Trilogy fluorometer

Chla_in-vitro were determined 4 months after collection. For a fluorometric-based analyses, Chl a was extracted in 100% methanol after 2 hours placed on a dark fridge at -10°C, and determined using the Turner Designs Trilogy fluorometer (model: 7200–000) and the non-acidification method [36 (link)].
In-situ surface (<10 m) FChla_AUV and FChla_FRRf are often spurious. Such measurements are not always representative of phytoplankton biomass concentrations because of a phenomenon named non-photochemical quenching (NPQ). NPQ is a photo-physiological response of live cells to high light levels, where the excess of energy is dissipated as heat, instead of being used for photochemistry [37 (link)]. Thus, a reduction (quenching) of the FChla_AUV/FChla_FRRf signal is often induced by high light conditions, particularly at surface waters during daytime hours [38 (link)]. To cross-check for the presence or absence of NPQ, Chla_in-vitro measurements from discrete water samples (1, 10, 20, 40 and 60 m) were taken to validate the data retrieved from sensors.

A subsurface CF maximum could be biased by non-photochemical quenching (NPQ) [45 (link)], exhibiting as a subsurface peak, by quenching the signal close to the surface rather than representing the true distribution of chlorophyll a in the water column. We compared CF predictions from in situ AUV data and shipborne CF measurements with the in vitro concentrations of chlorophyll a from water samples. To validate the data retrieved from sensors, discrete water samples for measurement of in vitro chlorophyll a were collected using 10 L Niskin bottles [46 ] on board the work boat. Sample depths were selected based on layers of relatively high, medium and low chlorophyll a concentrations estimated by the model generated in the operator hub from data from the AUVs. Seawater was filtered (0.5 L) onto Whatman GF/F glass fiber filters [5 (link)]. In the laboratory, chlorophyll a was extracted in 100% methanol after 20 hours at −20°C in darkness. The chlorophyll a concentrations were determined using a Turner Designs Trilogy fluorometer (model: 7200–000) following the non-acidification method [47 (link)].

Samples for chlorophyll-a analysis (~100 mL) were filtered onto 0.7 µm nominal porosity GF/F filters and extracted into 90% acetone for 24 h in the dark at 4 °C prior to measurement. Chlorophyll-a concentration was determined on a precalibrated Turner Designs Trilogy fluorometer (66 ) while at sea within 48 h of collection.

Seawater samples were collected on seven research cruises aboard the Canadian Coast Guard Ship (CCGS) Sir Wilfrid Laurier annually in July from 2013–2019 at DBO designated stations (Fig 1). DBO1 stations are located southwest of St. Lawrence Island and the DBO2 region is within the Chirikov Basin, which is north of St. Lawrence Island. Water was collected from the CTD rosette (Sea-Bird SBE25/33) using a SBE32 Carousel 12-bottle water sampler with 8-L bottles. Water samples were collected at set depth increments (i.e. 5, 15, 25, 35, 50, 75 m, bottom depth, and at the chlorophyll maximum). Chlorophyll a and pheophytin collection methods followed National Estuarine Research Reserve System Centralized Data Management Office protocols [36 ]. Briefly, seawater samples (200 mL) were immediately filtered on 25 mm Whatman GF/F filters in the dark. Filters were frozen shipboard (-20°C) and analyzed within three months post-cruise at Clark University. Chlorophyll a was extracted from filters using 90% acetone, and vials with acetone and filters were stored wrapped in foil in a freezer for at least 48 hours prior to measurement on a calibrated Trilogy Fluorometer (Turner Designs, San Jose, California). Pheophytin was determined following acidification of the samples. Cruise data are available at https://arcticdata.io/catalog/portals/DBO/Data.

Chlorophyll a, as an indicator of phytoplankton biomass, was measured on subsampled samples by filtering 50 mL of sample water onto Whatman glass fiber filters (GF/F). Filters were frozen at −20 °C and subsequently extracted using a tissue grinder in 90% acetone [107 ,108 ]. Chlorophyll a in extracts was measured using the non-acidification method of Welschmeyer [109 (link)] on a Turner Designs Trilogy fluorometer calibrated with pure Chlorophyll a standards (Turner Designs, Sunnyvale, CA, USA).

For the determination of chlorophyll‐a, the methodology of Jeffrey and Humphrey (1975 (link)) was followed. Pellets were resuspended in 2 mL of 90% acetone and vortexed. The samples were then stored in the dark at 4°C for 24 h. After this, samples were centrifuged for 5 min, and 2 mL were transferred into two glass cuvettes (1 mL in each), for two replicate readings. Samples were individually measured at two fixed wavelengths of 663 and 630 nm using a Trilogy Fluorometer (Turner Designs) fitted with a chlorophyll‐a module. Each sample was measured three times. All analyses were performed in a dark room. Measurements were normalised to the initial volume of the sample and to the number of algal cells per colony.