3.3 Incubation design and sampling
Incubation regeneration experiments were carried out in the subantarctic
Southern Ocean south of Tasmania (IN2018_V02) in acid-cleaned 20 L LDPE
cubitainers with Teflon-lined caps. Three regeneration experiments were
conducted, with differences in incubation temperature and the particle
collection depth (supplemental material). Each experiment consisted of
two treatments, each in triplicate. Cubitainers were filled with
filtered (0.2 µm) surface (~5 m) seawater collected
using a trace metal-clean tow fish, and aliquots were collected for
initial (T0) nutrients, [Cr] and
δ53Cr before particle inoculation. Filled cubitainers
were stored in the dark at 1° C until inoculating with particles (within
8 hours of filling). Ambient suspended particle samples from the oxic
water column were collected onto acid-cleaned 1 µm polycarbonate filters
using in-situ pumps (McLane). Particles were resuspended from these
filters, forming a particle-rich slurry that was added to the individual
cubitainers to result in particulate concentrations approximately 2-4
fold above natural levels. Because only resuspended particles were added
to the incubaitons, the high Cr blanks associated with many filter
materials (e.g. Scheiderich et al., 2015) will not impact the
incubations.
Particle-inoculated cubitainers were incubated in the dark at either 4°
C (ambient) or 1° C (cold treatment) for five days, with three
subsampling time points. For subsampling, the cubitainers were
transported to a Class 100 clean van, gently mixed, and subsampled into
acid-cleaned LDPE bottles (for trace metals) or polycarbonate tubes (for
macronutrients). Subsamples for trace metals were transported to a
laminar flow hood and filtered through acid-cleaned 0.4 µm polycarbonate
filters using a Teflon vacuum filtration apparatus (Savillex). Loss of
Cr to container walls has been shown to be minimal over short timeframes
(e.g. Semeniuk et al., 2016) and was not considered here.
Chemically-labile particulate metal and P concentrations were determined
with magnetic-sector ICPMS following solubilisation with a heated
mixture of reductant (hydroxylamine hydrochloride) and weak acid (acetic
acid) designed to dissolve biogenic material and Mn and Fe oxides
(Berger et al. 2008). Analytical procedures followed those previously
described in Rauschenberg and Twining (2015).