Instrument Type: Refractometer

Push cores were collected across multiple dives of HOV Alvin during cruises AT18-02 and AT26-13 aboard the R/V Atlantis. Cores were also collected during cruises EN527, EN528, EN559, and EN586 aboard the R/V Endeavor.

Cores were collected immediately upon retrieval and stored at 4 degrees C until further processing. Sediment samples were collected from select cores in 3 cm intervals (reported as average depth below seafloor, in centimeters) for geochemical and rate analysis, using aseptic techniques at in situ temperature (4 degrees C). Methane samples were collected first: A 3 ml subsample was collected in a cut-end syringe, placed in a 20 ml serum vial containing 4 ml 2M NaOH to stop microbial activity, sealed with a butyl rubber stopper, crimp sealed, vortexed to homogonize, and stored at room temperature until analysis. Porewater samples were extracted from whole sediment under pressure into acid-washed syringes using a custom argon-purged squeezer as described by Joye et al., 2004. A subsample was collected for sulfide determination and analyzed on board using Cline (1969) colorometric methods. DIC samples were collected in 20 ml Hungate tubes and sealed with butyl stoppers. Samples were preserved with 1 ml saturated CuSO4 and 1 ml of 56 mM NaMoO4 in 10% v/v H3PO4. The headspace DIC was methanized and analyzed by GC-FID. Subsamples were collected for determination of porewater pH and salinity: pH was measured on board using a ROSS pH electrode, calibrated with salinity corrected buffers (pH 4, 7, and 10) chilled to in situ temperature (Bowles et al., 2011), and salinity was measured visually on a 500ul subsample using a handheld refractometer (Cole-Parmer RSA-BR60). The remaining sample was filtered-sterilized through a washed 0.2 um Target filter and subsampled further for ammonium, major ions, and nutrients.

A 2 ml ammonium subsample was immediately analyzed on board using the method of Soloranzo (1969). A 5 ml subsample for major ions (sodium, magnesium, potassium, calcium, sulfate, and chloride) was preserved with concentrated nitric acid (0.1 micromole per L final concentration) and stored at room temperature until analysis by ion chromatography (Joye et al., 2004). The remaining nutrient subsample was frozen at -20 degrees C until shore-based laboratory analysis. Total dissolved nitrogen (TDN) was analyzed via the oxidative combustion-chemolumiescence technique of Salgado and Miller (1998) using a Shimadzu TOC 5000 coupled to an Antek model 7020 NO analyzer (Joye et al., 2004). NOx (nitrate + nitrite) was analyzed on a Lachat FIA 8000 Autioanalyzer using method 31-107-04-1-A, phosphate via the molybdate blue colorometric method, total dissolved phosphate (TDP) by high-temperature combustion and hydrolysis (Monaghan and Ruttenberg 1999), dissolved inorganic carbon (DIC) by gas chromatography (flame ionization detector; GC-FID), dissolved organic carbon (DOC) by oxidative combustion-infrared analysis, methane by GC-FID, and stable 13C isotopes of DIC by headspace analysis using a Picarro G2201-i isotope analyzer (Joye et al., 2004, Bowles et al., 2016). Whole sediment subsamples for radiotracer sulfate reduction rates and anaerobic oxidation of methane rates were collected in triplicate from a parallel core and were handled, injected, incubated, and analyzed using the methods described in Joye et al., 2004.

Sediment cores were retrieved from bioturbated, intertidal sediments at low tide with a 7.5 cm (inner diameter) clear Plexiglas liner by pushing it directly into the sediment with minimum pressure as not to artificially force the sediment horizons together. The end of the core (i.e., the deepest horizon) was plugged with a rubber stopper and the sediment core was placed on ice. Typical transport back to the laboratory for pore water extraction was 0.5 hours. Sediment temperature and bottom water salinity were recorded at the time of sampling with an alcohol thermometer and refractometer, respectively.

Once back to the laboratory, the cores were removed from ice and 5 cm Rhizons (0.16-0.19 um pore size) were inserted into pre-drilled 7 mm holes at 1 cm depth intervals to a depth of 10 cm. Pore waters were extracted by pulling negative pressure on the Rhizon with a 10 mL sterile syringe and holding the syringe plunger in place with a small wooden block placed between the syringe body and the plunger. Once pore water was extracted in the syringe, it was removed from the Rhizon, dispensed into a 15 mL centrifuge tube, and 250 uL of pore water was immediately transferred to 250 uL of Ferrozine buffer (10 mM in 50 mM HEPES buffer) and read on a MultiSkan MCC plate reader at 562 nm absorbance. The sediment core was then extruded and sliced into 1 cm intervals and dried in an oven at 70-80 degrees C for 24 hours, and then poorly-crystalline iron oxides (i.e., ferrihydrite and lepidocrocite) were extracted with 1 M hydroxylamine HCl in 25 % acetic acid (v/v) for 48 hours on a rotating shaker at 200 rpm. The extractions were allowed to settle for a few hours, then 10 uL was diluted into 990 uL (1:100 dilution) of distilled water containing 100 uL of Ferrozine buffer. The samples were read as above at 562 nm on the Multiskan MCC plate reader.

Note: data were not collected for months of August and October .