Lachromelite system

After gentle filtration onto glass fiber filters (<20 mmHg, GF/F), samples were immediately frozen and stored at −80°C until analysis. Pigments were extracted with 90% acetone (v/v) for 24 h at 4°C in the dark. Total pigment concentrations were determined via high‐performance liquid chromatography (LaChromElite^® system, VWR, Darmstadt, Germany) using a Spherisorb ODS‐2 column (5 μm particle size; Waters, Milford, MA, USA) and applying a gradient following Wright et al. (1991). Peaks were identified and quantified via cochromatography of pigment standards (DHI Lab Products, Hørsholm, Denmark) and the software EZChrom Elite ver. 3.1.3. Cellular pigments were separated into the categories light‐harvesting pigments (LHP: chlorophyll a [Chl a], chlorophyll c₂, fucoxanthin) and light‐protective pigments (LPP: diadinoxanthin and diatoxanthin). Pigment contents were normalized to filtered volume and cell densities to yield cellular quotas.

HPLC was carried out on a VWR-Hitachi LaChrom Elite system (pump L-2130, autosampler L-2200, diode array detector L-2455), equipped with a ZORBAX Eclipse Plus column (C18, 4.6 × 100 mm, 3.5 μ; Agilent). The solvents used were 0.1% formic acid in water (A) and acetonitrile (B). The gradients were as follows. Incubations with pentahydroxybenzophenone and tetrahydroxyxanthones: 4 min 5% B, 12 min 20% B, 25 min 27% B and 28 min 100% B; incubations with trihydroxybenzophenone, tetrahydroxybenzophenone and trihydroxyxanthones: 4 min 5% B, 12 min 30% B, 25 min 34% B and 28 min 100% B; and incubations with mono- and dihydroxybenzophenones and dihydroxyxanthones: 4 min 5% B, 10 min 30% B, 22 min 40% B and 26 min 100% B. The flow rate was 1 ml min⁻¹ and the detection wavelength was 260 nm.