Electronic Supplemental Material

Details of UPLC methods and respiration experiments

1) Ultra Performance Liquid Chromatography (UPLC) pigment measurements

Samples for this analysis were snap frozen in liquid nitrogen directly at the end of the experiment and stored at -80°C until further processing. Specimens were extracted in

800 l of pre-chilled (4°C) buffered methanol (98% MeOH/2% 0.5 M tetrabutylammonium acetate [TBAA] pH 6.5) and briefly crushed with a teflon pestle.

Samples were then sonicated for 10 s and extracted for 60 min on ice in the dark.

After centrifugation (5 min, 12000 RPM, 0°C), the supernatant was carefully removed and placed in a separate tube. The remainder was extracted for a second time with 800

l buffered methanol and supernatants combined. Supernatants were subsequently filtered (0.2 µm) and stored at -20°C in amber glass vials until injection into the

UPLC on the same day. Immediately prior to injection, 400 µl of the filtered sample was diluted with 1000 µl of 28 mM TBAA, pH 6.5. Chromatography was carried out on a Waters Acquity UPLC system fitted with photodiode array (PDA) detection using a 35 µl injection volume (diluted extract). Pigments were separated on an

Acquity UPLC BEH C8 column (2.1 x 150 mm; 1.7 µm) at a constant flow rate of

0.45 ml min-1 and a column temperature of 60°C using a binary gradient run over 20 min with solvent A (70:30, methanol: 28 mM TBAA ph 6.5) and solvent B (50:50, methanol: acetonitrile). The gradient conditions were as follows; initial conditions,

95% solvent A, 5% solvent B; 3.74 minutes, 45% solvent A, 55% solvent B; 5.09 minutes, 45% solvent A, 55% solvent B; 7.47 minutes, 5% solvent A, 95% solvent B;

10.50 minutes, 5% solvent A, 95% solvent B; 11.50 minutes, 95% solvent A, 5% solvent B and 20.00 minutes, 95% solvent A, 5% solvent B. Certified reference pigments were sourced from DHI (Denmark) and calibration curves for external calibration were prepared under the same running conditions. Pigments in samples were quantified using these calibration curves after identification via retention time and PDA spectral confirmation.

2) Optode based respirometers

Glass incubation vials (volume: 6.6 ml, ESM Figure 1 A) were equipped with a small stand on which individual foraminifera were placed. A Perspex® stand was needed to separate the foraminifera from a small glass coated stirrer bar (VWR, UK) at the bottom of each vial, while small holes in the stand allowed for free water exchange.

The vials were closed with translucent Perspex® lids containing the oxygen spots on the inside. On the outside of the spot a collar allowed the fiber-optic cable connecting to the measurement unit to remain in a stable position on top of the spot. Each of the four vials was placed in a custom made water bath (1 l volume, ESM Figure 1 B) sitting on a stirring unit. The water bath was connected to the pump of a temperature control (± 0.1°C) unit (Lauda, Germany). The stirrer unit consisted on a small electric engine driving four gears equipped with magnets. These magnets drove the stirrer bars in the vials. The oxygen concentration in each vial was recorded every 15 s using standard settings on the operating software. Oxygen spots were calibrated at the beginning and the end of the measurements using water saturated air as 100% O2 and water treated with Na2SO4 as 0% O2 reference. A

B

Electronic Supplemental Material Figure 1. Details of custom made respirometer setup. A: Glas incubation vial (6.6 ml) with lid containing the optode spot. A: specimen of Marginopora vertebralis is visible on the Perspex stand in the vial. B: Custom made water bath and stirring unit.