Question

1. Predict the elution sequence (rank first to last) of a mixture of the following proteins in a gel filtration column using Sephadex G50 (exclusion size 4*10^4 Da) resin. If some proteins would elute at the same time, give them the same rankings. Indicate which proteins are in the void volume and which will elute in one total column volume.

Lysozyme (14,300 Da), Trypsinogen (24,000 Da), B-galactosidase (116,635 Da), Transferrin (75,190 Da), Leucine aminopeptidase (51,648 Da), myoglobin (17,080 Da)

2. Repeat question 1 for a column packed with Sephadex G200 (exclusion size 5*10^5 Da) resin.

Answer 1

In column chromatography...if we are using a polar stationary phase & a non polar mobile phase...then during solution process, we can collect the most polar component first from the mixture and followed by the next less polar component. And at the end we can collect the least polar component of the absorbed mixture.

If we take non polar stationary phase and polar mobile phase, then initially we could collect the least polar component and followed by the components with increasing order of their polarity.