Chromatography Forum

I am setting up a method for quantitatively measuring several steroid metabolites, aimed for research projects.

Should I prepare my standards in a biological matrix, or does diluting the stock solutions in H2O(or H2O:MeOH) suffice?

I can not find any good literature on this subject, and judged from the articles I have read there seems to be no standard way of doing this.

Of course, avoiding spiking "analyte free" biological matrices (serum, saliva and urine) is a lot easier. So, what kind av tests do I have to perform to prove that simple dilution in H2O is valid?

Regards,

Biological matrices have great effects on ESI, e.g., ion suppression or enhancement. When we deal with clinical specimen, we use both isotope-labeled internal standards and matrix-matched calibration standards to compensate the signal loss due to ion suppression or sample preparation. For examples, even the signal difference between synthetic urine and real urine is very significant.

If you want to use aqueous standards for clinical samples, I would suggest (1) use internal standards (2) use SRM to verify your accuracy.

Thanks for reply...

SRM = Standard Reference Material ....right?

And if i actually do find...hypothethicly speaking(i have no SRM yet)...that my method gives consistenly 93% of the value of SRM...can I just adjust my standardcurve? Or do I have to prove this relationship for the concetration range? (given that cross-over interference from other analytes are ruled out).

Sometimes it's quite tedious and requires lot of work and testing to prepare proper analyte free matrices.

I am using internal standard, isotopes of my analytes. So that should be OK.

There was a publication by Matuszewski and coworkers in Anal Chem a few years ago that specified exactly what you need to do. I don't have the complete reference handy at the moment, but it should be easy to find.

Exactly!

Million thx...