Chromatography Forum

Check out this SPE Method Development document from Waters:

http://www.waters.com/webassets/cms/lib ... a31829.pdf


And this section of Waters' SPE catalog:

http://www.waters.com/webassets/cms/lib ... s/lcSP.pdf


It's not just the SPE, it's the selection of solvents used to wash the SPE and then elute the analytes from the SPE that make the difference.

What exactly are you looking for in these bread extracts?

maillard reaction products.

Check out this SPE Method Development document from Waters:

http://www.waters.com/webassets/cms/lib ... a31829.pdf


And this section of Waters' SPE catalog:

http://www.waters.com/webassets/cms/lib ... s/lcSP.pdf


It's not just the SPE, it's the selection of solvents used to wash the SPE and then elute the analytes from the SPE that make the difference.

What exactly are you looking for in these bread extracts?

I'm sure if you contact Uwe Neue (or your local SPE specialist), they may have some ideas in terms of eliminating this co-extracted interference and optimizing your extraction.

can I use gpc for sample clean up?

There may be ways to suppress the elution of polymers from SPE by using selective solvents for elution. If you do not know what this is, this may amount to a trial-and-error exercise. I can help you with some ideas using Oasis SPE packings.

First, I defer to anything Uwe Neue would recommend over what I may postulate. He's the expert, I'm just a guy with a lot of time on my hands. You would do well by your analysis to consult him in optimizing your SPE procedure.

My answer should probably be 'maybe', as I'm not an expert at anything, so take everything I say with a giant grain of salt. I think that optimizing your SPE extraction would be easier than adding a cleanup step (and potentially losing analytes of interest), but if you can't get the SPE optimized, I assume you could inject your analytes of interest into a GPC system, note how they separate on your GPC column, and inject a 'blank' bread extract to see how it separates out. If you can separate the components of the bread extract that are causing the hump from your analytes of interest using GPC, you could collect the fraction in which your analytes elute, evaporate the solvent, then reconstitute in your initial mobile phase conditions.

Perhaps I could start the SPE optimization discussion - What type of SPE are you using? Did you try different proportions of H2O:Methanol as an elution solvent? For example, after the 90:10 H2O:MeOH wash, did you try 50:50 MeOH:H2O, 60:40, 70:30, etc? Did you try more than one SPE?

In principle, GPC could remove a lot of such background. On the other hand, I consider GPC as way to crude and finicky for this purpose.

i just use C18 catriage and used two steps of elution.
first step, use 10% methanol to elute
second step use 100% methanol.
discard the eluent from first step since it has a lot large polysacchride and protein.
and inject sample from second step.


since compounds cause hump baseline goes all over the gradient range, I doubt finer gradient from SPE can help.

any suggestion?

First, I defer to anything Uwe Neue would recommend over what I may postulate. He's the expert, I'm just a guy with a lot of time on my hands. You would do well by your analysis to consult him in optimizing your SPE procedure.

My answer should probably be 'maybe', as I'm not an expert at anything, so take everything I say with a giant grain of salt. I think that optimizing your SPE extraction would be easier than adding a cleanup step (and potentially losing analytes of interest), but if you can't get the SPE optimized, I assume you could inject your analytes of interest into a GPC system, note how they separate on your GPC column, and inject a 'blank' bread extract to see how it separates out. If you can separate the components of the bread extract that are causing the hump from your analytes of interest using GPC, you could collect the fraction in which your analytes elute, evaporate the solvent, then reconstitute in your initial mobile phase conditions.

Perhaps I could start the SPE optimization discussion - What type of SPE are you using? Did you try different proportions of H2O:Methanol as an elution solvent? For example, after the 90:10 H2O:MeOH wash, did you try 50:50 MeOH:H2O, 60:40, 70:30, etc? Did you try more than one SPE?

so what do you think.
I wrote you one email for help for waters SPE, did you get my email?

In principle, GPC could remove a lot of such background. On the other hand, I consider GPC as way to crude and finicky for this purpose.

You had ended up in my spam folder, which I examine only rarely.

I send you a response to your questions.