[GoBlue]

Hello, all,

I've just found this forum and am hoping that someone can help me out.  I'm new to mass spec, and am attempting to identify an adduct of compound X and an amino acid.  Compound X is generated by live cells and secreted, where it binds to extracellular protein.

I expose the cells to a mixture of d0-substrate, d8-substrate, and trace amounts of 14C-substrate.  By collecting the supernatant, precipitating the proteins, digesting the proteins, and then purifying on a Sep-Pak tC18 cartridge, I think I'm enriching my sample w/ the X-amino acid adduct (well, at least I have 14C eluting under conditions that I would expect it to elute during purification).

When the resulting product is subjected to LC/ESI/MS (Full Scan, + ion mode), I'm hoping to find ions with m/z, (perhaps m/z+2), and m/z+8(?) that co-chromatograph....and then I'll work from there.  I put a (?) after 8 because it's entirely possible that I lose 1, 2, or 3 deuterium atoms when the adduct is formed.

Doing this manually is becoming quite tiresome, and I'm hoping there is an easier way.  I've basically been scanning each minute of the run (approx 12 min LC/MS run of analyzable data) at a time (in QualBrowser), looking for ions that are appropriately spaced.  I then look at the chromatographs of the ions to see if the "d0" and "d8" possibilities actually do co-chromatograph.  I haven't gotten lucky so far.

I'm wondering if there is software available -- freeware would be nice -- that would be able to automate at least SOME of this for me...perhaps find peaks separated by X mass units w/ the appropriate ratio?  I could then look at the chromatographs manually....

Any help would be much appreciated!
Jim

[Arkcon]

You've told us, just about everything we didn't need to know (analytical procedure,) and gave us only a hint to what we do need to know  -- what is the manufacturer of the instrument, and the version of the software it happens to use (Qualbrower means, what?).  The function you want may be there, or not, you should try asking the manufacturer.  Or give us better info, maybe someone uses the same instrument.

[GoBlue]

It's a ThermoFinnigan TSQ Quantum Triple Quadrupole Mass Spectrometer, and I'm using the latest version of Xcalibur.  QualBrowser is a subprogram within Xcalibur.

I included details of the analytic procedure because if I simply said "Can someone help me figure out how to find peaks of interest from an LC/MS spectrum on a ThermoFinnigan TSQ Quantum Triple Quadrupole Mass Spectrometer?" that would be even less helpful.

I didn't need responses such as, "You should start with pure substrates to get a cleaner result" (they're not available) or "You need to try and purify your product of interest a bit before LC/MS" (I am), etc.  That's why I included the details.  Also, you never know, someone might read what I was doing and say, "Hey, I worked for two years on a similar problem, and let me tell you how I finally achieved my goal."  That's why I included the details.

Thank you for pointing out that you'd need to know the manufacturer of the instrument and the version of software it uses. 

Jim

[Arkcon]

Well, stick around for another user of Xcalibur, and see if there's some simple steps you're missing.  Failing that, check on Monday to see if ThermoFinnigan can help you with their app.  For third party apps, you'll have to see what export formats are available with your software, and maybe someone wrote something that imports, and processes with better functions for you.  Again, maybe someone jumps in when you post your export format, or failing that try Goggling on the export format to see if something pops up.  'Course, someone's still likely to say, "clean up your sample more. k thx bi" despite your adequate explanation, so keep on plugging.

I don't do MS, but I find your particular procedure at least a little troubling.  You have to scan each minute for a particular MS spectrum, to find your adduct peak.  Do you have to do it that way?  Is it not possible to try some dry runs, with an optical detector, to narrow down the separation window of possible adduct peaks?

[GoBlue]

Re: the procedure....  I expect the adduct to be a very small percentage of what's actually in the sample, despite my attempts to enrich the sample.  Since I don't know the retention time of the adduct, I'm looking for candidate m/z ions over small increments of retention times, so that ions of interest don't get lost in the noise.

[mienaweis]

HI~

Just a thought. Have you infused your compound and looked for the exact mass.  I know it's one of the first things to do when dealing with an unknown compound, but at least this way you would know if the extraction you did worked or not.  Make the concentration as high as possible. Run the scan for a while and see what pops up.
What is your column??? what are your run condiations?? I haven't run a ThermoFinnigan Mass Spec, but always start with going back to the very basics.