Speaking of Solvents and Mobile Phases

[Jumpshooter]

*This SOP says prepare "wet Hexane" by adding 50 ml of DI to 2L of Hexane, then shake well, and let sit for 1 hr. I did this and the DI formed a bottom layer. My question is: when pouring this Mobile Phase mixture into the Solvent Reservoir of my LC 1090, do I care about preventing in the bottom layer of DI to go in as I pour? Or do I just pour into the Solvent Reservoir and let the liquids fall as they may? The SOP was unclear--so not sure what I should assume

Thnanks

[Mark Tracy]

You want only the upper hexane layer.

In normal-phase chromatography, water is a very strong solvent, and it is impractical to rely on anhydrous solvents remaining dry. So people add a controlled amount of water; saturating solvents with water is one commonly used way to do this.

[Jumpshooter]

I will attempt--this morning--to pour over this MP into my solvent reservoir--being careful NOT let any DI get in . The mixture I'm to prep is wet Hex:ethylether:ethylpalm. 95/4/1--fir use in Vit_A.

[Mark Tracy]

I notice that the MP contains ethyl ether. Does the SOP specify a particular grade? It is important. Ether frequently contains stabilizers and ethanol is a popular one, but it really changes the chromatography.

[DR]

Best method for wetting 2L hexane - use a 2L sep. funnel, drain water via petcock before adding hexane to MP.

[Jumpshooter]

Mark, the Ethyl Ether is stabilized with "BHT". There's a warning on the MSDS that says: "this liquid can violently combust at room temparture if peroxides are formed in it...".

[Jumpshooter]

DR, thanks for the tip on using a 2L separatory funnel to remove the water from the hexane. Initially I followed Mark's advice on NOT letting any water get into my reservoir; however, the use of separatory funnel is an approach I will use next time (Monday) when I have to prep up some more MP.

[Bruce Hamilton]

Depends on your assay, but using ethyl ether with BHT "Butylated hydroxytoluene = 2,6-di-tert-butyl-4-methylphenol" may affect your results. Certainly your method should state which ether grade to use, and which stabilizers are acceptable.

Some grades of diethyl ether use 1 - 10 ppm BHT, others use 2% ethanol, others use both, and some use no inhibiter - check out range offered by Aldrich. I prefer just the ethanol inhibitor, especially if ether content of mobile phase is high, but YMMV.

Please keep having fun,

Bruce Hamilton