I extracted my culture filtrate (cell-free) with the ethyl acetate. After I shook it for 5 minutes and let it separate, there were chunks water droplets inside the organic phase. Can anyone please tell me why this occurred and how to solve it?
Is it correct to add ethanol?
Probably an emulsion forms. Shake it gently and if necessary remove water by passing ethyl acetate by a small collumn of anydrous magnesium sulphate. Good luck..
Drain the ethyl acetate layer and add anhydro sodium sulphate to the ethyl acetate layer. Keep it standing and filter for 5-10 min. You will have dry ethyl acetate layer.
A correction. Read 'anhydrous sodium sulphate' instead of 'anhydro'
Thank you very much for the advice. Just would like to ask how about if adding ethanol to the mixture? Will it affects the sample? Thanks.
I beleive your culture filtrate (broth) is water based. If you add ethanol you tend to make water-ethylacetate layers more miscible. So avoid it. If you are thinking about using ethanol instead of ethyl acetate - ethanol being miscible with water, you will not be able to do paritioning. Either case avoid ethanol.
Couple of more points.
1. use a solvent like dichloromethane as a first partition solvent. Being heavy, it would sit at the bottom with the top layer being water. drain the dichloromethane layer
2. then use ethyl acetate for partition with the broth, which would be top layer.
Using two solvents which have different polarities, yet immiscible with water would provide exhaustive extraction.
While using some of these solvents for partitioning with the broth, even though you have filtered the broth, will give an emulsion which is difficult to separate and takes a long time to separate. In such instances, filter the whole solvent-broth mixture (emulsion) thru' celite in a sintered buchner funnel which will break the emulsion and provide two clear layers.
Thank Dr. Suresh for the opinion. You mean if use ethanol it will effects the sample too even I just add a small amount into the mixture? The misibility of the water-ethyl acetate layer will cause the more polar compounds to enter the organic phase? Thanks !
It sounds to me like the opening poster did not get a total separation of the organic from aqueous layer. I would suggest uding a centrifudge to get two clear layers.
- Badges
- Science topic
- Similar topics
- Analytical Chemistry
- Extraction