best method of degassing mobile phase? - Chromatography Forum

best method of degassing mobile phase?

Discussions about HPLC, CE, TLC, SFC, and other "liquid phase" separation techniques.

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Could somebody please explain which method is best for degassing mobile phase for HPLC - sonication / Helium sparging / vacuum filtration

I have 80% acetonitrile mobile phase to degas and thought vacuum filtration is the best method to degas it but some say it's sufficient to use sonication. But I do not agree as 30min - 1 hr of sonication will cause acetonitrile to be evaporated leading to change in mobile phase composition...but they say this won't be a problem as there's high portion of organic solvent in the mobile phase...

Thank you.
It's a combination! Generally start with filtration (~0.22 um), then when you install the mobile phase on the HPLC be sure the vacuum degasser is on and the Helium sparge is working. Even a filtered mobile phase will absorb atomospheric air.
Most "consider" helium sparging to be one of the best ways. But it is expensive and uses non-renewable helium.

Most convenient is vacuum degassing.

What have I used in my 40+ years before I was retired?
First, we filtered through membranes using vacuum, in the 1970s.
Then we got an HP 1084 which used heat and vacuum on the reservoir bottles.
Next, we got an HP 1050 which used helium sparging.
Next we got HP 1050 with vacuum degasser, and used Agilent or aftermarket degassers from then on with Agilent 1100, 1200, etc. We used HPLC grade solvents and we only vacuum-membrane filtered when we mixed a solid into a mobile phase.
This must be an April Fools Day question as it has been posted more times than I can remember and a quick search will turn up the answer (though some wrong ones will turn up too).

So again, the BEST method to degass any HPLC mobile phase is continuous sparging with high purity helium. Period. Do not turn it On, then OFF. Continuous sparging maintains the level of gas in the solution. Helium is not soluble in the mobile phase (which is what you want since we are not trying to turn the mobile phase into a fizzy drink. Never use nitrogen or argon). Helium displaces oxygen in the liquid (which vacuum and sonication methods do not). Displacing oxygen reduces low UV absorbance (if you are running in the low UV range, that is a good thing) and is also best for most Fluor det methods (quenching effects). Helium sparging may be more expensive, but it is THE BEST method.

Sonicating and or vacuum flask methods are better than nothing, but as soon as you take the solutions off the apparatus, the gas goes right back into the solution. This usually appears as baseline drift up or down over time. Not recommended for any validated or critical methods.

In-line electronic HPLC vacuum degassers provide a great deal of convenience in that they can maintain the level of degassing at a stable level over time (which equals a stable baseline). You just plug them in to operate, no tanks to order, exchange or hook up. However, the vacuum channels in modern degassers may have high internal volumes (lots of flushing and wasted solvent); are flow rate sensitive (the higher the flow rate, the lower the degassing efficiency) chemical compatibility problems (THF, methylene chloride, some strong acids, sodium azide, fluorinated solvents and/or Hexanes are not compatible with some models. **We see damaged vacuum degassers every week which have been used with inappropriate solvents and/or not serviced properly); pervaporation problems (esp with aqueous phases which can and do result in the destruction of the degasser components); and internal contamination of the mobile phase when used with incompatible liquids and/or operating too long without being serviced (a very common reason for failure and may result in very expensive contamination of MS sources). - So while they are super convenient to use, that convenience comes at a cost. As with anything, you must consider YOUR own applications, resources and needs in selecting the best method to us.
... just a comment on sonication versus vacuum. I've seen some users believe that degassing by sonication means standing the solvent in a sonicating water-bath for half an hour.
This sort of sonication does nothing to the level of dissolved oxygen (in contrast to application of a vacuum, which removes a dramatic amount of oxygen very rapidly). It's probably not violent enough to cause serious cavitation in the liquid, so there's little degassing effect. You can check this with a bottle of water and a hand-held oxygen monitor.
Whatever means you use to degas your solvent, it's really important to use gas-proof tubing between the degasser and the pump. Good manufacturers will have installed the system with appropriate tubing, but after someone's had a problem and replaced the tubing with something they found in the drawer...
If you sparge or sonicate a premixed phase for extended periods you will strip some of the organic component out of it, with interesting effects on retention times.

Peter Apps
Very true Peter and why we generally do not strongly sparge mixtures, for the exact reason you state. Probably not much risk today since most users today have never seen a He sparging system on an HPLC.
Hi Chemgc779,

I recommend these two references: ... hy-and-how

and ... -degassing

The answer to the posed question is He sparging.

That said, I am with Imh's recommendation regarding the nature of the tubing leading to/from the in-line degasser (I've seen the trouble caused by replacing the proper tubing with whatever tubing happened to be in the lab drawer) firsthand.

A Practical degassing procedure that has worked well for firms I've worked with is sonication of the mobile phase while simultaneously placing the solvent under vacuum for a time period of 2 - 5 minutes (depending on where I was working at the time)
"A Practical degassing procedure that has worked well for firms I've worked with is sonication of the mobile phase while simultaneously placing the solvent under vacuum for a time period of 2 - 5 minutes (depending on where I was working at the time)"

The above procedure is not recommended for reasons previously stated. The gas from the atmosphere will diffuse right back into the solution resulting in baseline drift over extended time periods. Baseline drift is responsible for poor reproducibility and poor accuracy. We are trying to avoid that in chromatography. To be effective, degassing must be performed in a continuous manner. In-line vacuum degassers and helium sparging system can provide this.
@ Multidimensional,

Typically where I've worked, the LC instruments were purchased with in-line degassers--the combination of simultaneous sonication whilst application of vacuum in combination with LCs with in-line degassing has performed satisfactorily as far as I've observed, including for detectors such as fluorescence and pulsed integrated amperometry.

I'll reiterate from my original post, He sparging is the most effective means of degassing LC solvent mixtures.

I've used He sparging in my previous work from time-to-time as well as the approach I noted above. As with much experimental work, actually trying out potential solutions other than the "gold standard" can be advisable.
For those of us who cannot do helium sparging or vaccum degassing, I have found that the Whatman (GE Healthcare) Aqueous IFD and Solvent IFD are effective enough to keep me up and running. I am using Waters 510 pumps and typically do gradient elutions on C18 columns starting with 90/10 ACN/H2O or MeOH/H20.
@ ecigtoxdoc,

My thanks. These are membrane-based cartridges? How often do you replace these?
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