Uptake Incubations:
Fe uptake incubations were performed on 12 Hawaii Ocean Time-series (HOT) cruises, onboard the R/V Kilo Moana throughout 2021-2023. All samples were collected at Station ALOHA (22°45' N, 158°0' W) following trace metal clean procedures. Incubations were performed and harvested following the method described in Hawco et al. (2022), with minor modifications. See the attached supplemental file for a listing of specific cruise IDs.

For all cruises except October 2021, trace metal clean seawater was collected using a powder-coated aluminum framed 12-place 'trace metal' rosette (Seabird) with 8-liter (L) external spring sampling bottles (Ocean Test Equipment), deployed using metal-free line (Amsteel). For the October 2021 cruise, samples were collected with C-Free bottles (Ocean Test Equipment) that were attached directly to the metal-free line and triggered with Teflon-coated messengers. Prior to collection, sampling bottles were subjected to a ca. 18 hour soak in seawater collected at a test station near Oahu. Prior to each cruise, all materials were soaked in Citranox for one day, soaked in 10% HCl for 7 days, and rinsed multiple times with 18.2 M-Ohm Milli-Q water (Millipore).

To condition the Fe spike at low nanomolar (nM) concentrations in seawater, seawater was collected at 25 meters (m) to "pre-incubate" the isotope spike with natural iron binding ligands present in seawater. Upon recovery, sampling bottles were taken to a positive pressure clean van onboard. For each "pre-incubation", 100 milliliters (mL) of seawater was filtered through a 0.2 micrometer (um) Acropak filter into an acid-cleaned 150 mL PC bottle. The seawater was amended with 100 microliters (uL) of a ~1 micromolar (uM) Fe double spike (Fe_DS) containing 57Fe and 58Fe and left in the dark at room temperature for approximately 24 hours.

Seawater for the incubations was collected after sunset from the mixed layer (25 m). The incubations were set up in the clean van under red light to minimize light shock. The 100 mL of pre-equilibrated solution was added to a 2 L acid-cleaned polycarbonate bottle, pre-rinsed with filtered seawater, which was then filled with seawater. The incubations were performed in triplicate, alongside a control incubation that was filtered at 0.2 um. Incubation bottles were placed in an on-deck flow-through incubator, shaded to light levels equivalent to a depth of 25 m and temperatures matching the surface mixed layer. At 24 hours, the incubations were removed and harvested via vacuum filtration onto acid-cleaned 0.2 um polyethersulfone filters (47 mm), with the filtrate collected in 1 L bottles.

Sample Purification and Measurement:
Filtrate from the incubations was acidified to pH 1.8 using Optima grade hydrochloric acid and left to equilibrate for a minimum of 2 months.

Laboratory processing of samples followed methods described in Hawco et al. (2022). Briefly, filters were digested overnight in 50% nitric acid (optima grade) in perfluoroalkoxyl vials (Savillex) at 95 degrees Celsius. Dissolved iron in acidified filtrates was extracted using Nobias PA-1 resin after adjusting sample pH to ~6, rinsed with water and eluted in 1M nitric acid. Both sample types were then evaporated to dryness, heated in a 1:1 HNO₃:HCl solution for 2 hours, dried down again, and then resuspended in 10 M HCl for column chemistry purification of Fe with AG-MP1 resin. Sample was loaded at 10 M HCl, rinsed at 5M HCl, and then Fe was eluted at 1 M HCl. Following resin extraction, samples were again dried down and resuspended in 0.1 M HNO₃ for MC-ICP-MS analysis.

Measurements were performed using a Thermo Scientific Neptune MC-ICP-MS at the University of Southern California, using the IRMM-014 standard to define the natural abundance ratio of Fe.