Can I combine calibrations into one uber mega-calibration?

Greetings forum denizens,

I'm running on Openlab CDS Version C.01.10[201] . I'm (still) working on setting up our GC for pesticide analysis. We're looking for about 30 compounds. As many of you know,  these can elute very closely. To make things easier, we are using two sets of standards: one compound mixture is "A', the other is "B". I have made calibration standard for both and entered them. To reduce paperwork, my boss would like to replicate an old method we used twenty years on a 5890 (!) using an old, old version of Chemstation. This method used one calibration that combined those of standards "A" and "B" ;  one curve with one report format that worked for both standards.

My question is simple: How can I combine the curves for both "A" and "B" in a single pesticide calibration? I played around and was unable to do so. I tried adding peaks. I tried adding one standard as separate levels (a dumb idea, but figured it was worth a try). I even hit upon the idea of overlaying chromatograms and trying to calibrate off this combination. Believe it or not, it worked for the first cal level, but I couldn't add more levels to the curve. Whether I can do this isn't going to bring calamity on my GC world. If worse comes to worse I can just use separate curves for the "A" and "B" standards. But we were capable of doing this on Chemstation , so I would think I can do it with my current OpenlLab CDS.

  • Hi

    OpenLab Chemstation that you're using should be able to do exactly what the old, old version did.

    Have you tried something like this:

    Load data for level 1 of the A standards and create a new calibration table.  Set up the default amount for level 1.

    Load data for level 1 of the B standards and Add Peaks to the table.

    You should then be able to add all of the other levels for one of the groups.  If you want, you could load up the other group and recalibrate at the appropriate levels for each of the datafiles.  I would probably just make sure the calibration table has the correct amounts for each compound at each level and reprocess the sequence, to calculate the calibrations for each compound.

    Hope that makes some sense!

    /Andy

  • It actually does. I tried that, and it sort of works. Let me explain what I'm doing:

    I load the chromatogram for "A, manually integrate whatever peaks I watn, remove things I don't want.  I do the same with the first chromatogram for "B". This will make it easier for me to see what I'm doing.

    I load "A" and start a new calibration table with an automatic setup .I set the first cal level and voila. I have all the compounds I want from A. It works like it always does. I assign compound names, then  I load the first chromatogram for "B". After loading "B" I have tried a couple things:

    1) I go to the calibration pull-down menu and click on the first "add peaks". I then add the cal level and the default amount in the window that pops up. The result is that it adds SOME peaks but not all. I considered that maybe this was a region where both standards will overlap and that might cause some problems. But the retention times ARE different, so I don't see why that would cause a problem.

    2) I go the cal pull-down menu and click the lower "add peaks". It says "add selected peaks from chromatogram". I enter the cal level. There is no window for a concentration. I figure I can scroll down the table when the peaks are added and add the concentrations manually. Nothing happens. No peaks are added. I take from this that the problem, is simple: I'm probably not selecting the peaks properly or not at all. I thought maybe just manually integrating them would select them. That's how I typically select my peaks when I calibrate. But I'm also typically calibrating off one standard, so  I'm certainly wrong in assuming integrating the peaks manually is a means of selecting them. The cal pull-down menu has "select peaks" highlighted (it's in the same are as "delete peaks", add peaks", and "recalibrate compounds)".I tried the help menu, but... well, let me just say that for my purposes it's improperly titled.

    3) I go down to the buttons near the cal table itself. There is one labeled "insert". I move the cursor to the peak I want and click "insert". A window comes up and I make sure it has the correct signal in the signal window. I enter a retention time. I get an entry there with the time and no compound label since I didn't enter one. however, the curve is flat . I try a linear curve fit. I still have no curve. so I scrapped that idea. I still think my second route is the correct one, I'm just not selecting the peaks. Sometimes when I click on the peaks in the chromatogram window the highlight with a blue rectangle. Sometimes the rectangle is grey Again, I peruse the help menu, but get nowhere..

  • Hmmm.... sounds like you're doing exactly what I would do, apart from my first step would be new calibration table.

    I've tried with some example HPLC data, and came up against a similar problem.  The only way that I found around it was to create a new calibration table from the "A" sample.  Add peaks from the "B" sample.

    I could then load up the "A" sample level 2 data and Calibration...Add level from the menu for each level, that deals with half of the peaks.  I couldn't find a way to do the same for the "B" samples.  Nothing seemed to work.

    All I could then do was to use the calibration table buttons to Insert levels.  Clicking to the left of the table allows the selection of a row.  Clicking the Insert button then allows the addition of a new level.

    Repeating this for each additional level for all of the remaining compounds manually builds a calibration table with all levels for all compounds.

    To add the correct response factors, I just reprocessed the sequence.

    Hope that helps.  It feels like there should be an easier way.

    Andy

  • Ok, so here's what I did: I started a new cal table and loaded up the first "A" standard the way I alsways have calibrated. Everything is fine. I then load the first "B" standard. As this is the first standard, I click "insert". When the window comes up I UNCLICK "insert peaks on all calibrated signals". . I then pick the first peak for "B" I would like to add. I Add it as a COMPOUND and enter the retention time. It adds the peak. I then make sure the peak has a name and an amount.

    I then load the next standard level. I I pick the peak and click "Insert" again. It tells me "Current Peak: ECD 1A: 5.659". This is the retention time of the peak I want, so I know OpenLab and I are looking at the same peak.  I add the second LEVEL and... It adds it! Great! I can get the second standard added to a peak. At least I know I'm getting somewhere. Here's the problem: I have two peaks for the compound. Each of the two cal levels have amounts. Still, I have no curve. I am using a linear fit forced through the origin, and my curve has no slope. It's just a horizontal line
    I  change the curve fits, but get nowhere. I didn't think it would. And I have no idea why a linear curve with three points with different "Y" axis values yields a flat, horizontal line. As the "Y" axis is area, I take it it's reading the are of the added peaks as zero as well. But I have no clue why.

    ETA: I think I have a solution to my area problem. A bit after typing the pervious message, I remembered I can add fields using the calibration table options, One of these fields is the peak area. Manually integrating the peaks gives areas on the report. I suspect I can manually integrate the peaks I want, print out the report, and use these area values in the field of the calibration table. If I can do this, then I should get a usable curve. Man, what a hassle. But if that's what I have to do, the a bit of extra work early on will save the effort opf having to switch between methods a couple times for each sample.

    ETA (again): I quickly threw together a new report that consists of sample id info, retention times, and peak areas. This will make it easier for me to get the peak areas and, again I hope, a usable curve. This will save me some effort. I won't know until I get some time tomorrow to try my idea out. But I don't see why it wouldn't work. I'll let you know how it goes.

    Still, can you think of a simpler way to get these areas into the curve?

  • Check that the sample type and levels are set correctly in the sequence table. Reprocessing the sequence should pull in all of the peak areas and show your calibration curve correctly.  Remember to set the values in the Update RF column in the sequence table to Replace before reprocessing.

    /Andy

  • I'll give that a shot when I get the chance. II take it you basically mean to enter the new calibration, save it as a new master method then save it as a sequence method to the appropriate data file and use that for reprocessing, right? Just out of curiosity, can you point me in a direction to where I can read more about the sample type and why it matters? To be honest, with the old software we use we never had anything of the sort, so I always assumed the sample type was more for information purposes than anything. Sorry for the extreme ignorange; I'm trying to rectify that.

  • I wonder if this would contain the information: https://www.agilent.com/cs/library/usermanuals/public/CDS_CS_Concepts.pdf

    Not sure about what you said at the beginning of your last message.  For me, the process is to add all of the compounds and levels to the sequence method where you have all of the data for the standards, then update the master version of this method.  Then, every time you use this method in a sequence, the calibration table will exist, and any injections made from calibration standards (sample type Calibration) will be used to update the calibration table.  It's important to show the Update RF column in the sequence table.  This column allows you to choose how the new response factors for these new standards, are used to update the calibration.  The options are to Replace (this is what I would do), Average (takes the current data and averages it with the existing data in the calibration table) or Ignore (just use the existing calibration data).

    Only the calibration sample type is treated differently to the others, the data from these injections being used to automatically update calibration tables.

    /Andy

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