Coelomic fluid: a complimentary biological medium to assess sub-lethal endosulfan exposure using 1H NMR-based earthworm metabolomics

Abstract

Endosulfan is an environmentally persistent pesticide and has been shown to be genotoxic, neurotoxic and carcinogenic to surrounding organisms. Earthworms are widely used in environmental metabolomic studies to assess soil ecotoxicity. Previous nuclear magnetic resonance (NMR)-based metabolomic studies have analyzed earthworm tissue extracts after exposure to endosulfan and identified some key metabolic indicators that can be used as biomarkers of stress. However, some metabolites may have been masked due to overlap with other metabolites in the tissue extract. Therefore, in this study, the coelomic fluid (CF) and the tissue extract of the earthworm, Eisenia fetida, were both investigated using ¹H NMR-based metabolomics to analyze their metabolic profile in response to endosulfan exposure at three sub-lethal (below LC₅₀) concentrations. Principal component analysis determined the earthworm CF and earthworm tissue extract to both have significant separation between the exposed and control at the two highest sub-lethal endosulfan exposures (1.0 and 2.0 μg cm⁻²). Alanine, glycine, malate, α-ketoglutarate, succinate, betaine, myo-inositol, lactate and spermidine in the earthworm CF and alanine, glutamine, fumarate, glutamate, maltose, melibiose, ATP and lactate in earthworm tissue extract were all detected as having significant fluctuations after endosulfan exposure. An increase in ATP production was detected by the increase activity in the citric acid cycle and by anaerobic metabolism. A significant decrease in the polyamine, spermidine after endosulfan exposure describes an apoptotic mode of protection which correlates to a previous endosulfan exposure study where DNA damage has been reported. This study highlights that earthworm CF is a complementary biological medium to tissue extracts and can be helpful to better understand the toxic mode of action of contaminants at sub-lethal levels in the environment.

In‐Phase Ultra High‐Resolution In Vivo NMR

“…Communications nearly identical to that of the aqueous buffer extract. [24] It also confirms that the real line-shape,p eak intensities,a nd the general spectra profile are well represented in the IP-iSQC. (a perfect correlation should not be expected, as as eparate worm was used to prepare the buffer extract.…”

“…[24] It also confirms that the real line-shape,p eak intensities,a nd the general spectra profile are well represented in the IP-iSQC. [24] It also confirms that the real line-shape,p eak intensities,a nd the general spectra profile are well represented in the IP-iSQC.…”

1H NMR Metabolic Profiling of Earthworm (Eisenia fetida) Coelomic Fluid, Coelomocytes, and Tissue: Identification of a New Metabolite—Malylglutamate

“…CF was extruded from the earthworms similarly to previously described methods. 7 , 12 Earthworms were dipped in EMD Millipore Simplicity ultrapure water to remove soil and patted dry. The earthworm was transferred to a 35 × 10 mm Falcon Petri dish containing 500 μL of 0.1% NaCl in ultrapure water.…”

“…The metabolomic impact of endosulfan on the CF and whole-tissue extract of E. fetida was measured via the Organization for Economic Co-operation and Development (OECD) filter paper test. 12 Decreases in α-ketoglutarate, malate, spermidine, and succinate were observed in the tissue and CF of exposed worms, while alanine, ATP, betaine, lactate, and myo -inositol levels increased. In the whole-worm extract, fumarate also decreased in comparison with the control.…”

Whole Transcriptome Analysis Using Next-Generation Sequencing of Sterile-Cultured Eisenia andrei for Immune System Research

“…Two earthworm species, Eisenia fetida and Eisenia andrei , have been used in European pesticide marketing authorization experiments since the early 1980s [ 6 ]. In these studies, several bioactive proteins were identified in the coelomic fluid of earthworms [ 7 – 9 ]. These proteins exhibit a variety of antibacterial, hemolytic, cytotoxic, hemagglutinating, and proteolytic activities [ 10 ].…”