Dataset: SO248: GF FLA hydrolysis rates
Deployment: SO248

Water was collected via Niskin bottles mounted on a rosette, equipped with a CTD.

Experiments on (operationally defined) particles were carried out by gravity-filtering water through 3 um pore size filters.  1/12th sections of the 3 um pore-size filters were submerged in 15 mL artificial seawater; enzyme activities were measured as described below.

The potential of the seawater microbial community associated with large particles (> 3 micrometer pore size filter) to hydrolyze six high-molecular-weight polysaccharides (arabinogalactan, chondroitin sulfate, fucoidan, laminarin, pullulan, and xylan) was investigated in surface and bottom water. For each substrate, three  15 mL falcon tubes were filled with autoclaved artificial seawater and one 15 mL falcon tube was filled with autoclaved seawater and a blank filter piece to serve as a killed control. Substrate was added at 3.5 uM monomer-equivalent concentrations, except for fucoidan, which was added at 5 uM concentrations (a higher concentration was necessary for sufficient fluorescence signal). Two 15 mL falcon tubes – one with seawater and one with autoclaved seawater – with no added substrate served as blank controls. Incubations were stored in the dark at as close to in situ temperature as possible. Subsamples of the incubations were collected at time zero, and at six subsequent timepoints. At each timepoint, 2 mL of seawater was collected from the 50 mL falcon tube using a sterile syringe, filtered through a 0.2 um pore size syringe filter, and stored frozen until processing.

The hydrolysis of high molecular weight substrate to lower molecular weight hydrolysis products was measured using gel permeation chromatography with fluorescence detection, after the method of Arnosti [1996, 2003]. In short, the subsample was injected onto a series of columns consisting of a 21 cm column of G50 and a 19 cm column of G75 Sephadex gel. The fluorescence of the column effluent was measured at excitation and emission wavelengths of 490 and 530 nm, respectively. Hydrolysis rates were calculated from the change in molecular weight distribution of the substrate over time, as described in detail in Arnosti [2003].

ara = arabinogalactan
chn = chondroitin sulfate
fuc = fucoidan
lam = laminarin 
pul = pullulan
xyl = xylan