The product will then be frozen dried to obtain the powder form. I have read that it has to be dialysed against distilled water, but I am not so sure.
Re-dissovle with solvent such as distilled water and then freeze dry.
Rotary evaporator is used to concentrate the solution, and not for completely removing the solvent. If you completely remove the solvent, a sticky product will be formed that will be difficult to take out from the flask. Concentrate in the flash evaporator and pour out the concentrate to as beaker or tray, from this concentrate making a pure product depends on the type of solvent that you have used.
Should you have dialyzed it, we hope it shall not diffuse through the membrane, but only the salts. That's is what dialysis is meant for. Other way to purify you "bio-surfactant" is to extract the crude solid left in your rotovap with a non polar solvent (ether) with the aid of ultrasound. Ultrasound disperses well most solid mixtures. You won't find any problem liophilizing the bio-surfactant extracted in ether. Take a good look at the solid left in the rotovap flask, and check with tetrahydrofuran to see if there is another bio-surfactant left. I wouldn't be surprised to find that several surfactants with different polarities are present in the crude product, then you may need different solvents.
I am a bit confused. Which is the best way? Either to evaporate all the solvents until dry and dissolve it with distilled water, then freeze dried? Or to just concentrate it and leaving the concentrate, then freeze dried. Sorry, I really do not know..
Bio-surfactants have a polar group chemically attached to a non polar group, lets talk in general. In biology there are always multiple related molecules, see for example sphingolipids. If you need a technique rather than a methodical answer, then go ahead and extract with hexane or heptane. However that doesn't warranty that you understand what you are extracting. As I said, according to the polarity of the solvent you may find that bio-surfactants are "a class of molecules" with tensioactive properties. As I said, evaporate water in a rotovap, and the extract the solid (or guy thing left) with different solvents of increasing polarity. Don't worry water soluble compounds wont be extracted, but different "bio-surfactants" will appear in ether than in hexane. In my own experience sugar-esterate does dissolve in ethylether, CHCl3 and THF in different amounts, but not in water where it forms an emulsion (droplets).
As a check up; try thin layer chromatography of the different extracted bio-surfactants.
Whoa, many different compounds, and they all depend on the solvent.
"I am a bit confused. Which is the best way? Either to evaporate all the solvents until dry and dissolve it with distilled water, then freeze dried? Or to just concentrate it and leaving the concentrate, then freeze dried. Sorry, I really do not know.."
Sorry I mislead you; you ask something more basic, technical stuff.
Method:
1st. evaporate your bio-concentrate or biological sample in a rotovap, until you form a gooey paste. It won't form crystals, possibly water is trapped asan emulsion, lets call it GOOEY EXTRACT.
2. wash the gooey extract with generous amounts of hexane. Some polar bio-surfactants won't leave the guy concentrate because it contains water.
3. Pour the hexane layer (extract) into another rotovap and dry it out. Label it 1st fraction.
4. Pour another generous amount of ethyl-ether over the gooey extract and extract it as you did before (you may use ultrasound to improve the extraction).
5. Take the upper natant to another rotovap and dry it out, call it fraction 2rd.
6. Repeat extraction with THF as you did before and dry it out as you did... Call it fraction 3rd.
7. Always keep the extract gooey in the original flask. Don't mix with fractions.
8. Run flash chromatography of the extracts 1st, 2nd and 3rd over silica and you will get an idea of what (bio-surfactant) you have in the original gooey extract.
You will have to decide which fraction is get what you is what you need. That is your job.
VCalvoP
Been there First try to get rid of all the water, if possible.
1. Your extract should be left overnight with a suitable dryer (anhydrous CaSO4)
2. then evaporate in a rotovap, down to a spot of surfactant.
3. Freeze it with liquid nitrogen, either outside or add liq N2 directly to the paste. This turns sticky things very fragile. You can use a cold spatula to scratch it under liq N2.
4. If not use lyophilization. Take the frozen extract from 2 and dry vacuum. This works for pharmaceuticals.
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