How to remove water droplets from fractions?

[gbalaji87]

I found water droplets in few of my fractions collected from the column and it interferes further purification steps. How can i remove water? Is Sodium sulphate advisable?

Suggestions are welcome.

[Don_Hilton]

This would depend on the solvent and analytes involved. Can you give a bit more detail?

[gbalaji87]

The fraction I am having is of Methanol. I want to dry the whole thing. There is a possibility of alkaloids and saponins in this fraction.

[Peter Apps]

Water is completely miscible with methanol and so it cannot make separate droplets - or do the droplets appear after you have evaporated the methanol ?

What is your mobile phase ?

Peter

[prepcolumn]

Peter is right..
I think you evaporate methanol and see water, right?
You can use dried sodium sulfate or magnesium sulfate to remove water (if it is trace amount)..

[chromatographer1]

I would evaporate the methanol to almost dryness and then add anhydrous salt.

Then I would rinse the slurry with absolute methanol before evaporating to dryness.

Rod

[gbalaji87]

Yes I have seen water droplets after evaporation of methanol. Okay I shall proceed with sodium sulfate. Will this sodium sulfate interfere in the later stages of purification or is there any chance of losing compounds while recovering the compounds from sodium sulfate?

[chromatographer1]

Remember methanol unless processed fresh is never free of water contamination.

There is always a possibility. Run tests to determine if this is happening.

Rod

[prepcolumn]

Will this sodium sulfate interfere in the later stages of purification or is there any chance of losing compounds while recovering the compounds from sodium sulfate?

You have to remove hydrated sodium sulfate with filtering from your sample after dissolving with a suitable solvent..
For recovery; at first glance no, but it also depends on the chemical nature of your compounds.

[Don_Hilton]

For alkaloids and saponins, sodum sulfate should be OK. There are compounds that can be lost on sodum sulfate.

An alternative is to add a solvent that forms an azeotrope with water to the sample before evaporation. Dichloromethane works well. Benzene was used for this purpose, back in the day.

[DJ]

You have yet to mention what vessel you are using to collect fractions, or how you are evaporating the methanol, but if you are doing it under vacuum, and see residual "water" droplets that persist after the bulk of the methanol is long-gone, I have to tell you that probably ain't water, friend.

If collecting in pp centrifuge tubes, try rinsing them beforehand with propanol or MeCN, or else look into buying a better quality tube.

[DJ]

There is still a possibility that your alkaloids dry as an "oil"

Let's hear some more about the chemical nature of your solute, your methods, etc.

[gbalaji87]

For alkaloids and saponins, sodum sulfate should be OK. There are compounds that can be lost on sodum sulfate.

An alternative is to add a solvent that forms an azeotrope with water to the sample before evaporation. Dichloromethane works well. Benzene was used for this purpose, back in the day.

Yes sir it is worth to give a try. I will share my experience soon

[gbalaji87]

You have yet to mention what vessel you are using to collect fractions, or how you are evaporating the methanol, but if you are doing it under vacuum, and see residual "water" droplets that persist after the bulk of the methanol is long-gone, I have to tell you that probably ain't water, friend.

If collecting in pp centrifuge tubes, try rinsing them beforehand with propanol or MeCN, or else look into buying a better quality tube.

What it could be that droplets then?

[DJ]

Some kind of lipid or fatty acid (amide of oleic acid is used as a mold-releasing agent).. I've had this happen to me when I bought "low protein binding" tubes.. They are coated with something that apparently leaches off in presence of organic.

Try the experiment--- get some high quality pp tubes, add 100 microliters or so of water, and see how long it takes to evaporate. In a decent vacuum centrifuge, that small volume will be dry in an hr or so. If your droplets remain, it's surely not water.

Heck, you don't even need a vacuum centrifuge.. throw a tube with a few droplets of water (or whatever is in them) in a desiccator. If it is water, it'll be gone by AM.