Residues of non-phthalate plasticizers in seawater and sediments from Osaka Bay, Japan.

NPPs (Non-phthalate plasticizers) are used as alternative plasticizers to phthalate esters, but there is limited knowledge on environmental residues, and they have not been reported in Japan. A method to analyze NPPs in seawater using solid-phase extraction was developed, and the residual burden of Diisobutyl adipate (DIBA), Acetyl tributyl citrate (ATBC), Di-(2-ethylhexyl) adipate (DEHA), Di-(2-ethylhexyl) sebacate (DEHS) and Trioctyl trimellitate (TOTM) in seawater and sediment from the Osaka Bay was measured. Using an Oasis Max column and acetone as the eluting solvent, the recovery of the target substances in seawater is >68 %. In Osaka Bay, no NPPs were detected in seawater. On the other hand, ATBC and TOTM were detected in the sediment at 36-69 ng/g and 47-131 ng/g, respectively, from about half of the 14 sites, while DEHA and DEHS were detected at 83 ng/g and 181 ng/g, respectively, from only one site.

Anthropogenic Microparticles in Sea-Surface Microlayer in Osaka Bay, Japan.

The abundance, distribution, and composition of microparticles (MPs) in the sea-surface microlayer (S-SML, less than 100 µm of sea surface in this experiment) and in bulk water (1 m under the sea surface) were investigated to evaluate the pollution level of MPs in Osaka Bay in Japan. Both seawater fractions were collected at eight sites including ship navigation routes, the coastal area, and the center of Osaka Bay for 2021-2023. MPs were filtered for four size ranges (10-53, 53-125, 125-500, and >500 µm) and then digested with H2O2. MPs' abundance was microscopically assessed; and polymer types of MPs were identified by a Fourier transform infrared spectrometer (FTIR). For the 22 collections performed along eight sites, the average MPs' abundance was 903 ± 921 items/kg for S-SML, while for the 25 collections performed along the same sites, the average MPs' abundance was 55.9 ± 40.4 items/kg for bulk water, respectively. MPs in both S-SML and bulk water exhibited their highest abundance along the navigation routes. The smallest MPs (10-53 µm) accounted for 81.2% and for 62.2% of all MPs in S-SML and in bulk water among all sites, respectively. Polymethyl methacrylate (PMMA) was the major type of MPs identified while minor ones were polyethylene, polyesters, polystyrene, polypropylene, polyvinyl chloride, polyamide, etc. PMMA comprised 95.1% of total MPs in S-SML and 45.6% of total MPs in bulk water. In addition, PMMA accounted for 96.6% in S-SML and 49.5% in bulk water for the smallest MP category (10-53 µm). It can be assumed that the MP sources were marine paints-primarily APPs (antifouling paint particles)-as well as land coatings. Sea pollution due to microparticles from ship vessels should be given proper attention.

Impact of acetyl tributyl citrate on gonadal sex differentiation and expression of biomarker genes for endocrine disruption in Japanese medaka.

Plasticizers are broadly classified as phthalate or nonphthalate. Recently, acetyl tributyl citrate (ATBC), an environmentally friendly nonphthalate plasticizer, was revealed to have the ability to disrupt thyroid hormone activity in fish species. Therefore, we aimed to assess whether ATBC exhibits any sex hormone (i.e., androgenic or estrogenic) activities. First, we examined the effects of ATBC on gonadal sex differentiation. Subsequently, we analyzed the different expression of biomarker genes that respond to endocrine-disrupting chemicals (EDCs) with sexual hormone activity in the liver. We observed normal testes and ovaries after both XX and XY medakas were exposed to ATBC, indicating that ATBC is not an EDCs with strong sex hormone activity and that it does not induce intersex (testis-to-ova or ovo-to-testis) or sex changes in Japanese medaka. The vitellogenin 1 (vtg1) and vitellogenin 2 (vtg2) mRNA expression levels in the liver of XX medakas were significantly reduced compared with those in the control group, whereas the expression levels of these genes in the liver of XY medakas remained unchanged. Finally, we examined the changes in the expression of biomarker genes that respond to EDCs with sex hormone activity in the gonads. The expression levels of biomarker genes did not differ significantly from that of the control group, although the expression levels of gsdf mRNA tended to increase while that of aromatase mRNA tended to decrease in the ovary of XX medakas following ATBC exposure. Conversely, the expression levels of gsdf and aromatase mRNAs in the testis of XY medakas remained unchanged. These results suggest that ATBC does not exhibit estrogenic activity, although it may have weak androgenic activity or no sexual hormone activity.

Effects of plasticizer diisobutyl adipate on the Japanese medaka (Oryzias latipes) endocrine system.

Plasticizer pollution of the water environment is one of the world's most serious environmental issues. Phthalate plasticizers can disrupt endocrine function in vertebrates. Therefore, this study analyzed thyroid-related, reproduction-related, and estrogen-responsive genes in Japanese medaka (Oryzias latipes) to determine whether non-phthalate diisobutyl adipate (DIBA) plasticizer could affect endocrine hormone activity or not. Developmental toxicity during fish embryogenesis was also evaluated. At a concentration of 11.57 mg/l, embryonic exposure to DIBA increased the mortality rate. Although abnormal development, including body curvature, edema, and lack of swim bladder inflation, was observed at 3.54 and 11.57 mg/l DIBA, growth inhibition and reduced swimming performance were also observed. In addition, DIBA exposure increased the levels of thyroid-stimulating hormone beta-subunit (tshß) and deiodinase 1 (dio1) but decreased the levels of thyroid hormone receptor alpha (trα) and beta (trß). These results suggest that DIBA has thyroid hormone-disrupting activities in fish. However, kisspeptin (kiss1 and kiss2), gonadotropin-releasing hormone (gnrh1), follicle-stimulating hormone beta (fshß), luteinizing hormone beta (lhß), choriogenin H (chgH), and vitellogenin (vtg1) expression did not change dose-dependently in response to DIBA exposure, whereas gnrh2 and vtg2 expression was elevated. These results indicate that DIBA has low estrogenic activity and does not disrupt the endocrine reproduction system in fish. Overall, this is the first report indicating that non-phthalate DIBA plasticizer is embryotoxic and disrupt thyroid hormone activity in fish.

Effects of non-phthalate plasticizer bis(2-ethylhexyl) sebacate (DEHS) on the endocrine system in Japanese medaka (Oryzias latipes).

Water pollution due to plasticizers is one of the most severe environmental problems worldwide. Phthalate plasticizers can act as endocrine disruptors in vertebrates. In this study, we investigated whether the non-phthalate bis(2-ethylhexyl) sebacate (DEHS) plasticizer can act as an endocrine disruptor by evaluating changes in the expression levels of thyroid hormone-related, reproduction-related, and estrogen-responsive genes of Japanese medaka (Oryzias latipes) exposed to the plasticizer. Following the exposure, the gene expression levels of thyroid-stimulating hormone subunit beta (tshß), deiodinase 1 (dio1), and thyroid hormone receptor alpha (trα) did not change. Meanwhile, DEHS suppressed dio2 expression, did not induce swim bladder inflation, and eventually reduced the swimming performance of Japanese medaka. These findings indicate that DEHS can potentially disrupt the thyroid hormone-related gene expression and metabolism of these fish. However, exposure to DEHS did not induce changes in the gene expression levels of kisspeptin 1 (kiss1), gonadotropin-releasing hormone (gnrh), follicle-stimulating hormone beta (fshß), luteinizing hormone beta (lhß), choriogenin H (chgH), and vitellogenin (vtg) in a dose-dependent manner. This is the first report providing evidence that DEHS can disrupt thyroid hormone-related metabolism in fish.

Adverse effects of thyroid-hormone-disrupting chemicals 6-propyl-2-thiouracil and tetrabromobisphenol A on Japanese medaka (Oryzias latipes).

Thyroid-hormone-disrupting chemicals are increasingly attracting attention because of their potential harmful effects on animal health, including on fishes. Here, we investigated the effects of exposure to the thyroid-hormone-disrupting chemicals 6-propyl-2-thiouracil (PTU) and tetrabromobisphenol A (TBBPA) on swim bladder inflation, eye development, growth, swimming performance, and the expression of thyroid-related genes in Japanese medaka (Oryzias latipes). PTU exposure resulted in reductions in eye size, growth, and swim bladder inflation, and these effects led to poorer swimming performance. These phenotypic effects were accompanied by increased expression of the thyroid-stimulating hormone subunit beta (tshß) paralog tshß-like, but there were no significant changes in expression for tshß, deiodinase 1 (dio1), deiodinase 2 (dio2), and thyroid hormone receptor alpha (trα) and beta (trß). For PTU exposure, we identified the key event (swim bladder inflation reduction) and an adverse outcome (swimming performance reduction). No significant effects from TBBPA exposure were seen on swim bladder inflation, eye development, growth, or swimming performance. However, expression of tshß-like and tshß (significantly enhanced) and trα and trß (significantly reduced) were affected by TBBPA exposure albeit not in dose-dependent manners. There were no effects of TBBPA on the expression of dio1 and dio2. We thus show that the two thyroid-hormone-disrupting chemicals PTU and TBBPA differ in their effect profiles with comparable effects on the studied phenotypes and thyroid-related gene expression to those reported in zebrafish.

Bisphenol A alters sexual dimorphism and gene expression in marine medaka Oryzias melastigma.

Bisphenol A (BPA) is an endocrine disruptor that is present in freshwater and marine environments. However, conclusive evidence for the toxicity of chronic BPA exposure to marine fishes remains lacking. Therefore, we investigated the influence of BPA on male marine medaka (Oryzias melastigma). BPA exposure induced formation of testis-ova at 2610 µg/L, and male-type anal fins became more female type in a concentration-dependent manner. Some males with female-type anal fins had normal testes, indicating that anal fin shape is more sensitive to BPA. Gonadal soma-derived factor (gsdf) expression decreased after BPA exposure in the 746 and 2610 µg/L exposure groups, although the changes were not statistically significant. Additionally, liver vitellogenin (vtg) expression increased in a dose-dependent manner and was significantly higher in all exposure groups. vtg and gsdf are likely to be useful biomarkers for the impact of estrogenic endocrine disrupters in O. melastigma.

Thyroid hormone disruption by bis-(2-ethylhexyl) phthalate (DEHP) and bis-(2-ethylhexyl) adipate (DEHA) in Japanese medaka Oryzias latipes.

Pollution of water bodies with plasticizers is a serious environmental problem worldwide. In this study, we investigated the effects of plasticizers bis-(2-ethylhexyl) phthalate (DEHP) and bis-(2-ethylhexyl) adipate (DEHA) in Japanese medaka (Oryzias latipes). DEHP significantly increased the expression of all the genes tested: thyroid stimulating hormone beta subunit (tshß-like), tshß, deiodinase 1 (dio1), deiodinase 2 (dio2), and thyroid hormone receptor alpha (trα) and beta (trß). However, DEHA only significantly increased tshß at 7.4 µg/L but significantly decreased dio2 expression at 25.8, 111.1, and 412.6 4 µg/L, while other genes were not significantly affected. Both chemicals reduced eye size and total body length, but did not affect embryo development, hatching time and rate, and swimming performance. DEHA alone affected swim bladder inflation and not DEHP. This is the first report that not only DEHP but also DEHA disrupt thyroid hormone activity in fish. DEHP contamination (13.2 µg/L) was detected in tap water from Kobe, Japan; thus, tap water itself may disrupt thyroid hormone activity in Japanese medaka. Importantly, the effective concentration of DEHP for thyroid hormone-related gene expression and growth was close to or lower than DEHP concentrations reported in surface water elsewhere, indicating that DEHP contamination is a serious aquatic pollution.

Color preferences and gastrointestinal-tract retention times of microplastics by freshwater and marine fishes.

We examined ingestion and retention rates of microplastics (MPs) by two freshwater (Japanese medaka and zebrafish) and two marine fish species (Indian medaka and clown anemonefish) to determine their color preferences and gastrointestinal-tract retention times. In our ingestion experiments, clown anemonefish ingested the most MP particles, followed by zebrafish, and then Japanese and Indian medaka. Next, we investigated color preferences among five MP colors. Red, yellow, and green MP were ingested at higher rates than gray and blue MPs for all tested fish species. To test whether these differences truly reflect a recognition of and preference for certain colors based on color vision, we investigated the preferences of clown anemonefish for MP colors under light and dark conditions. Under dark conditions, ingestion of MP particles was reduced, and color preferences were not observed. Finally, we assessed gastrointestinal-tract retention times for all four fish species. Some individuals retained MP particles in their gastrointestinal tracts for over 24 h after ingestion. Our results show that fish rely on color vision to recognize and express preferences for certain MP colors. In addition, MP excretion times varied widely among individuals. Our results provide new insights into accidental MP ingestion by fishes.

Effect of thyroid hormone-disrupting chemicals on swim bladder inflation and thyroid hormone-related gene expression in Japanese medaka and zebrafish.

We compared the influence of thyroid hormone-disrupting chemicals (heptafluorobutanoic acid, PFBA and tris[1,3-dichloro-2-propyl] phosphate, TDCPP) and thyroid hormone (3,3',5-triiodo-L-thyronine, T3) on swim bladder inflation and thyroid hormone-related gene expression in Japanese medaka and zebrafish. The swim bladder of most larvae had inflated at 4 h post hatching (hph) in Japanese medaka and at 48 hph in zebrafish in controls. In both fish species, the swim bladder inflation was inhibited in larvae exposed to PFBA (lowest observed effect concentration [LOEC] in medaka: 40 mg/L; in zebrafish: 80 mg/L), TDCPP (LOEC in medaka: 1 mg/L; in zebrafish: 0.5 mg/L), and T3 (no inhibition in Japanese medaka; LOEC in zebrafish: 7.5 µg/L). We also examined the influence of PFBA, TDCPP, and T3 on the expression of thyroid stimulating hormone subunit beta (tshß) or thyroid hormone receptor alpha (trα) and beta (trß). No changes were observed in the expression of genes after PFBA and TDCPP exposure; however, T3 exposure upregulated trα and trß expression in both fish species. When the results were compared between Japanese medaka and zebrafish, swim bladder inflation in both species was found to be inhibited by exposure to thyroid hormone-disrupting chemicals. Our results show that inhibition of the swim bladder inflation at 4 hph in Japanese medaka and 48 hph in zebrafish is a potential indicator of thyroid hormone-disturbing activity of chemicals.

Floating particles with high copper concentration in the sea-surface microlayer.

This study sought to clarify whether suspended particles containing high Cu concentrations are present in the sea-surface microlayer (S-SML). For this reason, suspended particles (10-2000 µm) in the S-SML were collected periodically from a ship mooring pond during 2018-2020, and the acid-soluble Cu concentration in the suspended particles was measured as particulate Cu (P-Cu). The highest concentration of P-Cu in the S-SML of the pond was 75 µg L-1 with a 90th percentile value of 2.5 µg L-1. This is below P-Cu values reported for the S-SML in North American ports, but 140 times higher than this found in bulk seawater in the Atlantic Ocean. The highest P-Cu concentration in the S-SML of non-organism (abiotic) origin was 17 µg L-1, and the abiotic P-Cu to P-Cu ratio varied from 0.2 to 100%, likely depending on the quality and quantity of biogenic material in the S-SML samples. It is assumed that the S-SML particles examined here contain high Cu concentrations originating from ship antifouling paints.

Invasive Weed Asystasia gangetica as a Potential Biomonitor and a Phytoremediator of Potentially Toxic Metals: A Case Study in Peninsular Malaysia.

The invasive weed Asystasia gangetica was investigated for its potential as a biomonitor and as a phytoremediator of potentially toxic metals (PTMs) (Cd, Cu, Ni, Pb, and Zn) in Peninsular Malaysia owing to its ecological resistance towards unfavourable environments. The biomonitoring potential of PTMs was determined based on the correlation analysis of the metals in the different parts of the plant (leaves, stems, and roots) and its habitat topsoils. In the roots, the concentrations (mg/kg dry weight) of Cd, Cu, Ni, Pb, and Zn ranged from 0.03 to 2.18, 9.22 to 139, 0.63 to 5.47, 2.43 to 10.5, and 50.7 to 300, respectively. In the leaves, the concentrations (mg/kg dry weight) of Cd, Cu, Ni, Pb, and Zn ranged from 0.03 to 1.16, 7.94 to 20.2, 0.03 to 6.13, 2.10 to 21.8, and 18.8 to 160, respectively. In the stems, the concentrations (mg/kg dry weight) of Cd, Cu, Ni, Pb, and Zn ranged from 0.03 to 1.25, 5.57 to 11.8, 0.23 to 3.69, 0.01 to 7.79, and 26.4 to 246, respectively. On the other hand, the phytoremediation potential of the five metals was estimated based on the bioconcentration factor (BCF) and the translocation factor (TF) values. Correlation analysis revealed that the roots and stems could be used as biomonitors of Cu, the stems as biomonitors of Ni, the roots and leaves as biomonitors of Pb, and all three parts of the plant as biomonitors of Zn. According to the BCF values, in the topsoil, the "easily, freely, leachable, or exchangeable" geochemical fractions of the five metals could be more easily transferred to the roots, leaves, and stems when compared with total concentrations. Based on the TF values of Cd, Ni, and Pb, the metal transfer to the stems (or leaves) from the roots was efficient (>1.0) at most sampling sites. The results of BCF and TF showed that A. gangetica was a good phytoextractor for Cd and Ni, and a good phytostabilizer for Cu, Pb, and Zn. Therefore, A. gangetica is a good candidate as a biomonitor and a phytoremediator of Ni, Pb, and Zn for sustainable contaminant remediation subject to suitable field management strategies.

Bioaccumulation of zinc in edible tropical vegetables in Peninsular Malaysia and its human health risk assessment based on various ethnicities in Malaysia.

Along with the growing utilization of zinc (Zn) and Zn-containing nanoparticles in various industries, Zn ecotoxicological evaluation on human food supply is necessary even though Zn is generally considered safe and rarely concentrated ecotoxicologically. This study aimed to investigate the bioaccumulation of Zn in 18 species of vegetables (seven leafy, nine fruity vegetables and one species each of tuber and legume) collected from two farming sites in the west coast of Peninsular Malaysia. A human health risk assessment (HHRA) was also conducted. In addition to HHRA based on the general population, HHRA based on each major ethnic group of the Malaysian society was also determined considering that the food consumption pattern would definitely be varied across ethnicities and age groups (children and adults). The study results showed that Zn concentrations were significantly higher (p < 0.05) in leafy vegetables than in other types of vegetables. However, the target hazard quotient (THQ) values were all found to be < 1.0. Therefore, based on the Malaysian ethnicities and age groups with their respective vegetable consumption patterns, the results indicated insignificant noncarcinogenic human health risk of Zn via oral consumption of vegetables by the Malaysian population. As a metric of measurement of HHRA, a comparison of THQ values could yield previously unreported insights into HHRA differences among the compared populations. A comparison of THQ values among the consumer groups indicated higher HHR for Chinese Malaysians and children due to their higher vegetable consumption and lower body weight, respectively. A comparison the Zn intakes of all the consumer groups with the recommended nutrient intakes indicated that the oral consumption of the vegetable species collected in this study would not result in Zn-related hazards and would not be able to fulfil the Zn dietary need of the individual consumer.

Assessments of the Ecological and Health Risks of Potentially Toxic Metals in the Topsoils of Different Land Uses: A Case Study in Peninsular Malaysia.

Human activities due to different land uses are being studied widely in many countries. This study aimed to determine the ecological risks and human health risk assessments (HHRA) of Cd, Pb, Ni, Cu, and Zn in the topsoils of six land uses in Peninsular Malaysia. The ranges of the potentially toxic metals (PTMs) in the soils (mg/kg, dry weight) of this study were 0.24-12.43 for Cd (mean: 1.94), 4.66-2363 for Cu (mean: 228), 2576-116,344 for Fe (mean: 32,618), 2.38-75.67 for Ni (mean: 16.04), 7.22-969 for Pb (mean: 115) and 11.03-3820 for Zn (mean: 512). For the ecological risk assessments, the potential ecological risk index (PERI) for single metals indicated that the severity of pollution of the five metals decreased in the following sequence: Cd > Cu > Pb > Zn > Ni. It was found that industry, landfill, rubbish heap, and mining areas were categorized as "very high ecological risk". For HHRA, the land uses of industry, landfill and rubbish heap were found to have higher hazard quotient (HQ) values for the three pathways (with the order: ingestion > dermal contact > inhalation ingestion) of the five metals for children and adults, when compared to the mining, plantation, and residential areas. The values for both the non-carcinogenic (Cd, Cu, Ni, and Zn), and carcinogenic risks (CR) for inhalation (Cd and Ni) obtained for children and adults in this study showed no serious adverse health impacts on their health. However, of public concern, the hazard index (HI), for Pb of children at the landfill (L-3) and the rubbish heap (RH-3) sites exceeded 1.0, indicating non-carcinogenic risk (NCR) for children. Therefore, these PERI and HHRA results provided fundamental data for PTMs pollution mitigation and environmental management in areas of different land uses in Peninsular Malaysia.

Human Health Risk Assessments of Trace Metals on the Clam Corbicula javanica in a Tropical River in Peninsular Malaysia.

This study aimed to analyse ten trace metal concentrations in the edible part of the freshwater clam Corbicula javanica and to provide a critical assessment of the potential risks to human health through consumption of this clam as food based on well-established indices and food safety guidelines. The clams were captured from a pristine original site and transplanted to other sites with different environmental qualities. The trace metal levels in the edible total soft tissue (TST) of the clam were below those of the food safety guidelines referred to except for Pb, which exceeded the permissible limit set by the European Commission (2006) and the US Food and Drug Administration/ Center for Food Safety and Applied Nutrition); Interstate Shellfish Sanitation Conference. (USFDA/CFSAN; ISSC) (2007). The estimated daily intake (EDI) values of the clam were found to be lower than the oral reference dose and the calculated target hazard quotient (THQ) and total THQ were found to be less than 1. Therefore, in conclusion, the human health risk for consumption of TST of C. javanica at both average and high-level were insignificant regardless of the environment it was exposed to.

Early developmental responses of three sea urchin species to tralopyril and its two degradation products.

Tralopyril (TLP) is a newly emerged antifouling biocide which rapidly degrades in water. The scientific data on its possible adverse effects to biota is very limited, and even more limited is the ecotoxicity data of its degradation products (DPs). In the present study we investigated the toxicity of TLP and its two main DPs on fertilization and embryogenesis of three sea urchin species: Clypeaster japonicus, Pseudocentrotus depressus and Hemicentrotus pulcherrimus. The species sensitivity to chemicals was investigated and compared. Additionally, the stability of TLP in test medium was examined. TLP in test medium degraded into one single degradation product. The degradation was slower at 17 °C (average incubation temperature for winter species H. pulcherrimus) than at 20 °C (incubation temperature for the other two species). Both DPs at 100 µg/L did not appear toxic, however, TLP highly affected larval development of all tested species. Sensitivity was similar for C. japonicus and P. depressus, but higher toxicity was noticed for H. pulcherrimus. The increased TLP toxicity on exposed H. pulcherrimus embryos could be attributed to the higher TLP stability at lower incubation temperature. Results suggest higher vulnerability to TLP for species spawning in colder seasons. Fertilization test appeared to be less sensitive than the embryotoxicity test.