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We used to have issues with viper and nanoviper lines but now we use a torque screwdriver (H4DETSD) with a hexagonal adaptor bit with a cutout for the line to tighten the lines to ~30 cNm and ~20 cNm for metal viper and glass nanoviper connections respectively. We also do a near fully connected leak check with a pre-detector plug (closed system). This has resolved all viper tightening leak issues for us.
Also not a statistician, but I think this is unpaired t-test for sure. Like you say, you have different numbers of samples in each time point. I guess you want to see if there is a systematic difference (increase or decrease) from season to season, which would be a good case for paired. If it was paired, it would have to be on the means, since the individual replicates can’t really be linked to each other I don’t think. Do you have to do ANOVA also?
The state of California is working with one, but I’m not working on that project. Perhaps @mccord can share?
Have you considered running these through any in silico fragmenters? Even the “MassSpec Scissors” in ChemSketch could be enlightening.
I’d personally reach out to Angela Batt since she has a Waters QTOF. This just got published, but if I remember correctly points to other papers for some of the methods parameters. https://doi.org/10.1021/acs.est.3c07514
*Disclaimer, not a statistician*
Volcano plots? This plots the fold change versus p-value (significance). Anything outside some set criteria in both axes would be the ones to look at.I’m not a very good statistician, but your setup reminded me strongly of this one: https://doi.org/10.1021/acs.est.3c07514
Brunelle, L. D., et al. (2024). “De facto Water Reuse: Investigating the Fate and Transport of Chemicals of Emerging Concern from Wastewater Discharge through Drinking Water Treatment Using Non-targeted Analysis and Suspect Screening.” Environmental Science & Technology 58(5): 2468-2478.I just read the paper this week, and don’t know that they performed stats tests, but rather qualitatively reviewed the data. Perhaps that’s a good approach here?
The only NTA method info I have found from Waters is their PFAS NTA Applications Note that uses a Xevo G2-XS QToF MS. However, it sounds like you are trying to do something more inclusive than PFAS. The note can be found at https://www.waters.com/content/dam/waters/en/app-notes/2021/720007184/720007184-en.pdf (Application Note: Approaches to Non-targeted Analyses of Per- and Polyfluoroalkyl Substances (PFAS) in Environmental Samples)
My generic method has stepped NCE at 30, 65, and 100. We’re running on a Q Exactive HF which uses an m/z of 500 for normalization. The degree of fragmentation will totally depend on the individual compounds; with this setup, I still see occasional compounds with only a precursor ion peak.
It may also help in your decision-making to consider what, if any, outside libraries for spectra searching you may be using. Incorporating the stepped energies can add some additional complexity since it’s a composite. mzCloud is supposed to be able to generate a composite spectrum to match, as closely as possible, your experimental spectrum. I have had some challenges, though, comparing my spectrum to spectra found on MoNA or Massbank where a single energy was used and it’s not clear if the spectrum was generated with absolute or normalized energies.
I don’t know of a quick way, but you can use the Advanced Search in ChemSpider (https://www.chemspider.com/FullSearch.aspx) and set the data source to whichever list you want to return the complete list of analytes in that list.
High Jargon Answer: You can directly access the spectra from the SQLite database and parse the spectra binary streams to produce a text output in whatever format you need.
Low Jargon Answer: Your results file has the spectra stored in a compressed format and Thermo doesn’t have a way to export them. As boettger mentioned, I have a script that pulls out spectra from an mzVault file, but the same method should work to pull them out of a CD results file. I would just need a test file to work with and some spare time to modify the script. If you send me an email or share a test file I can try to scrape something together.
According to the help menu, a “DIA only” flag happens when “Only data‑independent [all ions fragmentation (AIF)] scans are available.” I don’t know if a “DIA only” flag is possible if the data were collected with a DDA method. The “DDA for other ion” is dependent on the settings for “Ions” in the Detect Compounds node and for “Preferred Ions” in the Group Compounds node. For example, if you set your Ions to be [M+H]+ and [M+NH4]+ in Detect Compounds but only set your Preferred Ion to [M+H]+ in Group Compounds, you will get a “DDA for other ion” flag if there is an MS2 spectrum for an ion that the software assigned as an [M+NH4]+ adduct (since this wasn’t one of the Preferred Ions, it’s an “other ion”).
I know Logan Roach with James McCord has been working on an R script to process spectra and send them off to Tony Williams. I’m pretty sure he was starting from spectra in Compound Discoverer, so he must have a method of exporting the desired MS2 spectra from Compound Discoverer directly.
If you want to see live pump pressure plotting before starting a sample run that can be done in the Direct Control software. You will need to click on the settings cog along the top bar. This will open a window and you can scroll down to the eluting pump or loading pump. Click on eluting pump pressure, then click the commands tab. Turn on AqcOn. This will start plotting a live stream of the pump pressure. You will need to make sure to turn this off prior to running your sample list. Otherwise, the system will not run the sample list. Hopefully this helps!
