QQQ vs HRMS: Which Mass Spec Wins for Impurity Quantitation? Emery Pharma’s Bridging Study Reveals

Foreign. Welcome to our Emery Pharma video series. Today we're going to talk about something really exciting. This is a case study, but more specifically a bridging study. In this bridging study we're going to talk about how we quantitate impurities or look for certain impurities within drug products. But only this time we're not only going to use our accurate instrumentation, but do a comparative analysis between two different types of instruments. And in this case, our QQQ or our triple quad LC Ms. Ms. Or our high resolution mass spec or orbitrap explorist 240. So to introduce this scope of work into this case study, we are ideally trying to compare these two different technologies. We're evaluating not just standard performance parameters such as linearity, accuracy, precision, but also the ability to detect and quantify specific impurities within drug products. Both of these methodologies we want to make sure adhere and comply with our FDA guidelines and any sort of regulatory alignments. And lastly, we want to determine if both of these instruments perform the same, not just in those parameters, but in the actual quantitation itself. So one of the first parameters we look at is selectivity and specificity. Right. So what we're doing here in the upper left chromatogram is, is we're looking at an overlaid MRM from our QQQ instrument. So we're comparing the actual impurity of interest versus a splank solvent. As you can see here, when we look at signal versus retention time in our pink, we see a nice big peak representing our standard. And in blue, which is going to be just showing that baseline down there, we are just looking at our blank solvent, indicating to us that our method is can accurately and specifically identify the standard of interest. Similarly, if we look over here on the right side, this chromatogram, we're again looking at the same intensity versus retention time, except this time we're using a different viewpoint, which is our selective ion monitoring. From our HRMs here in Orange, we see the standard with a nice peak and again, when compared to our black solvent peak, here again indicating the specificity of our method. Data wise, you can see in this data table here what that represents. So what we're looking at is the different peak areas from both of these instrumentations of the same standard compared to a blank, both of which show high specificity for our standard or for our impurity of interest. Really the acceptance parameters that we look for is any sort of interference with the retention time or ensuring that the blank signal has to be less than 110 of the signal of the standard, both of which parameters are met with both of these different technologies. So, moving forward, we now want to focus on our linearity and range. This is the range of which we can accurately quantitate our analyte of interest or our impurity of interest, and the range of which that linearity or that response or that correlation between signal and concentration stays linear. So here we're looking again at response, except now we're looking at concentration. So you can see from our QQQ throughout this range, again redacted for this case study purposes, we have an R squared value greater than 0.9, which again indicates a strong correlation between signal and concentration. Similarly, on the right for our HRMs, we have the same linear range and again the same linearity as represented by that greater than 0.9 correlation coefficient. A standard data table would look something like this where you have the different standards, the calculated concentration from the instrument, and then the corresponding percent accuracy based on what we're getting versus what we should be. And again, the same thing happens for QQQ, as are HRMs. Both of our instruments, as you can see, have accuracy values well within acceptable criteria and again demonstrating that linearity greater than 0.9. Now we move to accuracy. So how accurate are these methods? Right, that calibration curve, how can it accurately quantitate external standards? So Here we have three different QCs, typically a low, medium and high, and again a calculated concentration and a corresponding percent accuracy for both QQQ and hrms. As you can see from this case study, we were able to successfully get accurate values for both of these instruments. What's really nuanced when you do sort of drug testing is your ability to detect these impurities or these analytes within a complex matrix. A lot of methods out there can do standard analysis in neat solvents, but really the relevance of that isn't that translatable. What we're trying to do is detect analytes in complex matrices like these drug products themselves. So what we did here was actually an in matrix accuracy or recovery study. We took the unspiked drug product. So we naturally just took this drug product and assessed for this amyloid of interest. Knowing the amount of that pre existing analyte is going to be important when you start doing these recovery studies. We took that same drug product and spiked it with three known amounts, again a low, medium and high. Knowing how much was pre existing from the unspiked plus the amount that we spiked, we were able to determine what our percent accuracy was based on those theoretical yields for both QQ and HRMs. Interestingly, both of those accuracy values fell well within the acceptance criteria, making us feel very comfortable that our methods can quantitate this impurity or these analytes accurately within whatever drug product we are testing. So really the big takeaway from this case study is that we were able to not only demonstrate the side by side parallelism of both of these methods in terms of their specificity, their selectivity, their the linearity and range, their accuracy as well as their in matrix accuracy, we were also able to demonstrate that just amongst themselves they all demonstrated strong linearity and well within acceptable accuracy limits or at least within the acceptance criteria for both methods. This gave us the confidence to move forward with our bridging study being complete, being able to now quantitate this analyte or this impurity in that drug product. I would encourage you to check out our website subscribe for any more videos or more content about anything that Emery Pharma offers. We'd be happy to get on the phone and discuss your your future projects. Thank you for watching.