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BCH 2333 Introduction to Biochemistry Lab 4 Corrected Lipids and Membranes
Two membrane fractions were prepared from a rat liver tissue: a microsomal enriched fraction and a mitochondrial enriched fraction. The lipids were extracted from the microsomal fraction, separated using 2D thin layer chromatography and were then compared with lipids extracted from the entire tissue. The 2D TLC plate is composed of mildly polar silica gel, which will bind polar compounds more tightly than non-polar ones. The first dimension of the plate was developed in a basic solvent and the second dimension was done in a basic solvent. The purpose of these 2 dimensions was to be able visualize the various polarities of the lipids and how they react on the silica gel in varying pH environments. In the second part of the experiment, the lipid/protein ratio in the mitochondrial fraction was analysed. Brilliant Blue Dye R was used to estimate the lipid concentration of the fraction and an absorbance assay was used to estimate the protein concentration.
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