Determination of Microalgal Lipid Content and Fatty Acid for Biofuel Production
Table 4
Processes of TLC running for the measurement of microalgal lipid content.
Preparation for TLC analysis
TLC analysis
Ref.
Prepare the solvent by shaking and degassing; add solvents to the TLC tank and equilibrate TLC tank.
Add samples to plates; dry the samples and run the plates in hexane : diethyl ether : acetone (60 : 40 : 5, v/v/v); dry plates and run in chloroform : methanol : ammonium hydroxide : water (60 : 35 : 4 : 1, v/v/v/v) to 18 cm; score plates and break apart with hand pressure; determine the line on a different plate or spray only the outside lane of the plate to be cut; rotate the lower portion by 180° and run in chloroform : methanol : acetic acid : water (85 : 12.5 : 12.5 : 3, v/v/v/v) in the original orientation; dry the plates and spray with a solution of primulin dye; visualize lipid spots and scan the plates by laser-excited fluorescent detection; quantify spots and compare to standard curves.
Elute silica gel with chloroform; elute lipids with chloroform to yield NLs; dry NLs and re-suspend in chloroform; activate plates in an oven.
Subject the NL fraction to TLC for lipid class separation and identification; use hexane/diethyl ether/acetic acid (70 : 30 : 1, v/v) for lipid separation; co-chromatography with pure standards; stain bands of lipid classes with 2, 7-dichlorofluorescein; visualize under UV light.
Elute silica gel with chloroform; elute lipids with chloroform to yield NLs; dry NLs and resuspend in chloroform; activate plates in an oven.
Use 80 : 20 : 1 hexane/diethyl ether/acetic acid for lipid separation; spray the plate with a solution of primulin dye; illuminate the plate with UV light.
