Since soil water content influences the springtails’ body water content (Supplementary Fig. S2) we had to estimate the internal concentration of phenanthrene on a dry weight basis as described by Wang et al. (2023 (link)) using the linear relationship between exposure time and the water content of springtails exposed to noncontaminated soil (Supplementary Fig. S2) to transform the fresh weight of phenanthrene-exposed springtails to dry weight and calculate an internal concentration of phenanthrene on a dry weight basis.
Phenanthrene concentrations in animal tissues were measured according to the method described by Holmstrup et al. (Holmstrup et al., 2014 ). In brief, the adults from each replicate were transferred to a 1.5 mL brown glass vial, and 500 µL of acetonitrile (VMR international, USA) was added. The vials were placed in a sonicator (Thermo, Germany), sonicated for 90 min on ice, kept at room temperature for 24 h, frozen at -18 °C for 24 h and kept at room temperature for another 24 h. The samples were sonicated again for 90 min on ice and then transferred to 1.5 mL tubes for brief centrifugation (3 min at 2,400 g). The supernatant from each tube was transferred to an autosampler vial and stored at -80 °C until phenanthrene analysis by GC‒MS (GCMS-QP2010, Shimadzu, Japan). Phenanthrene standards, including blanks, were run in parallel and subjected to the same extraction procedure.
Phenanthrene in soil samples (1 g fresh weight) was extracted with 4 mL of acetonitrile by shaking at 200 rpm for 24 h, followed by centrifugation at 1000×g for 5 min. The supernatants were transferred to autosampler vials and analysed as described above. For quality control, blank medium and uncontaminated soil were analyzed using the same procedures.
The limit of detection (LoD) and the limit of quantification (LoQ) of phenanthrene in animals were 4.5 − 10.5 and 16.5 − 37.8 mg phenanthrene/kg dry weight, respectively. The LoD and LoQ of phenanthrene in soil were 0.11 and 0.36 mg phenanthrene/kg dry soil, respectively. Recovery was tested by spiking uncontaminated animal material with known amounts of phenanthrene and ranged between 93.2 and 108.4%, with an average (± standard deviation) of 101 ± 6%.
Phenanthrene concentrations in animal tissues were measured according to the method described by Holmstrup et al. (Holmstrup et al., 2014 ). In brief, the adults from each replicate were transferred to a 1.5 mL brown glass vial, and 500 µL of acetonitrile (VMR international, USA) was added. The vials were placed in a sonicator (Thermo, Germany), sonicated for 90 min on ice, kept at room temperature for 24 h, frozen at -18 °C for 24 h and kept at room temperature for another 24 h. The samples were sonicated again for 90 min on ice and then transferred to 1.5 mL tubes for brief centrifugation (3 min at 2,400 g). The supernatant from each tube was transferred to an autosampler vial and stored at -80 °C until phenanthrene analysis by GC‒MS (GCMS-QP2010, Shimadzu, Japan). Phenanthrene standards, including blanks, were run in parallel and subjected to the same extraction procedure.
Phenanthrene in soil samples (1 g fresh weight) was extracted with 4 mL of acetonitrile by shaking at 200 rpm for 24 h, followed by centrifugation at 1000×g for 5 min. The supernatants were transferred to autosampler vials and analysed as described above. For quality control, blank medium and uncontaminated soil were analyzed using the same procedures.
The limit of detection (LoD) and the limit of quantification (LoQ) of phenanthrene in animals were 4.5 − 10.5 and 16.5 − 37.8 mg phenanthrene/kg dry weight, respectively. The LoD and LoQ of phenanthrene in soil were 0.11 and 0.36 mg phenanthrene/kg dry soil, respectively. Recovery was tested by spiking uncontaminated animal material with known amounts of phenanthrene and ranged between 93.2 and 108.4%, with an average (± standard deviation) of 101 ± 6%.
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