Permafluor e

¹⁴C-hexadecane and ¹⁴C-naphthalene radiotracer assays were applied for all microcosms to monitor the microbial degradation rate of n-alkanes and PAHs in water samples, respectively, as described previously [29 ]. For each sample, 8 mL of water subsample was transferred to a headspace-free scintillation vial, and amended with ¹⁴C-hexadecane or ¹⁴C-naphthalene (American Radiolabel Chemicals; ARC). The unit of radioactivity per 8 mL sample was 1.76 nCi. Killed controls were amended with 2 mL 2 M NaOH solution prior to tracer addition. The incubation was also halted by adding 2 mL 2 M NaOH solution. Afterwards, 1 g of activated carbon (Sigma Aldrich, Burlington, MA, USA) was added to the water for absorbing the remaining ¹⁴C-hexadecane/¹⁴C-naphthalene. The water was then transferred to a 250 mL flask. A total of 5 mL H₃PO₄ (≥80% wt) was added to release the ¹⁴C-dissovoled inorganic carbon (DIC). The ¹⁴C-CO₂ was trapped by Carbo-Sorb (Perkin Elmer, USA) in an acid digestion system as described [29 ]. The radioactivity measurements were performed with a Beckman LS-6500 liquid scintillation counter (Beckman, Fullerton, CA, USA) with scintillation cocktail (Permafluor^® E+, Perkin Elmer, USA). The rate of ¹⁴C-hexadecane/naphthalene oxidation was calculated as described previously [29 ].

Cellular uptake of ¹⁴C-glucose can be used to evaluate the glucose-uptake capacity of tumor cells. Cells were cultured in 12-well plates, then detached, and washed twice. Subsequently, they were incubated in 500 µL of DMEM containing 2 µCi/mL of ¹⁴C-glucose for 1 h at 37°C. Then, the cells were washed twice with ice-cold PBS. Cell lysates were produced using 500 µL of 0.1 M NaOH. Then, the radioactivity of the whole-cell lysates was assayed using a scintillation counter (LS 6500; Beckman Coulter, Fullerton, CA, USA). These readouts were normalized to the corresponding protein amounts (Beyotime Institute of Biotechnology, Beijing, China). Experiments were carried out independently in triplicate.
For ¹⁴CO₂-release assay, cells cultured in 100-mm dishes were placed in an airtight chamber. Five milliliters of DMEM containing ¹⁴C-glucose (2 μCi/mL) were added and the cells were incubated for 2 h at 37°C with constant airflow (5% CO₂ and 95% air, 15 mL/min); ¹⁴CO₂ was trapped using 16 mL of an amine-based absorber. Four milliliters of absorber-trapped ¹⁴CO₂ were transferred to vials containing 12 mL of Permafluor^® E+ (Perkin Elmer, Waltham, MA, USA) and radioactivity was counted using the liquid scintillation counter. The cell residue was used to determine protein content.

Trifluoroacetic acid (TFA), dimethylsulfoxide (DMSO), and acetonitrile used for chromatographic analysis were high-performance liquid chromatography (HPLC) grade or above and were purchased from Thermo Fisher (Waltham, MA, USA) or another commercial supplier. Combustaid, Carbo-Sorb E absorber, PermaFluor E, Ultima Gold, and Ultima-Flo M scintillation fluids were obtained from PerkinElmer (Waltham, MA, USA). All other reagents were analytical or American Chemical Society reagent grade.

Dry plant and soil material was ground and ¹⁴C in samples quantified by liquid scintillation counting (LSC) of ¹⁴CO₂ produced by dry combustion (Packard 307 sample oxidizer; 6 ml Carbosorb E mixed with 12 ml Permafluor E, Perkin Elmer).
Soil microbial C was determined by chloroform fumigation‐extraction (Vance, Brookes, & Jenkinson, 1987), with some modifications (Supporting Information). For L. alpina, ¹⁴C in microbial extracts was below the detection limit, most likely because of the lower amount of ¹⁴C applied during pulse‐labelling.