WOBEC SOP Water column CTD

Weddell Sea Observatory of Biodiversity and Ecosystem Change – WOBEC Standard Operating Procedures Technical Documentation December 2025 To be cited as Lenss, Megan, Nils Van den Steen, Anette Wold, Sebastien Moreau, Karley Campbell, Maria van Leeuwe, Jacqueline Stefels, and Anton Van de Putte. ‘WOBEC SOP Water Column CTD’. Zenodo, 15 December 2025. https://doi.org/10.5281/zenodo.17936552 Co-funded by the European Union (c) The authors This report is licensed under the Creative Commons Attribution International license Versions Date Comment Responsible Approved 4 December 2024 SOPs for water column sampling of parameters related to protists and biogeochemistry M. Lenss S. Moreau A. Wold K. Campbell M.A. van Leeuwe J. Stefels 12/12/2025 Formatting N. Van den Steen Table of Contents 1 Description of sampling gear .......................................................................................................... 4 2 Sampling gear deployment .............................................................................................................. 4 2.1 Deployment ............................................................................................................................................. 4 2.2 Calibration ............................................................................................................................................... 5 2.3 Post-Deployment ................................................................................................................................... 5 2.4 Common Issues ...................................................................................................................................... 5 3 Sample sorting-collection-preservation procedure: Water samples from Niskin bottles .................................................................................................................................................................... 6 3.1 Salinity samples ..................................................................................................................................... 6 3.2 Dissolved in organic carbon / total alkalinity ............................................................................ 6 3.3 Oxygen isotope ratios (δ18O) ............................................................................................................ 6 3.4 Methane .................................................................................................................................................... 7 3.5 Inorganic nutrients ............................................................................................................................... 7 3.6 Particulate organic carbon / nitrogen and their stable isotopic δ13C and δ15N compositions. ................................................................................................................................................. 8 3.7 Chlorophyll a ........................................................................................................................................... 8 3.8 Biogenic silica ......................................................................................................................................... 9 3.9 Phytoplankton taxonomy................................................................................................................... 9 3.10 Flow cytometry ................................................................................................................................... 9 3.11 Net community production using O2 optodes ...................................................................... 10 3.12 DMSP .................................................................................................................................................... 11 3.13 Net community production by recording uptake of the 13C stable isotope. ........... 11 4 Sample analysis method ................................................................................................................ 12 4.1 Salinity samples .................................................................................................................................. 12 4.2 Dissolved in organic carbon / total alkalinity ......................................................................... 12 4.3 Stable oxygen isotopes ..................................................................................................................... 12 4.4 Methane ................................................................................................................................................. 12 4.5 Inorganic nutrients ............................................................................................................................ 13 4.6 Particulate organic carbon / nitrogen and their stable isotopic δ13C and δ15N compositions. .............................................................................................................................................. 13 4.7 Chlorophyll a ........................................................................................................................................ 13 4.8 Biogenic silica ...................................................................................................................................... 14 4.9 Phytoplankton taxonomy................................................................................................................ 14 4.10 Flow cytometry ................................................................................................................................ 14 4.11 Net community production using O2 optodes ...................................................................... 14 4.12 DMSP .................................................................................................................................................... 14 4.13 Particulate organic carbon – 13C. ............................................................................................. 15 Method responsible: Sebastien Moreau/Jacqueline Stefels 1 Description of sampling gear Water column structure is sampled with the CTD-rosette installed aboard R/V Polarstern, which is equipped with a temperature and conductivity sensor, a pressure sensor, oxygen and fluorescence sensors, and an altimeter, among others. Processing of several samples taken from the CTD-rosette will include chemical use. A list of the necessary chemicals and their corresponding parameter is provided below. It is the responsibility of the individual using the chemical and processing the sample to operate in a manner that is safe for themselves, others on the ship, and the environment being sampled. Safety Data Sheets (SDS) should be readily available in the labs and proper chemical storage systems should be decided on and implemented before the cruise, including locked chemical storage cabinets and designated workspaces for radioisotopes. Chemical Parameter Safety consideration Saturated mercury chloride (HgCl2) DIC and CH4 Highly toxic Methanol Chlorophyll a Carcinogenic Glutaraldehyde Taxonomy and flow cytometry Carcinogenic Formaldehyde Taxonomy Carcinogenic and mutagenic Hydrochloric acid (HCl) Several Corrosive Ethanol Several Toxic NaOH DMSP Corrosive 2 Sampling gear deployment 2.1 Deployment Deployment should follow typical routines aboard R/V Polarstern. Here we provide a few points from deployment routines aboard R/V Kronprins Haakon. 1. Start data acquisition just before the CTD leaves deck to help with assessing surface pressure adjustment 2. Soak for 1 minute at 5 m (10 m if very cold and there is a risk of freezing during deployment/deck sampling), bring back up to surface (just submerged) and leave there for 3 minutes before lowering to max depth. Rest at max depth for at least 1 minute 3. Lower speed should be between 0.8 and 1 m s-1 and if LADCP is mounted lowering speed should be reduced to 0.7 m s-1 4. Water sampling should be taken at standard depths that are pre-determined before deployment, that is except for the depth of the maximum CHl-a which is determined in real time. Niskin bottles are closed on the upcast by stopping the CTD at desired depth and waiting for at least one minute before firing the bottle. 5. Stop data acquisition when the CTD is back on deck. 2.2 Calibration Annual calibration of all CTD sensors should be documented and samples for calibration of conductivity cell(s) should be taken regularly throughout the cruise. 2.3 Post-Deployment Postprocessing should follow standard routines aboard R/V Polarstern with raw data available and accessible for postprocessing by individual researchers. Metadata of the cast should be recorded in the CTD file and on the CTD sampling sheet. A sampling log sheet should be used for overview of type of water sample taken and sample number. The order of the samples on the CTD log sheet should reflect the order in which samples should be taken from the Niskin-rosette. Below is a suggested ordering of sampling from a ship CTD taken from the Norwegian Polar Institute 2024 Arctic Ocean cruise. 1. Methane 2. DIC/AT 3. DMSP 4. Stable oxygen isotopes 5. Inorganic nutrients 6. CDOM 7. Particle absorption 8. TSM 9. Chlorophyll a and phaeopigments 10. POC/PON 11. FCM 12. Phytoplankton taxonomy 13. Salinity 2.4 Common Issues Well organized and intuitive log sheets and labeling systems should be used commonly throughout the cruise. Water sampling budgets should be agreed on well before deployment and sampling order should be made clear before the Rosette is back on deck. Vinyl gloves (not nitrile) should be used during sampling and readily available in the CTD hanger. 3 Sample sorting-collection-preservation procedure: Water samples from Niskin bottles Water samples taken from the ship’s Niskin bottles are described below in separate subchapters. 3.1 Salinity samples • Rinse bottle and bottle cap three times. • Dry off bottle neck and cap with paper towel before closing the bottle. • Remember to place plastic insert before closing the cap. Pay special attention to threads on the bottle and in the cap when screwing close. • Note down cast number/station number, Niskin bottle number and sample bottle number on the sampling log. 3.2 Dissolved in organic carbon / total alkalinity • Total DIC measures the sum of bicarbonate, carbonate, carbonic acid and dissolved CO2 in seawater. • Use a tube to gentle fill a 250 mL borosilicate bottle directly from the Niskin bottle. Take care to avoid air bubbles in the tubing. • Overfill sampling bottle 1-2 volumes for rinsing and removal of air bubbles. Carefully remove the tubing out of the bottle and close the cap. Some headspace (a few mL) is okay. • After all bottles have been filled, add 60 µL saturated mercuric chloride (HgCl2) to each sample by submerging the pipette tip into the sample. Close the bottle with the blue cap. Do not shake or mix. • Store samples in a cool and dark place. Do not freeze. 3.3 Oxygen isotope ratios (δ18O) • Ratios of 16O to 18O (δ18O) in the H2O molecule are measured to a very high accuracy. Samples of δ18O are collected to determine the fractions of river water and sea-ice meltwater in the ocean. This sample is not related to dissolved oxygen. • Sample rinse a 20 mL glass scintillation vial and cap three times. • Fill the vial with seawater directly from the Niskin creating a convex meniscus. • Carefully apply the cap and check for small air bubble. • When all δ18O samples have been collected, dry the vials, tighten the caps and seal with Parafilm. • Store δ18O samples at room temperature or in a fridge, avoid freezing. 3.4 Methane • Methane is a volatile and relatively insoluble trace gas and the concentration in the sample will be affected by prolonged contact with air. Therefore, it should be sampled immediately after retrieval of the rosette. • Attach a flexible silicone sampling tube (about 30 cm long) to the Niskin bottle and flush the tube with sample to expel all air bubbles. • Rinse a 160 mL serum bottle with sample water then place the end of the tube at the bottom of the bottle. Allow the bottle to overflow by at least two volumes. • Slowly withdraw the tubing from the bottle and reduce flow towards the neck of the bottle by pinching the tube. The bottle should be completely full, free of bubbles, and have a slightly convex meniscus over the opening. • Immediately add 50 µL saturated mercuric chloride (HgCl2), then seal the bottle with a metal seal and butyl rubber septum using the crimping tool. • Store the samples in a cool, dark place (refrigeration at +4C). • Rinse crimping tool in freshwater after sampling. 3.5 Inorganic nutrients • Set up a clean working space and use vinyl gloves. Acid wash sampling vials (20 mL scintillation vials), syringes, and swinnex before sampling. Rinse sampling vials, syringes, and swinnex with MiliQ after acid bath and before use. • Assemble a swinnex with a pre-combusted GF/F filter. Place syringe into undiluted melt (chemistry core) and take up a small amount of sample to rinse the syringe. Repeat three times. • Take up a full syringe of sample (about 60 mL), push out air, and attach swinnex. Using steady but light pressure, gently push sample through the filter at a drip-dripdrip speed. Rinse the filter with sample, and then rinse the sampling vial and cap with filtered sample 3 times. • Fill sampling vial with filtered sample, leaving a small amount of headspace for freezing. • Store samples at -20C. 3.6 Particulate organic carbon / nitrogen and their stable isotopic δ13C and δ15N compositions. • Rinse a 2 L clear plastic bottle with sample three times using seawater directly from the Niskin. • Completely fill sample bottle and take to the lab for filtration. • Filter up to 2 L of sample (depending on particle concentration) onto precombusted 25 mm GF/F filters. Filters should be combusted at 450C for 12 hours before the cruise. Remember to gently mix the bottle before filtering to ensure that particles are suspended. Also ensure that the mesh side of the filter is facing up on the filter holder, not the soft waves side (back side). • Filter under low vacuum pressure (about -30 kPa) and cover the funnels with aluminum foil when filtering. Rinse the funnel with filtered seawater once the sample has been filtered. Take care not to let the filters dry out. • After filtration, use forceps to carefully place GF/F into Pall filter slides. Place slides into oven set at 60C and allow them to fully dry (about 24 hours). • Store samples at room temperature. Wrap together filter slides from one station in aluminum foil and keep them in a labelled Ziploc bag. • Prepare a reference filter (blank) for each station by filtering filtered seawater onto a pre-combusted filter. The reference filter will get a normal running number, so make sure to make a note on the CTD log sheet to avoid confusion with numbering on following casts. 3.7 Chlorophyll a • Rinse a 1 L brown plastic bottle with sample three times using seawater directly from the Niskin. Chl a is light sensitive, so keep samples in the dark as much as possible. • Completely fill sample bottle and take it to the lab for filtration/extraction. • Filter up to 1 L of sample depending on biomass (a light color on the filter is enough) onto 25 mm GF/F filters. Remember to gently mix the bottle before filtering to ensure that particles are suspended. Also ensure that the mesh side of the filter is facing up on the filter holder, not the soft waves side (back side). • Filter under low vacuum pressure (about -30 kPa) and cover the funnels with aluminum foil when filtering. Rinse the funnel with filtered seawater once the sample has been filtered. Take care not to let the filters dry out. • Use forceps to lift the filters into plastic tubes for extraction. • Add 5 mL of methanol to the extraction tubes, close with lid, and cover with aluminum foil to block light. Place in refrigerator (dark, +4°C). • Allow pigments to extract into methanol for 12-24 hours, making sure to note the start and end time of extraction. • Rinse filtration funnels and manifold with freshwater between stations. 3.8 Biogenic silica • Rinse a 1 L plastic bottle with sample three times using seawater directly from the Niskin. • Filter up to 1 L (depending on diatom biomass present) onto a polycarbonate membrane filter (0.8 µm pore size, 25 mm diameter) under low vacuum pressure (- 30 kpa). • Place filter into prelabeled plastic petri dish and place in oven at +60°C to dry, leaving petri dish slightly open to allow for evaporation. • Once filters are dry (ca. 24 hours in the oven), seal petri dishes with parafilm and wrap filter slides from one station together in aluminum foil. Place in labelled Ziploc bag and store at room temperature. • Prepare a reference filter (blank) for each sampling event by filtering a similar volume of filtered seawater onto a filter and treating it the same as the samples. 3.9 Phytoplankton taxonomy • Fill 190 mL of sample directly from Niskin bottle into a measuring cylinder. Sample rinse cylinder before filling. • Decant 190 mL sample into a pre-labelled brown glass (or plastic) bottle). • Bring samples back to the lab for fixation with aldehyde mixture under the fume hood. Add 0.8 mL of 25% glutaraldehyde and allow to fix for approx. 5 min. Thereafter add 10 mL of 20% hexamine-buffered formaldehyde. The final concentration is 0.1% glutaraldehyde and 1% formaldehyde in a 200 mL solution. • Store the samples in cool, dark conditions. Do not freeze. 3.10 Flow cytometry • Prepare for station by adding 38 µl of 25% glutaraldehyde into each pre-labeled 2 mL cryovials (3 per depth) under the fume hood. • Using a syringe directly from the Niskin bottle, rinse a 50 mL falcon tube three times with sample filtered through a 40 µm cell strainer. Then fill the falcon tube with about 40 mL of filtered sample. • Take samples back to the lab and pipette 1.8 mL of filtered sample into the prepared cryovials. • Fix the samples for 2 hours in the fridge and then snap freeze in -80°C. Store samples in cryobox at -80°C.