Computational protocol: Fatty acid profiles of the main lipid classes of green seaweeds from fish pond aquaculture

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Protocol publication

[…] The main lipid classes were separated by analytical thin‐layer chromatography (TLC) in plates coated with 0.25 mm silica gel G and developed with a mixture of hexane:diethylether:acetic acid (50:50:2 by volume), based on the method described by Bandarra, Batista, Nunes, Empis, & Christie (). Extracted lipids were dissolved in chloroforom (10 mg/ml concentration). A mixture of standards (sigma chemical Co., St. Louis, Mo) was also prepared in chloroform with the same concentration. Specifically, glyceryltrioleate (TAG), glyceryl 1,3‐dipalmitate (diacylglycerol, DAG), DL‐α‐monoolein (MAG), oleic acid (FFA), L‐α‐phosphatidylcholine (PL), and monogalactosyl diacylglycerol (GL) were used. The samples and standards (10 μl) were applied to the plates and each plate was immersed in 102 ml of the elution mixture inside a developing chamber. The elution front was followed visually. After elution front reached the upper limit, plates were taken out from the chamber. The developed plates were then sprayed with 10% phosphomolybdic acid in ethanol (w/v). Identification of lipid classes (polar and apolar) was done by comparison with standards. Quantification was performed using a scanner and version 4.5.2 of Quantity One 1‐D Analysis software from Bio‐rad (Hercules, CA). There were always two replicates. [...] In order to test normality and variance homogeneity, the Kolmogorov‐Smirnov's test and Levene's F‐test, respectively, were used. Data fulfilled both of these parametric tests’ assumptions. The seaweed species (C. linum, R. riparium, U. intestinalis, U. lactuca, and U. prolifera) and the contrast between different lipid classes (TAG, PL+GL, MAG, and FFA) were the two studied factors. All statistically analyzed values were in percentage (% of total FA in the total FA comparison; % of total fat in lipid class distribution; and % of the FAs in each class in the respective lipid class). The parametric test, Tukey HSD, was done with STATISTICA 6, 2003 version (StatSoft, Inc., Tulsa, OK). For all statistical tests significance level (α) was 0.05. Whenever p was lower than α, statistical differences between species (total FA comparison; class distribution; specific class FA comparison) or between lipid classes for the same species (specific class FA comparison) were identified. In the former situation, lowercase letters were used, while, in the latter situation, uppercase letters were used. […]

Pipeline specifications

Software tools Quantity One 1-D, Statistica
Applications Miscellaneous, DNA fingerprinting
Organisms Danio rerio, Ulva prolifera
Chemicals Fatty Acids, Nonesterified, Triglycerides, Palmitic Acid, Monoglycerides