Spectrofluorometric quantification of neutral and polar lipids in zooplankton using Nile red

A high throughput Nile red method for quantitative measurement of neutral lipids in microalgae

“…Little information is available regarding determination of the absolute content of neutral lipids in algae using the conventional gravimetric method calibrated with a lipid standard (Alonzo and Mayzaud, 1999). Our results indicated no significant difference between the two methods based on the mean values, whereas, an absolute error (7%) and a relative standard error (15%) between the two methods in neutral lipid content from the same algal biomass was observed.…”

“…These wavelengths were different from the wavelengths 480 nm/ 580 nm used by several investigators (Genicot et al, 2005;Elsey et al, 2007), but similar to that reported by Alonzo and Mayzaud (1999), in which 530/580 nm was selected for the neutral lipid determination in zooplankton. The excitation and emission wavelengths for total lipid measurement ranged from 470 to 549 nm and from 540 to 628 nm, respectively, depending on the individual organisms tested, and the composition and content of intracellular lipids.…”

“…Therefore, prior to fluorescence measurement, optimal excitation and emission wavelengths for maximum fluorescence yield must be identified. Given that many classes of algae, particularly the green algae taxa (Chlorophyceae) contain large amounts of chlorophyll (1-4% of dry weight) which can considerably increase the fluorescence background, making it difficult, if not impossible, to use the Nile red method to quantify cellular polar lipids using the wavelengths of 560/620 nm, which were previously used to determine neutral and polar lipids in non-or low chlorophyll-containing cells, tissues, and organisms (e.g., mammalian cells, zooplankton, yeast, diatoms and chrysophycean species) (Cole, 1990;Alonzo and Mayzaud, 1999;de la Jara et al, 2003).…”

“…The extract was evaporated in a water bath (30°C) using a rotary evaporator (Büchi, Switzerland) to remove solvents. Crude lipids were further separated by column chromatography using silicon gel (60-200 mesh) (Merck Corp., Germany) according to Alonzo and Mayzaud (1999) using the following solvent sequence: 6 volumes of chloroform to collect the neutral lipid class and 6 volumes of methanol to collect the polar lipids. Each lipid fraction was transferred into a pre-weighed vial, initially evaporated in a water bath (30°C) using a rotary evaporator and then dried under high vacuum.…”

Effect of Salinity and pH on Fatty Acid Profile of The Green Algae Tetraselmis suecica

“…Nile red sample measurement test: Nile Red measurements on algal cells grown under different environmental conditions were done after performing optimisation test for Nile Red peak fluorescence and Nile Red concentration test based on the work done by Alonzo, Chen and Bertozzini [15][16][17]. Also, to find the optimise time for harvesting cells during the induction of lipid accumulation, measurements were performed on cells grown for 1 week, 2 weeks, 3 weeks and 4 weeks.…”