Overview of the project and objectives: To investigate whether nitrate uptake and processes other than nitrate uptake by phytoplankton are significant and show spatial variability possibly induced by varying availability of Fe and other parameters in the region, seawater was collected from CTD (Conductivity, Temperature and Depth) and TMR (Trace Metals Rosette) casts jointly with the nutrient sampling, as well as well as sea-ice collected from Bio ice-core types on Ice Station, for analysis of nitrate d15N, d18O isotopic composition. Results have been interpreted in the light of prevailing nitrate-nutrient concentrations (Belgian team) and N-uptake regimes for the Ice Stations (new vs. regenerated production and nitrification; see Silicon, Carbon and Nitrogen in-situ incubation Metadata file). Methodology and sampling strategy: Samples for isotopic composition of nitrate were collected from the CTD rosette, TMR and Bio ice-core jointly with the nutrient sampling. Sea-ice sampling: sampling strategy follows ice stations deployment via Bio ice-core type. Most of the time we worked close to / directly on the Trace Metal site following precautions concerning TM sampling (clean suits etc.). When we worked close to the TM site, precautions were not such important because we don't need the same drastic precautions for our own sampling. We work together because we want to propose a set of data which helps to characterize the system of functioning in close relation with TM availability (for that, sampling location have to be as close as possible). All samples were filtered on 0.2 microns acrodiscs and kept at -20 degrees C till analysis in the home-based laboratory. We applied the denitrifier method elaborated by Sigman et al. (2001) and Casciotti et al. (2002). This method is based on the isotopic analysis of delta 15N and delta 18O of nitrous oxide (N2O) generated from nitrate by denitrifying bacteria lacking N2O-reductase activity. As a prerequisite the nitrate concentrations need to be known (nutrients analysis in the home lab.) as this sets sample amount provided to the denitrifier community. Briefly, sample nitrate is reduced by a strain of denitrifying bacteria (Pseudomonas aureofaciens) which transform nitrate into N2O, but lack the enzyme to produce N2. N2O is then analysed for N, O isotopic composition by IRMS (Delta V, Thermo) after elimination of CO2, volatile organic carbon and further cryogenic focusing of N2O (Mangion, 2011). Casciotti K.L., D.M.Sigman, M.G. Hastings, J.K. Bohlke and A. Hilkert, 2002. Measurement of the oxygen isotopic composition of nitrate in seawater and freshwater using the denitrifier method, Analytical Chemistry, 74 (19): 4905-4912. Mangion P., 2011. Biogeochemical consequences of sewage discharge on mangrove environments in East Africa, PhD Thesis, Vrije Universiteit Brussel, 208 pp. Sigman D.M., Casciotti K.L., Andreani M., Barford C., Galanter M. and J.K. Bohlke, 2001. A bacterial method for the nitrogen isotopic analysis of nitrate in seawater and freshwater, Analytical Chemistry, 73: 4145-4153.

Water samples of 1 to 2L from Niskin bottles filled close to the surface, mid mixed layer depth and bottom of the mixed layer were drawn cleanly through a 210um mesh to exclude zooplankton. All samples were filtered as two size fractions, 1.2 to 20um (larger particles excluded by 20um Nitex mesh) and a separate 1.2 to 210um total sample. The filters were 1.2um silver membranes (Sterlitech) 13mm diameter. The samples were preserved by drying at 60C in a dedicated clean oven. Prior to encapsulation, a 5mm diameter subsample was taken for biogenic silica analysis, which is delayed until there has been evaluation of the particle data from the flow cam and UVP. Samples were encapsulated in silver (Sercon sc0037) after acidification and drying. The decarbonated encapsulated POC samples were analysed by elemental analyser at the CSL UTAS by Dr Thomas Rodemann (EA TCD 960C, single point standardisation every 12 samples).  EA detection limit 0.001umol POC. POC and PON are presented as molar units. Blanks were process blanks (seawater) and 7% of the average for the combined data n=177. 1sd=0.12uM. The ctd casts were all given the prefix K, so K001, K002 etc. Not all stations were sampled due to budget constraints. Niskin is the Niskin bottle number.

Overview of the project and objectives: To investigate whether the Nitrogen - Silicon - Carbon biogeochemical system functions in the Antarctic Marginal Ice Zone and shows spatial variability possibly induced by varying availability of Fe and other parameters in the region. This toolbox is part of project 4051 - samples were taken (1) on the same sea-ice site or very close than the one used for Trace Metal sampling; (2) via Trace Metal Rosette TMR; (3) via Conductivity Temperature and Depth CTD Rosette. It is also part of project 4073 since some intercalibration studies were conducted in collaboration with the primary production team. Three main tools were used which can be either independently or intricately studied. For this reason the complete set of sampling done for this stable isotope toolbox is summarized in one excel file which is duplicated and attached to three child metadata records. Same reasoning for raw data acquired on boar and on field information. This parent metadata record has thus three child metadata records. Each of the child metadata files explain individually the different approaches which were treated together by the same team to resolve the main question of sea-ice biogeochemical system functioning via the use of stable isotope ratio tools. The details of each are in the respective metadata records. The data are attached to this metadata record. METADATA FILES are: - 13C, 15N, 30Si in-situ incubation experiments during SIPEX 2 - Nitrogen and oxygen isotopic composition of nitrate during SIPEX 2 - Delta13C signal of brassicasterol and cholesterol in the Antarctic Sea-ice / Is there particulate barium in sea-ice?

Sediment cores were collected from the East Antarctic margin, aboard the Australian Marine National Facility R/V Investigator from January 14th to March 5th 2017 (IN2017_V01; (Armand et al., 2018). This marine geoscience expedition, named the “Sabrina Sea Floor Survey”, focused notably on studying the interactions of the Totten Glacier with the Southern Ocean through multiple glacial cycles. The cores were collected using a multi-corer (MC) and a Kasten corer (KC). The MC were sliced every centimetre, wrapped up in plastic bags, and stored in the fridge. The KC was sub-sampled using a u-channel; and sliced every centimetre once back the home laboratory (IMAS, UTAS, Hobart, Australia). This dataset presents stable isotopes measured in total and fumigated (i.e. organic) sediment samples collected during the IN2017_V01 voyage. The data include the sampling date (day/month/year), the latitude and longitude (in decimal degrees), the seafloor depth (in meter), the sediment core ID, the sediment depth (in cm), the elemental concentration (in %) and the stable isotope (13C, 15N and 34S) compositions reported as delta values (in ‰). This dataset presents stable isotopes measured in fumigated (i.e. organic) sediment samples collected during the IN2017_V01 voyage. The data include the sampling date (day/month/year), the latitude and longitude (in decimal degrees), the seafloor depth (in meter), the sediment core ID, the sediment depth (in cm), the elemental concentration (in %) and the stable isotope (13C and 15N) compositions reported as delta values (in ‰). Sediment samples were dried in an oven at 40°C and ground using a pestle and a mortar. Thirty mg of sediment was weighed into a tin cup for elemental and stable isotope analysis at the Central Science Laboratory (CSL), University of Tasmania. Total carbon (C), nitrogen (N) and sulfur (S) content was analysed by elemental analyser using flash combustion (Elementar, vario PyroCube, Germany). The stable isotopes 13C, 15N and 34S were analysed by isotope Ratio Mass Spectrometry (IRMS, Isoprime100). A duplicate sample of 35 mg was weighed into a silver cup for organic C measurement. Fifty µL of MQW was added into this cup and the samples were fumigated with concentrated HCl within a desiccator for 24h (Komada et al., 2008) to remove inorganic C. Samples were finally dried in an oven at 60°C and analysed. Isotopic results are reported as delta values (δX; where X = 13C,15N or 34S): δX =(R_sample / R_standard -1)×1000 ‰ where R is the ratio 13C/12C, 15N/14N or 34S/32S respectively. The δ13C value is reported respective to the PDB (Pee Dee Belemnite) standard; the δ15N is reported with reference to air; and δ34S is reported respective to the CTD (Canyon Diablo troilite) standard. References - Armand, L. K., O’Brien, P. E., Armbrecht, L., Baker, H., Caburlotto, A., Connell, T., … Young, A. (2018). Interactions of the Totten Glacier with the Southern Ocean through multiple glacial cycles (IN2017-V01): Post-survey report. ANU Research Publications. - Komada, T., Anderson, M. R., and Dorfmeier, C. L. (2008). Carbonate removal from coastal sediments for the determination of organic carbon and its isotopic signatures, δ 13 C and Δ 14 C: comparison of fumigation and direct acidification by hydrochloric acid . Limnology and Oceanography: Methods, 6(6), 254–262.

The absolute abundances (cells per ml) of 22 hard-shelled phytoplankon taxa (comprised of species, genera or higher taxonomic groups) estimated from Scanning Electron Microscope survey of 52 samples collected through 11 austral spring-summers (2002/3 to 2012/13) (part of the L' Astrolabe collection) from the seasonal ice zone of the Southern Ocean (between latitude 62 and 64.4 degrees south, and longitude 135.8 and 150 degrees east) also included are environmental covariables for each sample: three constructed SAM indices, SST, Salinity, NOx, PO4, SiO4, and the sampling date, time, and location. Fifty-two surface-water samples were collected from the seasonal ice zone (SIZ) of the Southern Ocean (SO) across 11 consecutive austral spring-summers from 2002/03 to 2012/13. The samples were collected aboard the French re-supply vessel MV L’Astrolabe during resupply voyages between Hobart, Tasmania, and Dumont d’Urville, Antarctica between the 20th October and the 1st March. Most samples were collected from ice-free water, although some were collected south of the receding ice-edge. The sampled area was in the high latitude SO (Figure 1b) in the south-east corner of the Australian Antarctic Basin, spanning 270 km of latitude between 62°S and 64.5°S, and 625km of longitude between 136°E and 148°E. The area lies greater than 100 km north of the Antarctic continental shelf, in waters greater than 3,000 m depth. Samples were obtained from the clean seawater line of the re-supply ship from around 3 m depth. Each sample represented 250 ml of seawater filtered through a 25 mm diameter polycarbonate-membrane filter with 0.8 µm pores (Poretics). The filter was then rinsed with two additions of approximately 2 ml of MilliQ water to remove salt, then air dried and stored in a sealed container containing silica gel desiccant. Samples were prepared for scanning electron microscope (SEM) survey by mounting each filter onto metal stubs and sputter coating with 15 nm gold or platinum. Only organisms possessing hard siliceous or calcareous shells were sufficiently well preserved through the sample preparation technique that they could be identified by SEM, and included diatoms, coccolithophores, silicoflagellates, Pterosperma, parmales, radiolarians, and armoured dinoflagellates. The composition and abundance the phytoplankton community of each sample was determined using a JEOL JSM 840 Field Emission SEM. Cell numbers for each phytoplankton taxon were counted in randomly selected digital images of SEM fields taken at x400 magnification (Figure 2). Each image represented an area of 301 x 227 µm (0.068 mm2) of each sample filter, which was captured at a resolution 8.5 pixels/µm. A minimum of three SEM fields were assessed for each sample, with more fields assessed when cell densities were lower. On average, 387 cells were counted for each sample. Taxa were classified with the aid of Scott and Marchant (2005), Tomas (1997), and expert opinion. Cell counts per image were converted to volume-specific abundances (cells/ml) by dividing by 0.0348 ml of sea-water represented by each image. A total of 19,943 phytoplankton organisms were identified and counted: 18,872 diatoms, 322 Parmales, 173 coccolithophores, 81 silicoflagellates, and 45 Petasaria. A total of 48 phytoplankton taxa were identified, many to species level. Because the diatoms Fragilariopsis curta (Van Heurck) Hustedt and F. cylindrus (Grunow ex Cleve) Helmcke and Krieger could not be reliably discriminated at the microscope resolution employed, they were pooled into a single taxa-group. Other taxa were also grouped, namely Nitzschia acicularis (Kützing) W.Smith with N. decipiens Hustedt to a single group, and discoid centric diatoms of the genera Thalassiosira, Actinocyclus and Porosira to another. Rare species, with maximum relative abundance less than 2%, were removed from the data prior to analysis as they were not considered to be sufficiently abundant to warrant further analysis (Webb and Bryson 1972, Taylor and Sjunneskog 2002, Swilo et al. 2016). After pooling taxa and deleting rare taxa, twenty-two taxa and taxonomic-groups (species, groups of species and families) remained to describe the composition of the phytoplankton community. Phytoplankton abundances were related to a range of environmental covariates available at the time of sampling. These included the SAM, sea surface temperature (SST), salinity, time since sea ice cover (DaysSinceSeaIce, defined below), minimum latitude of sea ice in the preceding winter, latitude and longitude of sample collection, the days since 1st October that a sample was collected (DaysAfter1Oct), the year of sampling (year, being the year that each spring-summer sampling season began), the time of day that a sample was collected, and macro-nutrient concentrations: phosphate (PO4), silicate (SiO4) and nitrate + nitrite (hereafter nitrate, NOx). Water samples for dissolved macro-nutrients were collected, frozen on ship, and later analysed at CSIRO in Hobart using standard spectrophotometric methods (Hydes et al. 2010). Daily estimates of SAM were obtained from the US NWS Climate Prediction Center's website and are the NOAA Antarctic Oscillation Index values based on 700-hPa geopotential height anomalies (NOAA 2017). The variable DaysSinceSeaIce was defined as the time since sea ice had melted to 20% cover (after Wright et al. 2010) as determined from daily Special Sensor Microwave/Imager (SSM/I) sea ice concentration data distributed by the University of Hamburg (Spreen et al. 2008). To examine the lag in the expression of the SAM on phytoplankton community composition, two response surfaces were constructed relating the variance in phytoplankton community composition explained by the SAM to the temporal positioning of the period over which daily SAM was averaged. These were derived by evaluating separate CAP analyses (described below) based on daily SAM averaged across a range of days {1, 3, 5, … 365} centred on (i) each calendar day individually (1 Jan – 31 Dec) through the year associated with each sample; and (ii) lagged from 1 to 365 days prior to each sample collection date. Empirical identification of the time between variation in the SAM and the manifestation of this variation in the phytoplankton community structure revealed three modes (maxima) in phytoplankton community composition explained by the SAM. The first was an autumn seasonal SAM mode, which was determined to be the average of 57 daily SAM estimates centred on the preceding 11th March (11th Feb – 8th Apr). This mode explained up to 13.3% of the variance in taxonomic composition (SAM autumn). The second was a spring seasonal mode, which was determined to be the average of 75 daily SAM estimates centred on 25th October (20th Sep – 3rd Dec). This mode explained up to 10.3% of variance in taxonomic composition (SAM spring). Unlike the other modes that were related to the time of year, the third mode was timed relative to the date of sample collection for each sample and comprised the average of the 97 daily SAM estimates centred 102 days prior to each sample collection date. It explained 9.9% of the variance in phytoplankton composition (SAM prior). The mean standard error on estimates of SAM were 0.14 SAM index units for SAM autumn and SAM spring, and 0.13 for SAM prior. Note that SAM prior and SAM spring temporally overlapped to varying extents across the 52 samples and so were not entirely independent covariates: for example, a sample collected in the summer had previous days contributing to both SAM prior and SAM spring.

Midwater fish nitrogen isotope data from the Kerguelen Axis ecosystem study (AAS_4344): These data are based on samples collected as part of the Kerguelen Axis marine ecosystem study (AAS_4344), chief scientist Andrew Constable. This research was supported by the Australian government under the (i) Cooperative Research Centre Program through the Antarctic Climate and Ecosystems Cooperative Research Centre (ACE CRC), (ii) Australian Antarctic Science Program (Projects 4343, 4344, 4347 and 4366), and (iii) Australian Research Council’s Special Research Initiative for Antarctic Gateway Partnership (Project ID SR140300001). The preferred citation is: Walters et al. Food sources and trophic structure of deep sea midwater (fish) food webs in the Southern Ocean as inferred from nitrogen isotopic compositions. Midwater fish samples were collected on board the R.S.V. Aurora Australis during the austral summer of 2016 (22 January-17 February) as part of the Kerguelen Axis marine ecosystem study (AAS_4344). Samples were collected from 9 sampling stations along one transect from the Antarctic continental shelf to the BANZARE Bank over the Kerguelen Plateau. Midwater fish were sampled from the surface to 1000 m depth using an IYGPT (International Young Gadoid Pelagic Trawl) net equipped with a MIDOC (Mid-water Open Close) multiple cod-end device. Analyses focused on mesopelagic and bathypelagic fish taxa. The nitrogen isotopic composition of individual amino acids was measured in muscle tissue from each fish. As of 2022-09-30, these data were still being worked up for publication.

Water samples were collected from the seawater line on the Aurora Australis during the K-Axis voyage. They were filtered so that two fractions of each sample were collected: a fraction that was between 1.2 and 210 um and a fraction that was between 210 and 1000 um. A 47 mm diameter 1000 um mesh was placed upstream of all samples, and this prevented larger particles (e.g. zooplankton) from entering the samples. The underway water was taken from the pCO2 rig at 1.4 to 1.5 atmospheres. All samples were collected on 25 mm diameter 1.2um Sterlitech silver membrane filters. The greater than 210 samples were collected on mesh and refiltered onto silver filters. The filters were stored frozen until they were processed in Hobart. Subsamples of the filters were analysed at the Central Science Laboratories, University of Tasmania to determine elemental N and C. The remainder of the filters were analysed by ANSTO (NSW) to determine delta15N and delta13C. Volumes are in litres, and the values for the nitrogen isotopes are presented as ratios.

Krill, salps and pteropods were collected with an RMT8 net during the K-Axis cruise. Specimens were removed from the samples, measured and frozen at -20C until ready for analysis in Hobart. Individuals of known species were dried at -60C, ground to a fine powder, encapsulated into tin cups and analysed with an ICP-MS in the Central Science Laboratories, University of Tasmania. Samples were analysed for delta15N and delta13C. The salp was the common Southern Ocean species Salpa thompsoni and the krill were Euphausia superba, E. triacantha, E. frigida and Thysanoessa macrura. A small number (2) of the siphonphore Diphyes antarctica were also analysed. Pteropods analysed included both shelled (thecosomes) and naked (gymnosomes) pteropods. Columns E-O in the Pteropods worksheet in the spreadsheet are expressed as ratios.

Five (out of a possible 7) ice stations were sampled for the Main Biology Site, collected from -63.88S 119.9E off East Antarctic in September to November 2012 during the Sea Ice Physics and Ecosystems eXperiment (SIPEX) II. Sampled pack ice floes were several 100 meters to several kilometres apart, and indicated variation in the degrees of physical deformation and biological characteristics. The sampled sites were selected on each floe due to low snow cover disturbances, were level, and free from surface deformations (limited rafting). For the production and carbon allocation dataset, the bottom 2 cm of 3 x 5 - 8 (dependant on level of biomass) cores were collected and combined within a 4 m grid using a 9 cm diameter SIPRE corer. Biology was isolated from the ice by gently mixing with 0.22 micron filtered sea water collected from the site with a Niskin bottle, and pass through a sieve. The liquid was then analysed for bacterial and algae productivity. The corresponding dataset (P:\Data\ Copy of SIPEX C14_Ugalde_Raw Data_St 8) describes data (expressed in disintegrations per minute) directly input into an excel file from the scintillation counter measured on board the Aurora Australis. The additional dataset (P:\Data\ SIPEX C14_Chloro_SIPEXII_Updated to Station 8) describes data directly input into an excel file of volume filtered for chlorophyll analysis (expressed in mls) of both the ice and liquid fraction. For the Main Biology dataset, 6 cores were taken from the same site: Core 1: temperature profile Core 2: nutrients, extracellular polymeric substances Core 3 and 4: chlorophyll, pigments (HPLC), bacteria and cell counts Core 5: particulate organic carbon/nitrogen, dissolved organic carbon Each core was sectioned from the ice-water interface at 0 - 2 cm, 2 - 10 cm, and then the remaining core was quartered. Core 1 was discarded after the temperature profile was taken. Core 3 and 4 were slow-melted in at 6 degrees cold room with 0.22 micron filtered sea water collected from the site as above (200 ml per cm ice). Cores 2 and 5 were slow-melted as above without the addition of filtered sea water. Additional measurements included 5 replicates of snow thickness and freeboard level. After melting, samples were taken/filtered for the parameters above within 12 hours of melting. The corresponding dataset (P:\Data\Data Updated to Station 8\ Main Bio Data Sheet_SIPEX II_Updated to Station 8) describes descriptive and quantitative parameters of the above cores, directly input into a spreadsheet.

3 litres of seawater were collected every 2nd CTD (conductivity, temperature and depth) cast on every CTD transect of the BROKE-West voyage. 7 CTD transects were completed on the BROKE-West voyage, all on southwards legs. Samples were collected at 6 depths in the top 200 m of the water column using niskin bottles. 2 litres were filtered through polycarbonate filters and 1 litre was filtered through a fibreglass filter. Chemical digestion of the polycarbonate filter enabled us to determine the particulate silicon concentration for each sample (using the nutrient autoanalyser onboard the Aurora Australis, see hydrochemistry section), fibreglass filters have been dried and stored for CHN analysis back on shore. This work was completed as part of ASAC projects 2655 and 2679 (ASAC_2655, ASAC_2679).