Mercury is a globally distributed toxicant to aquatic animals and mammals. However, the potential risks of environmental relevant mercury in terrestrial systems remain largely unclear. The metabolic profiles of the earthworm Eisenia fetida after exposure to soil contaminated with mercury at 0.77 ± 0.09 mg/kg for 2 weeks were investigated using a two-dimensional nuclear magnetic resonance-based (¹H-¹³C NMR) metabolomics approach. The results revealed that traditional endpoints (e.g., mortality and weight loss) did not differ significantly after exposure. Although histological examination showed sub-lethal toxicity in the intestine as a result of soil ingestion, the underlying mechanisms were unclear. Metabolite profiles revealed significant decreases in glutamine and 2-hexyl-5-ethyl-3-furansulfonate in the exposed group and remarkable increases in glycine, alanine, glutamate, scyllo-inositol, t-methylhistidine and myo-inositol. More importantly, metabolic network analysis revealed that low mercury in the soil disrupted osmoregulation, amino acid and energy metabolisms in earthworms. A metabolic net link and schematic diagram of mercury-induced responses were proposed to predict earthworm responses after exposure to mercury at environmental relevant concentrations. These results improved the current understanding of the potential toxicity of low mercury in terrestrial systems.

Mercury (Hg) is recognized as a dangerous contaminant due to its bioaccumulation and biomagnification within trophic levels, leading to serious health risks to aquatic biota. Therefore, there is an urgent need to unravel the mechanisms underlying the toxicity of Hg. To this aim, a metabolomics approach based on protonic nuclear magnetic resonance (1H NMR), coupled with chemometrics, was performed on the gills of wild golden grey mullets L. aurata living in an Hg-polluted area in Ria de Aveiro (Portugal). Gills were selected as target organ due to their direct and continuous interaction with the surrounding environment. As a consequence of accumulated inorganic Hg and methylmercury, severe changes in the gill metabolome were observed, indicating a compromised health status of mullets. Numerous metabolites, i.e. amino acids, osmolytes, carbohydrates, and nucleotides, were identified as potential biomarkers of Hg toxicity in fish gills. Specifically, decrease of taurine and glycerophosphocholine, along with increased creatine level, suggested Hg interference with the ion-osmoregulatory processes. The rise of lactate indicated anaerobic metabolism enhancement. Moreover, the increased levels of amino acids suggested the occurrence of protein catabolism, further supported by the augmented alanine, involved in nitrogenous waste excretion. Increased level of isobutyrate, a marker of anoxia, was suggestive of onset of hypoxic stress at the Hg contaminated site. Moreover, the concomitant reduction in glycerophosphocholine and phosphocholine reflected the occurrence of membrane repair processes. Finally, perturbation in antioxidant defence system was revealed by the depletion in glutathione and its constituent amino acids. All these data were also compared to the differential Hg-induced metabolic responses previously observed in liver of the same mullets (Brandão et al., 2015). Overall, the environmental metabolomics approach demonstrated its effectiveness in the evaluation of Hg toxicity mechanisms in wild fish under realistic environmental conditions, uncovering tissue-specificities regarding Hg toxic effects namely in gills and liver.

The efficiency of biopurification systems to treat pesticide-contaminated water was previously studied in microcosms. To validate the obtained results, macrocosm systems were set-up. Four pesticides (linuron, isoproturon, bentazone, and metalaxyl) were continuously applied to ten different organic substrate mixes. Retention of the pesticides was similar and in some cases slightly lower in the macrocosms compared to the microcosms. Differences in retention between the different mixes were however minimal. Moreover, the classification of the retention strength of the pesticides was identical to that observed in microcosms: linuron > isoproturon > metalaxyl > bentazone. Monod kinetics were used to describe delayed degradation, which occurred for isoproturon, metalaxyl and bentazone. No breakthrough of linuron was observed, thus, this pesticide was appointed as the most retained and/or degraded pesticide, followed by isoproturon, metalaxyl and bentazone. Finally, most of the matrix mixes efficient in degrading or retaining pesticides were mixes containing dried cow manure. Transport of pesticides in macrocosm containing organic substrates.

Arsenic (As) is a metalloid element that is ubiquitous in the marine environment and its contamination has received worldwide attention due to its potential toxicity. Arsenic can induce multiple adverse effects, such as lipid metabolism disorder, immune system dysfunction, oxidative stress and carcinogenesis, in animals. Inorganic arsenic includes two chemical forms, arsenite (As (III)) and arsenate (As (V)), in natural environment. As (V) is the dominant form in natural waters. In the present study, metabolomic and proteomic alterations were investigated in juvenile rockfish Sebastes schlegelii exposed to environmentally relevant concentrations of As (V) for 14 d. The analysis of iTRAQ-based proteomics combined with untargeted NMR-based metabolomics indicated apparent toxicological effects induced by As (V) in juvenile rockfish. In details, the metabolites, including lactate, alanine, ATP, inosine and phosphocholine were significantly altered in As-treated groups. Proteomic responses suggested that As (V) could not only affected energy and primary metabolisms and signal transduction, but also influenced cytoskeleton structure in juvenile rockfish. This work suggested that the combined proteomic and metabolomic approach could shed light on the toxicological effects of pollutants in rockfish S. schlegelii.

¹H NMR-based metabolomics was used to examine the response of the earthworm Eisenia fetida after exposure to sub-lethal concentrations of phenanthrene over time. Earthworms were exposed to 0.025 mg/cm² of phenanthrene (1/64th of the LC₅₀) via contact tests over four days. Earthworm tissues were extracted using a mixture of chloroform, methanol and water, resulting in polar and non-polar fractions that were analyzed by ¹H NMR after one, two, three and four days. NMR-based metabolomic analyses revealed heightened E. fetida responses with longer phenanthrene exposure times. Amino acids alanine and glutamate, the sugar maltose, the lipids cholesterol and phosphatidylcholine emerged as potential indicators of phenanthrene exposure. The conversion of succinate to fumarate in the Krebs cycle was also interrupted by phenanthrene. Therefore, this study shows that NMR-based metabolomics is a powerful tool for elucidating time-dependent relationships in addition to the mode of toxicity of phenanthrene in earthworm exposure studies.

It is highly likely that the toxicity of water accommodated fractions (WAF) will influence marine microalgae, and consequently lead to potential risk for the marine ecological environment. However, it was often neglected whether WAF can influence the transformation of relative compounds in organisms. The metabolism of amino acids (AAs) can be used to track physiological changes in microalgae because amino acids are the basis of proteins and enzymes. In this study, using marine Chlorophyta Platymonas helgolandica as the test organism, the effects of different concentrations of WAF on AA compositions and stable carbon isotope ratios (δ¹³C) of individual AAs of Platymonas helgolandica were investigated. The results showed that the WAF of #180 fuel oil had an obvious suppressing effect on the growth and chlorophyll a content of microalgae. The growth inhibitory rate at 96 h was 80.66% at a WAF concentration of 0.50 mg L⁻¹ compared with the control. Furthermore, seven among the 16 AAs, including alanine, cysteine, proline, aspartic acid, lysine, histidine and tyrosine, had relatively high abundance. Under the glycolysis pathway, the cysteine abundance was higher than control, meaning that the biosynthesized pathway of alanine through cysteine as a precursor could be damaged. Phosphoenolpyruvate (PEP) was an important synthesis precursor of alanine (leucine) and aromatic AA family (Phenylalanine and tyrosine), and played an important role in δ¹³CAAₛ fractionation under the WAF stress. Under the TCA pathway, to protect cell metabolism activities under WAF stress, the δ¹³C value of threonine and proline abundance in microalgae with the increase in WAF stress. Therefore, δ¹³CAAₛ fractionation can be used as a novel method for toxicity evaluation of WAF on future.

Nitrated polycyclic aromatic hydrocarbons (nitro-PAHs) have been widely studied for their mutagenic and carcinogenic effects. This study aims to investigate whether exposure to nitro-PAHs is associated with biomarkers of carbohydrate metabolism, an underlying risk factor for metabolic disorder. Early morning urine and blood samples were longitudinally collected two times with a four-week interval from 43 healthy adults. Five urinary amino-PAHs (1-aminonaphthalene, 2-aminonaphthalene, 9-aminophenanthrene, 2-aminofluorene, and 1-aminopyrene) were measured as biomarkers of nitro-PAH exposures. We measured plasma concentrations of glucose and six amino acids that can regulate insulin secretion, including aspartate (Asp), glutamate (Glu), glutamine (Gln), alanine (Ala), Arginine (Arg), and ornithine (Orn). We found that increasing concentrations of 9-aminophenanthrene were significantly associated with increasing glucose levels and with decreasing Asp, Glu, Ala, and Orn levels. We estimated that 26.4 %–43.8 % of the 9-aminophenanthrene-associated increase in glucose level was mediated by Asp, Glu, and Orn. These results suggest that exposure to certain nitro-PAHs affects glucose homeostasis, partly resulting from the depletion of insulin-stimulating amino acids (Asp, Glu, and Orn).

The accumulation of cadmium (Cd) in rice grains is closely associated with the content of mineral nutrients and amino acid metabolism, but the causal link among them is unclear. Profiles of amino acids (AAs) and quantities of essential nutrients in grains from early and late rice cultivars grown at four sites with different Cd levels were analyzed in the present study. Hazard quotients (HQs) for consumers by intake of rice from late cultivars were much higher than that from early cultivars at sites with soil Cd content of 0.25, 0.61 and 0.84 mg kg⁻¹. Cadmium accumulation in grains resulted in a sharp reduction of total essential AAs and non-essential AAs in both early and late rice cultivars. High-Cd-accumulating (HCA) cultivars had significantly higher level of glutamate (Glu) than low-Cd-accumulating (LCA) cultivars when rice Cd content was less than 0.20 mg kg⁻¹. However, Glu level in grains dramatically declined with the accumulation of Cd, which subsequently leaded to the reduction of other AAs. Cadmium content was well predicted by five amino acids (i.e., Glu, Alanine, Phenylalanine, Glycine and Threonine) or four essential elements (Ca, Fe, Mn and Zn) when rice Cd was less than 0.80 mg kg⁻¹. Amino acids played more important roles than nutrients in Cd accumulation. When Cd content was in the range of 0.40–1.16 mg kg⁻¹, the Mn content in rice increased significantly with the increase of Cd content, while the Glu content dropped down synchronously. Remarkably, the ratio between Mn and Glu displayed the highest direct path coefficient on Cd accumulation than any single cation or amino acid. These results indicate that high capacity in synthesizing Glu and concentrating Mn is the determinant factor for Cd accumulation in rice grains, and abundant Glu in aleurone layer may alleviate Cd toxicity by forming Glu-Cd complex.

Free amino acid concentrations and nitrogen (N) isotopic composition in new current-year (new), mature current-year (middle-aged) and previous-year (old) Masson pine (Pinus massoniana Lamb.) needles were determined to indicate atmospheric N deposition in Guiyang (SW China). In different areas, free amino acids (especially arginine) concentrations in new and middle-aged needles were higher than in old needles, and the variation of free amino acids (especially arginine) concentrations in new and middle-aged needles was also greater than in old needles. This indicate that free amino acids in new and middle-aged needles may be more sensitive to N deposition compared to old needles. Moreover, concentrations of total free amino acids, arginine, histidine, γ-aminobutyric acid and alanine in middle-aged needles exhibited a strong relationship with N deposition (P < 0.05). Needle δ¹⁵N values showed a strong gradient from central Guiyang to the rural area, with more positive δ¹⁵N (especially in old needles) in the city center (0–5 km) and more negative δ¹⁵N (especially in old needles) in rural area (30–35 km). These suggest that N deposition in the urban center may be dominated by ¹⁵N-enriched NOx-N from traffic exhausts, while it is dominated by isotopically light atmospheric NHx-N from agriculture in rural area. Soil δ¹⁵N decreased slightly with distance from the city center, and the difference in δ¹⁵N values between the soil and needles (especially for old needles) increased significantly with the distance gradient, indicating that atmospheric N deposition may be an important N source for needles. This study provides novel evidence that free amino acids in needles and age-dependent needle δ¹⁵N values are useful indicators of atmospheric N deposition.

1H NMR metabolomics was used to monitor earthworm responses to sub-lethal (50-1500 mg/kg) phenanthrene exposure in soil. Total phenanthrene was analyzed via soxhlet extraction, bioavailable phenanthrene was estimated by hydroxypropyl-β-cyclodextrin (HPCD) and 1-butanol extractions and sorption to soil was assessed by batch equilibration. Bioavailable phenanthrene (HPCD-extracted) comprised ∼65-97% of total phenanthrene added to the soil. Principal component analysis (PCA) showed differences in responses between exposed earthworms and controls after 48 h exposure. The metabolites that varied with exposure included amino acids (isoleucine, alanine and glutamine) and maltose. PLS models indicated that earthworm response is positively correlated to both total phenanthrene concentration and bioavailable (HPCD-extracted) phenanthrene in a freshly spiked, unaged soil. These results show that metabolomics is a powerful, direct technique that may be used to monitor contaminant bioavailability and toxicity of sub-lethal concentrations of contaminants in the environment. These initial findings warrant further metabolomic studies with aged contaminated soils.