Chromatography Forum

Dear colleagues,

I have a problem with my separation. The root of the problem is the guar gum contained in the sample, it apparently precipitates at the column when the organic solvent ratio increases during the gradient elution and gradually increases the backpressure at the column. I tried to flush the column with water afterwards but it does not seem to help. From my experience the guar seems to have some retention so it is not possible to pass it through the column by a short isocratic step at the beginning of the gradient, on the other hand it is not so slow to be precipitated and trapped on the precolumn.

I see two possibilities - to precipitate (or separate it in other way) during the preparation of the sample or to find a way how to dissolve it and get it out of the column at the end of sequence. Do you have any experience with the guar in this way? According to the literature it is insoluble in almost everything except water. However, except of this general statement I did not find any detailed solubility information source. I'm planning to try DMSO or DMF, do you have any other idea?

Thanks.

Thickeners such as guar gum are usually quite large. My first thought is that you can remove by filtering. I don't have experience removing this type of material, but this was my first thought.

Ron J

I would try to remove it from the sample by precipitation and filtration. We've done similar in the past with gel agents where the stuff would slowly build up over a sequence and later overpressure the system.

You can use a guard column to trap your gum and wash it off during the run:
http://www.sielc.com/upload/file/pdf/SI ... r_2004.pdf

We used this approach to analyze toothpaste and milk without any sample prep. If you send me your email I will share "Direct Analysis of Melamine in Milk" and" Analysis of Amino Acid in Toothpaste" In both cases samples were injected without SPE or precipitation.

My limited experience with these used IPA to dissolve and as mobile phase with no issues.

My thanks to all of you!
Precipitation and filtration was of course one of the first ideas. However, smaller amounts of alcohol did not work sufficiently. Now I am trying higher amounts of alcohol to precipitate guar. However, I will have to check if it won't deteriorate peak shape of early eluting impurities as I will use solvent with higher eluting strength than is my mobile phase at the beginning of the run.

Vlad Orlovsky> Thank you but your solution would require additional hardware which is unfortunately unacceptable for me.

Most of modern LC systems have a switching valve, we have Agilent 1100 units which are 15 years old and they are equipped with switching valve.

Most of modern LC systems have a switching valve, we have Agilent 1100 units which are 15 years old and they are equipped with switching valve.

Yes, I know. But to keep the method as simple as possible I would preferably employ a precipitation step followed by filtration (which is necessary anyway) so I will try this approach first.

Just to report the results, I tried the precipitation-and-filtration way and it worked well, no pressure increases any longer and recovery of impurities not affected. Thank you all!

If you want to try and cut the percipitation+filtration step you could try a monolithic column they are much less suseptible to blockage by gunk like this.