Water samples were taken monthly from August 2022 - December 2023. One water sample was taken per reef. The samples were analyzed for phosphate (µmol/L) and nitrate + nitrite (µmol/L). Water samples were analyzed by the University of Hawaiʻi at Mānoa SOEST Laboratory for Analytical Biogeochemistry. They were analyzed for: Total Nitrogen, Total Phosphorous, Phosphate, Silicate, Nitrate + Nirite, Ammonia, and Chlorophyll using a Seal Analytical AA3 HR Nutrient Autoanalyzer.

Sample collection: Water samples were collected in sterile 60ml plastic bottles, each rinsed three times from the sample area before a sample was taken.

Seal Analytical AA3 HR Nutrient Autoanalyzer:
This instrument is a fully automated/computerized analyzing system for nutrients in environmental waters. It is a five-channel segmented-flow continuous analyzer consisting of a sampler, a pump, five-reagent mixing and reaction manifolds and five photometers. S-LAB utilizes the AA3 to simultaneously measure dissolved inorganic nitrate, nitrite, ammonium, phosphate, and silicate at low μM levels. Detection limits offered by this state-of-the-art instrument fulfill the detection needs of researchers wishing to analyze oligotrophic (low nutrient concentration) seawater, while also providing the capability of analyzing fresh and brackish waters, including soil water, sediment pore water, and groundwater. (Description from https://www.soest.hawaii.edu/S-LAB/equipment/slab_autoanalyzer.htm

The following describes the measurements in more detail.
* excerpt from University of Hawaiʻi at Mānoa SOEST Laboratory for Analytical Biogeochemistry's "Seal Analytical AA3 HR Nutrient Autoanalyzer" page https://www.soest.hawaii.edu/S-LAB/equipment/slab_autoanalyzer.htm)

Ammonium
Ammonium is measured fluorometrically following the method of Kerouel and Aminot (1997). The sample is reacted with o-phthalaldehyde (OPA) at 75°C in the presence of borate buffer and sodium sulfite to form a fluorescent species in a quantity that is proportional to the ammonium concentration. Fluorescence is measured at 460 nm following excitation at 370 nm.

Nitrate and Nitrite
Nitrate and Nitrite are analyzed via the diazo reaction based on the methods of Armstrong et al (1967) and Grasshoff (1983). This automated procedure involves reduction of nitrate to nitrite by a copper-cadmium reductor column. The nitrite then reacts with sulfanilamide under acidic conditions to form a diazo compound, which then couples with N-1-naphthylethylene diamine dihydrochloride to form a purple azo dye. The concentration is determined colorimetrically at 550 nm.

Silicate
Silicate measurement is based on the reduction of silicomolybdate in acidic solution to molybdenum blue by ascorbic acid (Grashoff and Kremling 1983). Oxalic acid is introduced to the sample stream before the addition of ascorbic acid to minimize interference from phosphates. The concentration is determined colorimetrically at 820 nm.

Phosphate
This automated procedure for the determination of orthophosphate is based on the colorimetric method of Murphy and Riley (1962) in which a blue color is formed by the reaction of orthophosphate, molybdate ion and antimony ion followed by reduction with ascorbic acid at a pH of 1. The reduced blue phospho-molybdenum complex is determined colorimetrically at 880 nm.

Total Phosphorus
Following the method developed by the University of Hamburg in co-operation with the Ocean University of Qingdao, this automated procedure for the determination of dissolved phosphorus in seawater takes place in three stages. First, the sample is irradiated in a UV digestor. In this digestion step organically bound phosphorus is released. Second, acid persulfate is added, which further promotes breakdown of orgniac matter that persists after UV digestion, and polyphosphates are converted to ortho-phosphate by acid hydrolysis at 90°C. Third, the ortho-phosphate is determined by reaction with molybdate, antimony and ascorbic acid, producing a phospho-molybdenum blue complex which is determined colorimetricallyat 880 nm.

Total Nitrogen
Following the procedure developed by the University of Hamburg, inorganic and organic nitrogen compounds are oxidized to nitrate by persulfate under alkaline conditions in an on-line UV digestor. The nitrate is reduced to nitrite in a cadmium column and then determined using the sulfanilamide/NEDD reaction with colorimetric detection at 520 nm.