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Offshore interstitial water sampling and analysis

Pore water sampling

Cores were sampled for pore water immediately on recovery, using either Rhizon samplers or squeezers. Rhizon samplers (CSS-F 5 cm; Rhizosphere Research Products, Netherlands) are narrow elongated cylindrical filters (0.2 µm pore size; 5 cm long; 130 µL volume) with a stiff plastic core (Seeberg-Elverfeldt et al., 2005; Dickens et al., 2007). Before use, each Rhizon was soaked for several hours in ultrapure water (Elga Purelab Classic UV), which was subsequently ejected under pressure with a syringe and discarded prior to sampling. A 3.8 mm hole was drilled through the plastic core liner using a spacer on the drill bit to avoid penetrating the core itself. A Rhizon sampler with the same diameter as the hole was then inserted into the core. Negative pressure was applied by attaching a 20 mL all-plastic pulled-back syringe to the Rhizon sampler, in which the pore water was collected. The sediment cores remained capped throughout this process to maintain an environment as similar to the original sediments as possible.

When Rhizon sampling took place before core logging (i.e., paleoenvironmental holes), Rhizon samplers remained in the cores for 2–3 h. When Rhizon samples were taken after Fast-track core logging (i.e., microbiology holes), the Rhizons remained in the core for as long as 9 h. Usually, two Rhizons were used per core section, spaced 10 cm apart to obtain sufficient pore water from a single interval while minimizing overlap.

Whole-round sections 10 cm in length obtained from the microbiology cores were squeezed in a Teflon-lined titanium squeezer of piston-cylinder design (described in detail at Whole rounds were nearly always cut from the bottom of the first core section (unless both sections were sampled), as this material was most likely to be undisturbed. Before the whole-round samples were prepared for squeezing, a preprepared, rinsed Rhizon was inserted to obtain the first 1–2 mL of pore water to fulfill a sample request for postcruise research. Whole rounds were scraped clean on all surfaces to remove drilling mud and material from the outer surface that may have been smeared down the core liner. They were then cut to <54 mm diameter to fit into the squeezer. Once in the squeezer, sediment was contained when under pressure (up to 10 tons/inch2, equivalent to 137,900 kPa, applied by a hydraulic press) by a circular titanium screen at the base of the cylinder overlain by nylon mesh and a paper filter (Whatman 1; 55 mm), through which the water was pushed to an online filter (Sartorius; 0.2 µm; 25 mm; nylon) and through that into a 20 mL all-plastic syringe. All parts of the squeezer that come in contact with either the sediment or pore water are made of polytetrafluoroethylene (PTFE), titanium (grade 2), or polyamide plastic (Delrin).

We collected pore water reference samples, including seawater from each site, tap water, artificial seawater, and the different mud types and additives used while drilling. These samples were treated like pore water samples, analyzed, and split to cover the IODP required measurements.

After sampling, all syringes were placed in a glove bag under nitrogen, and the extracted pore water was transferred to a 20 mL polypropylene vial. When multiple Rhizon samples were taken from one core section, pore water samples were combined in a single vial. Oxygen was monitored in the glove bag using a Drager Pac 5500 oxygen sensor and was typically <1 vol% (Sites M0059–M0062). At Site M0063, however, it was not possible to maintain such low oxygen concentrations, possibly as a result of degradation of the closure mechanism on the glove bag. For those samples and for all following sites, one of the Rhizon syringes was transferred directly to a cation vial. The sample was subsequently split into six fractions: a pore water sample for total sulfide (add 1 mL of sample to a vial prefilled with 0.4 mL 5% Zn acetate), cations and trace elements (1–3 mL of sample acidified with 1% concentrated trace metal grade HNO3 per sample), pH/alkalinity (0.5 mL), ammonium (0.2–0.5 mL), salinity (0.5 mL), and anions (1–3 mL).

Pore water analysis

Pore waters from all holes were analyzed on board for pH, alkalinity, ammonium, and salinity. In addition, sulfide was analyzed on the first paleoenvironmental hole of every site where a microbiology hole was cored (Sites M0059, M0060, M0063, and M0065). Pore water pH was measured using an ion-specific electrode (Mettler Toledo) with a 2-point calibration on 0.2–0.5 mL of pore water. The same sample was then utilized for the measurement of alkalinity by single-point titration to pH 3.95 with 0.01, 0.05, or 0.2 M HCl according to standard procedures (Grasshoff et al., 1983). Ammonium was measured by conductivity after separation as NH3 through a PTFE membrane in a flow-through system. In the latter technique, modified after Hall and Aller (1992), ammonia is stripped from a 100 µL sample by an alkaline carrier solution (0.2 M Na citrate in 10 mM NaOH), passed through a 200 mm × 5 mm PTFE membrane, and redissolved as NH4+ in an acidic solution (1 mM HCl). Ammonium ions were then measured as the resulting conductivity signal in the acidic carrier in a microflow-through cell. All samples collected for pH, alkalinity, and salinity analyses were stored at 4°C, and measurements were conducted within 24 h or, for ammonium, within 24–48 h of sample collection.

Total sulfide was measured spectrophotometrically applying the methylene blue method of Cline (1969) using a DR 6900 Hach Lange (Berlin, Germany) spectrophotometer. Total sulfide is referred to as H2S throughout the text for simplicity, though H2S was not quantified. Salinity was determined by optical refraction (Krüss Optromic digital refractometer DR 6300), calibrated with International Association for the Physical Sciences of the Oceans (IAPSO) seawater standards by OSIL having salinities of 10, 30, 35, and 38.

Subsamples for cations, trace elements, and anions were stored at 4°C for further onshore analysis at the University of Bremen (Germany) via inductively coupled plasma–optical emission spectrometry (ICP-OES; Al, B, Ba, Ca, Fe, K, Mg, Mn, Na, P, S, Si, Sr, Ti, Li, Mo, Rb, V, and Zr) and ion chromatography (Cl, Br, and SO4) measurements.

Methane sampling and analysis

Methane samples were taken from all cores when the core material was intact and thus suitable for sampling. Methane concentrations were determined for each core of the first paleoenvironmental hole on the microbiology sites (Sites M0059, M0060, M0063, and M0065) for chemical zonation, following standard protocols for headspace sampling and analysis. A 5 cm3 sediment sample was collected with a cut-off disposable syringe from the freshly exposed end of every section within the uppermost 20 m of each hole. Deeper than this, the sampling frequency was reduced and one sample was taken from the bottom of the first section within each core. The sample was extruded into a 20 mL glass vial filled with 8 mL of 1 M NaOH solution, immediately crimp-sealed with a gas-tight septum, and stored upside down at room temperature prior to analysis. After the sample was shaken and left for gas equilibration, 250 µL of headspace volume was manually injected using a gas-tight glass syringe into an Agilent A7890 gas chromatograph (GC) equipped with a PLOT 5A column (30 m × 0.53 mm, inner diameter = 50 µm) and coupled to a flame ionization detector (FID). Helium was used as carrier gas with a constant flow of 5 mL/min. The oven was held isothermal at 30°C for 3 min. Quantification of methane was achieved by comparison of its chromatographic response with an external three-point calibration curve using 0.01%, 0.1%, and 1% methane standards.

The concentration of methane in interstitial water was derived from the headspace concentration by the following equation (Shipboard Scientific Party, 2003), where methane that remains undetected because dissolution in the aqueous phase is minimal (e.g., Duan et al., 1992), is not accounted for:

CH4 = [χM × Patm × VH]/[R × T × ϕ × VS],


  • VH = volume of sample vial headspace,
  • VS = volume of whole sediment sample,
  • χM = molar fraction of methane in headspace gas (obtained from gas chromatograph analysis),
  • Patm = pressure in vial headspace (assumed to be the measured atmospheric pressure when the vials were sealed),
  • R = universal gas constant,
  • T = temperature of vial headspace in degrees Kelvin, and
  • ϕ = sediment porosity (determined from moisture and density measurements on nearby samples).

Onshore science party chemical analyses

Pore water analysis

A total of 689 filtered (0.2 µm) and acidified (10 µL of concentrated HNO3/mL) pore water samples were analyzed for cations and trace metals using analytical equipment at the University of Bremen. All samples were diluted 10-fold with 1% HNO3 and analyzed for Al, B, Ba, Ca, Fe, K, Mg, Mn, Na, P, S, Si, Sr, and Ti using a Varian Vista Pro CCD ICP-OES equipped with a sea-spray nebulizer and a cyclon spray chamber. Trace elements (Al, Ba, Fe, Li, Mn, Mo, P, Rb, Ti, V, Zn, and Zr) were analyzed on a second set of 10-fold diluted samples using an Agilent Technologies 700 Series ICP-OES equipped with a K-style conical nebulizer. Standardization was performed against multielement solutions prepared from commercial standards with either a 1% HNO3 matrix adjusted to a NaCl concentration similar to the matrix in the samples (for trace element samples with salinity >5) or 1% HNO3 only (all other samples). Calibration standards for major elements and trace elements were prepared using IAPSO seawater and National Institute of Standards and Technology Certified Reference Material (NIST CRM). Measurement precision was ±3% for major elements and ±5% for trace elements.

Trace element analyses for Al and Zn were either below the detection limit or showed contamination and therefore are not reported for all sites. Rubidium, Mo, Ti, V, and Zr had concentrations above the detection limit, and results were consistent between holes for some of the sites. Table T2 indicates which sites had acceptable data (denoted by “Y”). These data are listed in the tables in each site chapter. For elements analyzed for both major and trace elements, reported values came from the major element analyses, with the exception of Ti.

A total of 711 filtered and unacidified pore water samples were analyzed for anions (chloride, bromide, and sulfate) using a Metrohm 882 compact ion chromatograph at the University of Bremen. A 40-fold dilution of IAPSO seawater and standards prepared from commercial single anion standards were used for calibration. Measurement precisions were ±0.6% for Cl, ±3.5% for Br, and ±1.6% for S (1σ).

Bulk geochemical analysis of sediments

Approximately 10 cm3 of sediment was freeze-dried and ground to a fine powder using an agate mortar. Total carbon (TC), total organic carbon (TOC), and total sulfur (TS) were measured using a LECO CS-300 carbon-sulfur analyzer. Approximately 65 mg of the homogenized sample was weighed in a ceramic cup and heated in a furnace. The evolved CO2 and SO2 were then measured with a nondispersive infrared detector to provide a measure of the sedimentary TC and TS content. To determine the TOC content, sediments (~65 mg) were decalcified using 12.5% HCl to remove carbonate species and analyzed as described above. Total inorganic carbon (TIC) was determined by subtracting TOC from TC. All data are reported in weight percent (wt%) dry sample with an analytical precision <3% absolute based on replicate sample analysis.