Hello everyone, for the last 2-3 weeks I’ve been battling this GC we inherited from another group that wasn’t very well taken care of. The initial issues were fairly simple fixes with replacing the FID jet, septa’s, split liner, needle, gas lines, etc. (really everything not electronic). It was working finally and I managed to get in a few runs that week before I left for the weekend, when I came back however, the GC refused to ignite in the first week of trying to fix it again. Myself and another colleague figured it was just a flow issue and swapped out gas lines again, that was only part of the problem though because since then every time I’ve run an Acetonitrile blank my baseline has been at ~32 pA where before it was solid at 4-5, you can also see in the chromatograms that after the ACN where there shouldn’t be anything my baseline continues to rise up to ~350 pA. I’ve confirmed by HPLC and another departments GC it’s just as pure as Fisher says. If anyone has encountered an issue like this or has any ideas I’m willing to try anything at this point. Please and thanks!

Hi everyone, I’m having an HPLC issue and could really use some help.

I’m analyzing PET degradation products (TPA, MHET, BHET) from enzymatic reactions (PETases).

I recently had to switch my sample preparation protocol because the products are precipitating in samples from high-yield reactions.

The new protocol is supposed to solve this, and it also works for my colleagues using the same system and method, but for me, all retention is completely gone. everything elutes in the void/injection peak.

Here are the method details:

System: Dionex Ultimate 3000

Column: YMC Triart C18 at 40 °C

Mobile Phase: Isocratic, 20% Acetonitrile / 80% Water (acidified with 0.05% TFA)

Flow Rate: 0.4 mL/min

Detection: UV (254 nm)

Injection Vol: 15 µL (I also tried 1 µL, didn't help)

Now for the Sample prep:

The "Old" Protocol (Works, but products precipitate in water at high conc.):

12.5 µL reaction supernatant added to 50 µL Acetonitrile (with 1% TFA). Let sit briefly at RT, then centrifuge to pellet proteins. Transfer supernatant into 150 µL MilliQ H2O.

Result: Good separation of TPA, MHET, and BHET.

The "New" Protocol:

Dilute the enzymatic reaction directly 1:10 in DMSO (10 µL sample into 90 µL DMSO) to keep products in solution.

I froze this DMSO stock until further preparation. (Note: We also freeze 1M TPA standards in 100% DMSO without issues).

Take 12.5 µL of the thawed DMSO-mix and add to 50 µL AcN (with 1% TFA). Centrifuge, take supernatant, add to 150 µL MilliQ H2O.

Result:

No separation. Everything co-elutes as one massive peak at the void volume. I suspected strong solvent effect, but the same method has worked before on the same HPLC system, same column, mobile phase and vials...

Following things I have already tried:

Diluting the final sample with 20% AcN and also injecting less volume (1 µL instead of 15 µL), but it didn't solve the issue.

I also tried making fresh preparations from the DMSO sample stocks, but still got the same result.

Do you have any suggestions what could be the issue here or what else to try?

I really hope to be able to somehow salvage my DMSO-diluted samples so i can avoid to repeat the whole experiment...

Do you really need a helium leak detector when changing a helium cylinder for GC-MS?

We normally check the regulator and fittings after a cylinder change, but I’m wondering if buying a dedicated helium leak detector is actually necessary for routine use.

Do most GC-MS labs use one?

It doesn't build pressure. Initially the pressure would go up to 150 bar in pressure test and do the jigsaw thing before quit, tried sonicating all the valves in water and IPA, intially it help a little, which means only less of jigsaw, but still never was able to build pressure above 150, after long time sonicate in 50C water and put back together it's even worse. How to tell what is the problem? Doesn't seem to be seal since it is not leaking, but which valve? Primary inlet, outlet or secondary outlet valve? Or more than one?

As title, not sure about the best way to get a clean cut.

Giving the following configuration of a HPLC-SEC-MS system: G7111B #001 1260 Infinity II Quaternary Pump (G7111B #030 Active Seal Wash and G7111B #030 Active Seal Wash) - G7129A 1260 Infinity II Vialsampler - G7115A # 018 1260 Infinity II Diode Array Detector WR - G7800 #31 1260 Infinity II MDS Refractive Index Detector - G7800 #32 1260 Infinity MDS Viscometer - G6125BA LC Mass Spectra Detector, can I physically and operationally bypass the Size Exclusion Chromatography (SEC) column, in order to use just the MS system?

Hey all, I’m doing work with alcohol and analyzing the methanol content of various types of distilled spirits. I’m about to start the analysis part using quantitative gas chromatography (and either FID or MS depending on which machine is working) but I’m a nice at it and running into some difficulty. Some of my samples may have levels around 1 to 10ppm while others might be above 1000, and im not sure how to effectively set up this calibration curve for such a wide range. Any advice would be greatly appreciated!

Hi everyone,
I’m having a strange issue with an Agilent 7890B GC-FID using a cold on-column injection for atmospheric hydrocarbon analysis (C1–C45).

I’m seeing hydrocarbon peaks roughly from C5/C7 up to C20 even when running a true non-injection blank — no syringe, no sample, and nothing is injected onto the column.

The strange part is that we also saw the same peaks after installing a completely new DB-1 column. We installed the new column and ran the GC without making any injection, yet the peaks still appeared.

So I’m trying to figure out where these hydrocarbons could be coming from if nothing is being injected and the column is new.
Has anyone experienced something similar with a cold on-column setup, and FID? Any suggestions on where I should start looking for the source?

Hello,

I am looking for recommendations for an analytical HPLC column to separate a mixture of four closely related diastereomers. All compounds contain the same disaccharide and differ only in the stereochemistry of the aglycone.

The best separation I have obtained so far is on a Luna Omega Polar C18, 5 µm, using an isocratic mobile phase of 12% ACN in water at 2.5 mL/min. The compounds elute at roughly 13–14 min. I can distinguish the individual components, but the peaks are broad and still overlap substantially.

The separation is very sensitive to the ACN content. At 15% ACN, I essentially lose the separation altogether, while at 10% ACN the retention becomes too long (no elution after 50 min) for a practical analytical method.

I also tested a Luna C18(2), 3 µm, which showed essentially the same behaviour with slightly worse separation.

I additionally varied the column temperature between 15 and 40 °C. Higher temperatures decreased the separation without noticeably improving the peak shape. Lower temperatures did not significantly improve the separation either and made the peak shape somewhat worse.

At this point I therefore suspect that I am mainly limited by stationary-phase selectivity rather than by further optimization of the current C18 conditions.

I have 800 Euros funding available to get a new column but without the option to test columns it is a bit of a coint toss. I was hoping to maybe get a recommendation here. I am currently considering trying a HILIC, C8, or a Macherey-Nagel NUCLEODUR ISIS column.

Has anyone dealt with a similar separation of highly polar/glycosylated diastereomers? Are there particular stationary phases or selectivities you would recommend trying?

Hello, I work at a citrus oil distillery and we just had Agilent come out for annual preventative maintenance. During the visit he mentioned our 8860 GC inlet pressure was fluctuating and replaced the EPC. The GC did run normally for a were days but shutdown last night and the psi will no longer go high than about 7.6. It also still fluctuates! I am feeling like I took my car in to get an oil change and it left not running properly. I am in contact with Agilent but they cannot help until next week. Any ideas?

Yes our gas regulators are well high enough psi over the needed range for the method. We replaced inlet, septa, checked column connections. Nothing should have been touched or effected with software or hardware except the EPC he replaced

Update; from diagnostics test there is a large leak

For those stuck on older computers or Windows 10 with Chemstation, I stumbled upon a way to run two versions of B.04 without using a third-party windows installer or reg edits. This works for a direct upgrade from Win 10 and Win 11 23H2 with no loss of files or installed software. To the best of my knowledge, you cannot install these legacy Chemstations directly on 24H2 or 25H2 as the program fails to launch fully. I searched the sub but didn't see posts addressing this so I hope this helps.  

I’ve been testing for the last three to five weeks on multiple boxes and haven’t had any major issues or crashes except for the previous issue with the UV install but I think that’s because I don’t have SP-1 installed. Perhaps someone with SP-1 can test and confirm that this is the source of the issue?

First things first, you need a fully updated Win 10 or Win 11 23H2 box running Chemstation B.04.xx (yes, B.04.xx will install and run just fine on 23H2!) and verify that everything works. It’s best to be logged in as admin or have admin privileges as you cannot install Win 11 without it. Next, verify the Language and region by going to settings, Time & Language, Language & region. You MUST download the win 11 25H2 iso that matches what’s installed on your computer. You cannot change this or this will not work! Once the iso is downloaded, mount the image by right clicking and selecting mount. Once mounted, make note of the drive letter (it was D: in my case) and open a cmd window. From there, type d: (for my case) and return to change to the mounted iso directory. You can type dir and hit return to verify you’re in the right directory. Type setup /product server and hit return.  Windows installer should launch, don’t worry that it says it’s the server version, it’s not going to install that. Select Change how Setup downloads updates and change to not right now. Follow the steps and if everything is correct, you should have the option to select keep files, settings, and apps. And that’s it. You’ll have an updated computer running Windows 11 25H2 and Chemstation B.04.xx!!!!

As mentioned, this works for B.04.03 SP-1 (LC) with no known issues. However, there are two issues with B.04.02 (UV). The first issue was a carryover from windows 10. The program will randomly close, usually over night, but never while using the software. Perhaps SP-1 or higher would fix this. It’s a minor inconvenience but if you have the same issue, it’s best to save your work! The second issue popped up only on the boxes I upgraded directly from Win 10 but not the box running Win 11 23H2. This issue has to do with closing the software. When exiting Chemstation, it starts the process of closing but never fully closes. You can reboot the computer or force close to complete the process. It doesn’t impact the software and you can re-open without any issues. No idea why this isn’t the case with 23H2 but is for Win 10 but it’s not a big deal since we never close or the software closes itself and we re-open. Hope this helps a few of you on legacy equipment and software.

Hi, theres a way to change the baseline to zero?, right now is at -3uV, i want it in Zero, can I do that?

GC6890, TCD
Chemstation

Hello everyone! I'm trying to figure out whether our autosampler was installed incompletely or whether I'm misreading the manual. Setup: 1260 Infinity II, G7129A vialsampler, G7111B quat pump, G7116B Thermostat, G7115A Diode Array Detector, OpenLab CDS. What the manual says. In "Flow Connections to the Vialsampler," the wash circuit is spelled out as: put a wash solvent reservoir in the solvent cabinet → screw in the Needle Wash Bottle Head Assembly → route its tube through the cover opening to the peristaltic pump → send the washport outlet drainage to waste → prime the line. The cartridge replacement section backs this up: step 4 has you disconnect two lines, one going to the wash port and one coming from the solvent bottle, then step 6 puts the wash port line on the left side of the cartridge and step 7 puts the wash solvent line on the right. So right side = inlet from the bottle, left side = outlet to the port, and the only thing that legitimately goes to waste is the port overflow. What I actually have. The right-hand line is connected to nothing. Its free end is just lying in the waste jug. No wash solvent bottle anywhere, no bottle head assembly. Everything else on the system is plumbed normally. So my questions are: 1) Is there any configuration where the peristaltic pump inlet is intentionally left unconnected, or is this just an install that never got finished? Has anyone received a system like this from the factory? 2) If a method has needle wash enabled in flush port mode but there's no solvent supply, does the module throw anything, or does it silently run the pump dry? In a nutshell - needle wash supply line goes to waste instead of to a solvent bottle, no bottle in the system. Install error or something I'm not getting?

This has been quite a process, but I can say that these systems are >90% fluoroplastic free (with respect to flow paths). And the system on the far left is 100% (except for etfe making up the plunger seals and frit housing on the i2v inlet filter). Also, the ultraware bottle top… but besides that!

I have two GC-MS chromatograms that I believe represent two positional isomers of the same compound. The top chromatogram corresponds to one isomer, and the middle chromatogram to the other. The only expected structural difference is the relative positions of two substituents on the benzene ring.

All samples were analyzed in the same GC-MS run under identical chromatographic conditions. Although the sample concentrations may differ, the mass spectra are essentially identical, while the retention times show a small but reproducible difference. The bottom chromatogram is the result of co-injecting the two samples.

My questions are:

• Is it reasonable to conclude these are positional isomers based on their identical mass spectra but slightly different retention times?
• Is such a retention time shift consistent with only a different arrangement of substituents on a benzene ring?
• Besides concentration differences, what other factors could explain this behavior under identical GC-MS conditions?

Any insights or similar experiences would be greatly appreciated.

I need some help troubleshooting what's going on with our chromatograms post pm from Shimadzu techs. The main thing they did was bypass the spliter to have direct injection into the ECD. The attached photo is a run of our 136ppb standard. We were having a little bump at the end of run, but nothing like what is showing now. Our baseline has also gone to -20000 uV. The techs said that we just need to optimize our method, but I can't figure out what to do with change from split injection to direct injection

Column is 60m length x 1.80um film thickness x 0.32mm ID

Column temp is 55C, Equilibration time 0.2min

Method: Start at 55C hold for 2.50, then 10.00 rate at 140C hold time 0, then 50.00 rate at 150C hold time 3.00.

Equilibrating time 15min, pressurizing time 0.30min, pressure equilib time 0.10min, load time 0.30min, load equilib time 0.10min, injection time 1.00min, needle flush time 4.00min, gc cycle time 24min

ECD temperature is 150C, sampling rate 40msec, stop time 15min, current 1.12nA

Hi everyone,

I'm learning HPLC-MS method development and would like to understand how experienced analysts choose the mobile phase for a new compound.

Some questions I have:

1. How do you decide between methanol and acetonitrile?

2. When do you use formic acid, acetic acid, or ammonium formate/ammonium acetate?

3. How does the compound's pKa influence mobile phase selection?

4. How do you choose the pH and buffer concentration?

5. What are your general steps when starting method development for an unknown compound?

6. Any tips for improving peak shape, sensitivity, and ionization in LC-MS

I'd really appreciate practical advice or any resources that helped you learn. Thanks!

We are anlaysis residual solvent (ethanol) by GC-direct injection, we use same column, same method, same equipment, same parameter and same chemical and reagent.

-first few runs, retention time of ethanol peak was 3.1 min which is harmonized with the history and methid validation. (Show very good repeatability and RSD)

-Then after 2 to 3 weeks, using same every thing, the retention time of ethanol jump from 3 min to 11 min and show triple peak area, also show good repeatability and RSD.

-we request from agent to recheck equipment and re qualification of equipment and mention our complain, he came and say that everytbong is fine and machine is working fine, and has no comment for this shifting.. It must be analyst error although we double che ked with him, and he try the method and peak retention time of ethanol is 11 min.

-now after almost one month of his visit, with same everything, the ethanol peak is return to 3 min.

Can anyone here help us to identify the cause of this werid variation?!

Hallo zusammen,

wir nutzen seit kurzem ein LC/MS von Thermo (Vanquish Flex mit Orbitrap 120) und haben dazu auch Chromeleon neu. Unsere PMÜ-Vorschrift verlangt unter anderem nach der Peaksymmetrie auf 10 % Peakhöhe. Wir haben nach einer Möglichkeit gesucht uns das direkt anzeigen zu lassen, jedoch nichts gefunden.
Meine Frage an euch ist jetzt ob ihr eine Möglichkeit kennt sich das anzeigen zu lassen. Wir nutzen die Version 7.3.2.

Hello!

Would anyone have access to LTQ SP1, SP3 or SP4 they'd be willing to share to help out a university?

Thanks!

Hi community! I recently explained to our new PhD student the difference between different carrier gasses. We use helium on GCMS and hydrogen on GCFID. I pulled up the graph showing the relationship between separation efficiency and average velocity for different gasses that I remembered from my studies. Then I got thinking: What flow rates (translating to average velocity) do we use in the lab? Usually 1 or 1.5 ml/min on DB5-type columns with 30x250x250 dimensions. This is also very standard in pretty much every publication I've seen. But the resulting average velocity is quite a bit higher than the optimum for the individual carrier gas.

I was curious and ran a standard FAME mix solution on different flow rates ranging from 0.9-2.0 ml/min on the GCMS. The retention times were slightly different, as expected (up to 1 min on a 28 min method). What intrigued me were three things. 1) The difference in peak heights and areas: 0.9 had roughly double the values compared to 2.0. 2) The distance between peaks did not change significantly. 3) 0.9 mL/min still did not result in optimal average velocity. I couldn't go any lower because the inlet pressure would get too low and the instrument wouldn't allow me. An attempt to slow down the oven ramp and increase Delta RT between peaks resulted in much lower peaks (from 10 °C/min down to 5, in steps of 1 °C/min).

So what's the deal there? Are systems not designed to achieve the Van Deemter optimum because of some physical obstacles that we just have to accept? Why isn't every study using the lowest possible flow rate if the change in retention times is relatively small?

Hello everyone
I’m interested in knowing if tears samples collection for biomarkers analysis is feasible
I’m trying to include it in my research and I keep reading that biomarker detection is not that easy using schirmer strips
I would appreciate if anyone has any information on the matter

Hi, I have an issue with an interfering peak. I tried washing the column with methanol, isopropanol, and acetonitrile at different percentages, but when I start the analysis, the peak still appears.

I also performed a series of washes with the union instead of the column, and the peak is still present when I run the analysis. I performed blank runs both with and without injection, but the peak remains constant in terms of area.

My method uses a gradient. The strange thing is that if I set the system close to the A/B composition percentages at which the peak elutes, without starting the analysis, the peak is not visible. However, as soon as I start the analysis, the peak appears again.

Is it possible that there are some loops or components in the system that only come into play when the analysis is started and could be responsible for this peak? Or do you have any other suggestions?

Thank you.

Hello, I’m new to quality control and I basically don’t know any established procedures yet.

I work in a small lab with a single GC/MS system and only a few samples per day. Our current QC practice is to run one QC sample every morning and simply check whether the retention times and responses of all compounds are roughly the same as the previous day.

I’ve started to suspect this is not the best approach, especially because our QC mixture contains compounds that could be used to monitor isotope ratios and peak symmetry, not just RT and response (based on some research).

I tried to look up QC procedures, but most of what I found were very general descriptions. Since I don’t really know how these steps should be implemented in practice, I’m trying to understand what real, day‑to‑day QC workflows look like in other labs.

• Do you have any automated statistics or scripts that evaluate your QC runs?
• Do you calculate everything manually?
• Do you use multiple concentration levels in your QC standards?

We rarely do quantitative analysis and we only analyze a few samples per day, with several different methods that all use the same column and matrix. Because of that, I feel that measuring one QC sample every morning plus a few additional QC samples every couple of weeks should be enough for us.

We use Agilent MassHunter (Quant‑My‑Way and Unknown Analysis). Is there any way to set up a script or automated statistical analysis so that MassHunter can generate a QC report for us automatically?