023;201:2636–2649. doi: 10.1007/s12011-022-03352-x. [DOI] [PubMed] [Google Scholar]
  52. Aschner M. Skalny A. Santamaria A. Buha-Đorđević A. Tizabi Y. Jiang Y. Lu R. Virgolini M. B. Tinkov A. A. From mechanisms to implications: understanding the molecular neurotoxicity of titanium dioxide nanoparticles. Front. Biosci. 2023;28:1–21. doi: 10.31083/j.fbl2809204. [DOI] [PubMed] [Google Scholar]
  53. Suthar J. K. Vaidya A. Ravindran S. Size, surface properties, and ion release of zinc oxide nanoparticles: effects on cytotoxicity, dopaminergic gene expression, and acetylcholinesterase inhibition in neuronal PC-12 cells. Biol. Trace Elem. Res. 2024;202:2254–2271. doi: 10.1007/s12011-023-03832-8. [DOI] [PubMed] [Google Scholar]
  54. Ranjan S. Dasgupta N. Chinnappan S. Ramalingam C. Kumar A. A novel approach to evaluate titanium dioxide nanoparticle–protein interaction through docking: an insight into mechanism of action. Proc. Natl. Acad. Sci. India B Biol. Sci. 2017;87:937–943. doi: 10.1007/s40011-015-0673-z. [DOI] [Google Scholar]
  55. Ranjan S. Dasgupta N. Sudandiradoss C. Ramalingam C. Kumar A. Titanium dioxide nanoparticle–protein interaction explained by docking approach. Int. J. Nanomed. 2018;13:47–50. doi: 10.2147/IJN.S125008. [DOI] [PMC free article] [PubMed] [Google Scholar]
  56. Wang Z. Li Y. Kong F. Li M. Xi M. Yu Z. How do trophic magnification factors (TMFs) and biomagnification factors (BMFs) perform on toxic pollutant bioaccumulation estimation in coastal and marine food webs. Reg. Stud. Mar. Sci. 2021;44:101797. [Google Scholar]
  57. Karki B. K. Philip L. Karki K. Ghimire A. Insight into urban river water quality using ecological risk assessment based on risk quotient. Water Conserv. Sci. Eng. 2024;9:56. doi: 10.1007/s41101-024-00289-1. [DOI] [Google Scholar]
  58. Zhu L. Zou L. Nie M. Xia B. Zhao J. Sui Q. Sun X. Xing B. Ocean acidification enhances TiO2 nanoparticle toxicity in Oryzias melastigma: Dominant role of size effects in driving bioaccumulation and hepatotoxicity. Water Res. 2025:124822. [Google Scholar]
  59. Burchio S. D. A. Natale S. Mulas M. Bevilacqua S. Provenza F. Anselmi S. Comis S. Renzi M. Falace A. Sunscreen-derived nano-TiO2 undermines viability and recruitment of the Mediterranean foundation alga Ericaria amentacea. Environ. Pollut. 2026:127687. doi: 10.1016/j.envpol.2026.127687. [DOI] [PubMed] [Google Scholar]
  60. Nugnes R. De Negri Atanasio G. Perata E. Lertora E. Dondero L. Robino F. Tardanico F. Capelli C. Ghioni F. Cai T. Comprehensive methodology for standardized ecotoxicological assessment of TiO2-based sunscreen leachates in aquatic environment. Front. Toxicol. 2025;7:1686954. doi: 10.3389/ftox.2025.1686954. [DOI] [PMC free article] [PubMed] [Google Scholar]
  61. Qian W. Chen C. C. Huang Y. Zhu X. Exposure concentration ratios and biological responses play a critical role in determining the joint toxicity of TiO2 nanoparticles and As (V) to the organism: The case study in marine algae Phaeodactylum tricornutum. Sci. Total Environ. 2024;909:168508. doi: 10.1016/j.scitotenv.2023.168508. [DOI] [PubMed] [Google Scholar]
  62. Zheng R. Li L. Wu Z. Xu A. Xu H. Hao Z. Yu S. Cai Y. Liu J. Distribution and source of titanium dioxide nanoparticles in seawater and sediment from Jiaozhou Bay, China. J. Hazard. Mater. 2025;483:136576. doi: 10.1016/j.jhazmat.2024.136576. [DOI] [PubMed] [Google Scholar]
  63. Keller A. A. Nanomaterials in sunscreens: potential human and ecological health implications. Int. J. Cosmet. Sci. 2023;45:127–140. doi: 10.1111/ics.12905. [DOI] [PubMed] [Google Scholar]

Associated Data

This section collects any data citations, data availability statements, or supplementary materials included in this article.

Supplementary Materials

RA-016-D6RA05133B-s001
RA-016-D6RA05133B-s001.pdf (847.2KB, pdf)

Data Availability Statement

The data supporting this article have been included as part of the supplementary information (SI). Supplementary information is available. See DOI: https://doi.org/10.1039/d6ra05133b.

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