Surface water samples for these experiments were collected from West Bay of the Neuse River Estuary, North Carolina USA (34°55'32.42" N, 76°21'54.25" W) and distributed into six microcosms. The microcosms were 60 liters. The experiments were conducted in biological duplicates under three light treatment incubations: 12-hour light-dark cycle of photosynthetically active radiation (PAR), 12-hour light-dark cycle of UV-B radiation, or darkness. For each 60-liter microcosm, water was sampled daily to weekly at the University of North Carolina over the month-long (September 2021) experiment or two-month-long experiment (April 2022). Water was sampled from the microcosms using a peristaltic pump (Masterflex) under gentle (75%) pressure through in-line 3 micron (um) and 0.2 micron, 47-millimeter (mm) polycarbonate filters (Millipore Sigma) which were flushed with 250 milliliters (mL) of Milli-Q water. All tubing, filter holders, and 60-milliter (mL) HDPE collection bottles (Fisher Scientific) were acid washed in 10% (v/v) HCl for six hours and triple rinsed with Milli-Q water. <0.2 um filtrate was collected from each tank for dissolved inorganic nutrient analysis and stored for one month at -20 degrees Celisus (°C). Acid-washed glassware was used for NOx and NH4 assays, and RBS phosphate-free detergent washed glassware was used for SRP assay.
NOx was quantified using 2% vanadium chloride in hydrochloric acid to reduce nitrate to nitrite, then using the Greiss reaction on 1 mL sample volumes following Garcia-Robledo et al. (2014). 10 standard solutions ranging from 0.1 to 10 micromolar (uM) were created in saltwater (sodium chloride in Milli-Q at in situ salinity).
NH4 was measured using a colorimetric alkalic nitroprusside reaction in 96-well plates as described in Bower and Holm-Hanson, 1980. 10 standard solutions of NH4 were created ranging from 0.5 to 25 uM using saltwater similarly to NOx standards. For summer samples, NOx and NH4 absorbance were measured with quartz cuvettes at 540 nanometers (nm) (NOx) and 630 nm (NH4). Spring samples were measured on a Tecan plate reader.
Soluble reactive phosphorus SRP was measured using a modified molybdenum blue assay from Murphy and Riley (1962). Seven standards were prepared ranging from 0.1 to 4 uM in saltwater similar to NOx and NH4. All measurements were quantified with quartz cuvettes at 885 nm.