Quantitation problem

[che313]

I'm working on forensic toxicology lab where our mission is to extract drugs from urine or blood.
We are working qualitatively utilizing Agilent GC-MS QQQ after immunoassay screening.
The immunoassay results are in ng/mL and for example if i got a positive result of 2000ng/ml in urine for morphine i can confirme it easly by gc/ms.

Now adays, I asked to do it quantitatively, and i got a morphine standard from restek with conc. 1000 ug/mL. I diluted 10 folds (i.e. 0.5 mL of stock is made up to 5 mL with methanol) and run it on the gc-ms with same condition that used for qualitative but surprising that gc-ms not detect it!!

How that could be happen? I can detect morphine in samples which are in ngs but can't detect standard which is in ugs??

Do I have to worked only on SIM or MRM (if using MS2)??

[Don_Hilton]

How did you confirm the identity of the morphine peak at the nanogram level?

Assuming that you are following the same steps other than concentration, iit is possible that you have overloaded the system. Try a sample that shoud put 1 ng morphene onto the column. (if you are using a split injection, you will have to have a more concentrated sample to account for the portion discarded)

[che313]

the identity is confirmed by library search.
the injection is in splitless mode

[Don_Hilton]

While the spectrum of morphine is fairly unique, a library search is only part of the confirmation process. You need to show that the peak matches up chromatograpically with the known compound. There are compunds with similar spectra, such as norcodene - to grab the first one that comes up. If you have not run a standard to confirm by retention time as well - the current work should help.

Using the chromatographic conditions that you have used to obtain a library match for your suspected morphene peaks, inject a small quantity of morphine (on the order of a nanogram) and let us know what the result is.

Also what solvent are you using when you make the chromatographic runs to obtain a library match to the suspected morphine?

[Ron]

I suspect the mass spec is grossly overloaded, and the ion ratios are not matching the library mass spectra. Try Don's suggestion and I would be very surprised if the library match isn't much better.

The quality of the library match also depends on the way the mass spectrometer is tuned. I am not sure of the current Agilent names for their tune files, but what Agilent calls (or used to call) Standard Autotune gives better library matching results than what they call Autotune.