Delta plus mass spectrometer

The rates at which carbon was fixed by the shallow and mesophotic coral colonies under their natural PAR levels were estimated using ¹³C- labelled bicarbonate according to Tremblay et al.^{4 (link)}. To take into account variation in PAR levels during the day, corals were incubated for 5 h, between 10 am and 3 pm, to cover the maximal daily irradiance⁴⁵ (Fig. S1). For each species and depth condition, 10 coral nubbins, from 10 different colonies were placed in individual beakers filled with 200 mL FSW enriched with 0.6 mM NaH¹³CO₃ (98 atom % ¹³C, #372382, Sigma-Aldrich, St-Louis, MO, USA). After this “pulse” period of 5 h, half of the corals were sampled and stored frozen at −20 °C until further analysis (T₀). The other half was transferred into 200 mL of non-enriched FSW for a chase period of 19 h (T₂₄). The determination of %¹³C enrichment, and total carbon content in the symbionts and host compartments were performed with a Delta plus Mass spectrometer coupled to a C/N analyser (Thermo Fischer Scientific, Bremen, Germany). The natural isotopic abundance of each species at each depth was determined from the corals used for the respiration measurements. Detailed calculations are described in Tremblay et al.^{4 (link)}. Data were normalized to ash-free dry weight of the organisms^{27 (link)}.

Samples for the Dissolved Inorganic Carbon (DIC) concentration and stable isotopic composition were collected from the filtrate of POC samples and processed as described in Remize et al. [33 (link)] and following the protocol of Assayag et al. [68 (link)]. The DIC was measured using a gas bench coupled with a Delta Plus mass spectrometer from Thermo Scientific, Bremen, Germany (GB-IRMS).

Samples for Dissolved Inorganic Carbon concentration and stable isotope composition were collected from the filtrate of POC samples (25 mL). Twelve milliliters was put into Labco Exetainer (Labco, Wales, UK) tubes, poisoned with 20 µL of mercury chloride (HgCl₂) and stored at 4 °C until Gas-Bench Isotope Ratio Mass Spectrometer (GB–IRMS) analysis. The sample preparation for DIC concentration and ¹³C-DIC measurements was made according to Assayag et al. [57 (link)]. Briefly, 1 mL subsample was added to the mixed tube, then 23 droplets of phosphoric acid (H₃PO₄) were introduced into a closed Exetainer tube (Labco, Wales, UK) that is flushed with ultra-pure helium gas to eliminate the residual air and avoid contamination. The H₃PO₄ addition converted all DIC species into CO₂ [57 (link)], and after 15 h, equilibration at room temperature CO₂ was measured in the headspace of a Gas Bench coupled to a Delta Plus mass spectrometer from Thermo Scientific, Bremen, Germany (GB–IRMS).