Heterogeneous Samples and Error

[GCgirl26]

Hi people,

I have more a scenario than a direct question. I'm looking for ideas/solutions from experienced chromatographers.

Currently, I am using a standard addition headspace method to analyze plant matter (prepared horseradish) for volatiles. I am being forced to throw away about 20 percent of my data because I can not get a uniform sample into vials 1, 2 and 3 (sample, spiked sample, more spiked sample). Software is flagging error outside of the acceptable range we have defined.

A simple answer would be to grind the sample to be more uniform, but my application is not that simple. Maceration activates, by enzyme activity, the production of more VOC of interest.

Does anyone have any ideas for improving accuracy under these conditions?

Thank you.

[James_Ball]

This would be similar to myrosinase activity on glucoraphanin to form sulforaphane I believe. The only way I have found to stop the activity of the enzyme is to place the sample into solvent and then grind the sample to release the bound intact active molecule instead of the final molecule after enzyme activity.

Since you are looking for volatile analytes it may be more difficult, our process was using HPLC so it is easier to control the enzyme activity with solvents. Heating or freeze drying may be other ways to deactivate the enzyme, but heat would probably cause loss of the volatiles.

[GCgirl26]

Oh wow. Thanks James. I did not realize that it could be that simple to manipulate enzyme activity. That definitely opens up some doors.

[Gizmo]

I recall using the addition of a saturated solution of CaCl2 to stop lipoxygenase activity when doing some work on the volatiles produced by that enzyme in fruit tissue, just to add other options to the table.

[James_Ball]

I recall using the addition of a saturated solution of CaCl2 to stop lipoxygenase activity when doing some work on the volatiles produced by that enzyme in fruit tissue, just to add other options to the table.

Interesting,

Anything that will denature proteins will usually stop the enzyme activity since most of the enzymes are proteins.

[Steve Reimer]

Have you considered freezing the samples while grinding? I do that for volatiles. Either dry ice or better, liquid nitrogen, will make the grinding easier and stop any reactions and loss of volatile compounds.
Our protocol for prepping tissue for ordnance compounds includes freezing with dry ice and grinding in a blender. Once the dry ice sublimes off you are left with a nice frozen powder.