Ionpac as18

Sulfate and chloride concentrations were measured by suppressed ion chromatography with an ICS2500 system (Dionex) equipped with an eluent generator (EG50) and KOH eluent generator cartridge (EGC III KOH). The column was a Dionex IonPac™ AS18 operated at 30°C. KOH concentration started at 20 mmol L⁻¹ and was raised to 32 mmol L⁻¹ at the end of the analysis run at 15 min. Hydrogen sulfide (sum of H₂S, HS⁻ and S²⁻) was determined spectrophotometrically at 670 nm (FLUOstar Omega, BMG Labtech GmbH, Orthenberg, Germany) on zinc-preserved porewater samples by the methylene blue method (Cline, 1969 ; Reese et al., 2011 (link)). DIC was measured immediately after the cruise on headspace-free porewater samples stored at 4°C. Samples were transferred to sealed exetainers and acidified with 85% (v:v) phosphoric acid. After 24 h of equilibration time, the produced CO₂ was measured from the headspace of the exetainer by a Delta V™ isotope ratio mass spectrometer (IRMS, Thermo Scientific).

The concentrations
of major ions were measured using two Dionex Integrion high-pressure
ion chromatography (HPIC, Thermo Fischer Scientific, USA) instruments.
Samples and blanks were prepared by filling ion chromatography vials
with 5 mL of the ∼10 mL final eluates from the solid-phase
extractions. The autosampler split the volume of each sample between
the two HPIC instruments, with one column to quantify the cations
(IonPac CS12A, Thermo Fischer Scientific, USA) and the other to quantify
the anions (IonPac AS18, Thermo Fischer Scientific, USA). Here, we
measured the concentrations of cations lithium, sodium, ammonium,
potassium, magnesium, and calcium and anions fluoride, acetate, formate,
chloride, nitrite, carbonate, sulfate, and nitrate in each solution.
The concentrations of the corresponding sample blanks were subtracted
from the samples. Concentrations in solution were calculated from
calibration curves for each ion of interest. Ion concentrations in
particle samples were derived using the total volume of air sampled.

The pH of the culture samples obtained during the incubation was measured using a pH meter (Seven Compact^™ S220, Mettler Toledo, United States). The cultures were then centrifuged for 10 min at 3,900 g (Allegra X-22R, Beckman Coulter, United States). The supernatant obtained after centrifugation was analysed for total alkalinity (TA), anions, cations, and total nitrogen concentrations. In brief, TA was measured using a G20 compact titrator (Mettler Toledo, United States); 40 ml of supernatant was taken in a beaker and titrated with 0.2N H₂SO₄ until the samples reached an endpoint of pH = 4.3 (Vadlamani et al., 2019 (link)). Bicarbonate and carbonate concentrations were calculated using the measured pH and total alkalinity (TA) (Vadlamani et al., 2017 (link); Vadlamani et al., 2019 (link)).
The nitrate concentration was measured using an ion chromatograph equipped with an IonPac AS18 anion and a conductivity detector (DIONEX ICS 2000; Thermo Fisher, United States) (Vadlamani et al., 2017 (link); Vadlamani et al., 2019 (link)). The ammonium concentration was determined colorimetrically as previously described (Sims et al., 1995 (link)). The total nitrogen concentration was assessed using a scaled down version of the Persulfate Digestion method (Hach Method 10071, Hach, United States) (Vadlamani et al., 2017 (link); Vadlamani et al., 2019 (link)).

Major cation and anion concentration of the hot spring water was measured using ion chromatograph (IonPac AS18 4 × 250 mm for anion, ICS 600, ThermoFisher, Carlsbad, CA, USA). The DOC and TOC contents were measured by using an NC 2100 Elemental Analyzer (multi N/C 2100, Analytic Jena, Jena, Überlingen, Germany). Before TOC measurements, the sampled sediments were fumigated with HCl to remove carbonates.