Pentobarbital on LC/MS/MS?

[mtorrey]

Hello

I'm wondering if anyone does Pentobarbital by LC/MS/MS.

To start with we just got this Waters Acquity LC/MS/MS (TQ) to do tricyclic's, pentos, benzo's and hopefully phospholipids (for fetal lung maturity). This is our first LC/MS/MS and I haven't even done LC for 15 years (HP 1090). Mostly we do GC/MS. Anyway I am really out of my element.

They (Waters) insured us we could do Pentobarbital on this. We wanted to move them off the GC/MS because the procedure is so long and the derivitizing agent is so bad for the column.

But I cannot get this drug to ionize in either ESI+ or ESI-. Cannot figure out what I'm doing wrong. I can get the tricyclics to ionize, almost have a MRM method worked up.

Cannot find anything on Pentobarbital by LC/MS/MS, it is mostly done by GC/FID or GC/MS.

So maybe this can't be done by ESI, wonder if anyone has any ideas. Please excuse my ignorance in LC/MS/MS, I admit I have no clue.
Been trying to infuse a 1 ug/mL solution in 30:70 water:ACN solution. I get nothing.

TIA

[Kostas Petritis]

Here is a reference for you:

Title: Method for screening and quantitative determination of serum levels of salicylic acid, acetaminophen, theophylline, phenobarbital, bromvalerylurea, pentobarbital, and amobarbital using liquid chromatography/electrospray mass spectrometry
Author(s): Hori Y, Fujisawa M, Shimada K, Hirose Y, Yoshioka T
Source: BIOLOGICAL & PHARMACEUTICAL BULLETIN 29 (1): 7-13 JAN 2006

They analyse the Pentobarbital in negative ion mode. I suggest that you try to duplicate the method with the chromatographic conditions and mobile phase additives suggested.

There are several details to pay attention when using an LC-MS-MS. It would be best if you had someone that knows the instrument for a week so you can develop with him/her a couple of methods (i.e. go through mobile phase that you can use, how to find the best MRM conditions for each molecule, how to fine tune the electrospray emmiters, how the different ion optic parameters change the ESI response for different molecules in MS and MS/MS mode and what to expect etc).

If ever you do not succed you may repost here...

[mtorrey]

Thanks, I will try to get that reference.

I had about 3 days training by the engineers on how to calibrate, tune and maintenance but about 25% of it was lost in translation, another 25% just didn't sink in. They never showed me negative mode ESI. Then because of some contract/purchasing snafu I won't be going to TQ school like I was suppose to.

I've tryed to get application help from waters on pentobarbital but they haven't gotten back to me. Either they don't care or don't know.

Frankly this has been one discouraging situation.