Dataset: Alkaline phosphatase activity over bioassay experiments with seawater from R/V Savannah cruise SAV-19-02 in the NW Atlantic Ocean in Spring of 2019

Release Date:2022-07-26Data not availableVersion 1 (2021-11-02)Dataset Type:Cruise ResultsDataset Type:experimental

Principal Investigator: Solange Duhamel (Columbia University)

Co-Principal Investigator: Julia Diaz (University of Georgia)

Contact: Kahina Djaoudi (University of Arizona)

BCO-DMO Data Manager: Amber D. York (Woods Hole Oceanographic Institution)


Project: Collaborative Research: Assessing the role of compound-specific phosphorus hydrolase transformations in the marine phosphorus cycle (P-hydrolase)


Abstract

Alkaline phosphatase activity over bioassay experiments with seawater collected during R/V Savannah cruise SAV-19-02 from March to April of 2019 in the Northwestern Atlantic from the surface to 50 m depth.

Sampling and analytical procedures:

 Bioassay Experiments consisted of incubating, over an incubation period of 48h, surface seawater (5m) with inorganic and organic phosphate compounds (20 µM; final concentration of P) including, polyphosphate (polyp), inorganic phosphate (Pi), nucleotides (ATP, AMP) and phosphonate (Mepn). In each incubation experiment, a control treatment (surface seawater) was considered as well in each bioassay experiment alongside a treatment amended with nitrogen (NH4Cl, NaNO3). These bioassay experiments were conducted at station 1 and stations 3. At each station, inorganic and organic phosphate amendments were performed on seawater with and without nitrogen enrichment.

Alkaline phosphatase activity  was determined fluorometrically using 4-methylumbelliferyl phosphate (MUF-P), as representative of phosphomonoesterase activity. Total APA was measured in each experimental bottles in triplicate. Hydrolysis rates were determined by separately incubating 200 µL of seawater with 8 different concentrations (0, 0.1, 0.2, 0.5, 1, 5, 10, 20 µM; final concentrations) of MUF-P in 96-well black microtiter plates. To ensure linearity of the hydrolysis rate, the increase of fluorescence was measured (excitation/emission wavelength: 359/449 nm) at multiple time points over an incubation period of 24 h.  Maximum hydrolysis rate (Vmax) was determined, using a nonlinear regression based on the rectangular hyperbolic function following V=Vmax x S/ Km+ S, where S and V are the concentrations of the substrate and the hydrolysis rates, respectively.    

Location:  Northwestern Atlantic surface waters. Depth: surface-50 m.

Instruments:  Reading of fluorescence was performed on a plate reader (SpectraMax® M2, Molecular Devices).


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