Loss of Signal Intensity in MS after cleaning the ESI Source

I´m working with an UPLC-System coupled to a Quattro Premier XE. Everytime the ESI-source (sample cone, cone gas cone, ion block and it´s compounds) was cleaned I had problems to get better values for peak area and peak height in comparison to pre-cleaned level. This Problem is also observed on the routine MS (Quattro Micro API). Recently I cleaned just the sample cone and the cone gas cone just because it was obviously contaminated. After replacement and about 1 hour re-equilibration I performed an UPLC-run seperating two substances. The same sample was the last run before cleaning the cone. And now I found just half of the area compared to the level before cleaning. But this only apears for one of the two substances. Substance stability can be excluded as the cause. Are there any ideas?For an earlier cleaning procedure I contacted Waters and they told me to tune the MS with the set-up solution (cat. no. 700003105) before and after cleaning. I tuned the solution by direct injection with 100% MeOH at a flow rate of 0,05 ml/min. Than I cleaned the source and also the probe tip. After replacement I tuned the set-up solution and adjusted the probe tip for maximum signal intensity. The values after cleaning were 2 fold increased for the set-up solution. But the next UPLC-run gave peak areas about 50% compared to the values before cleaning. I readjusted the probe tip but that was time consuming. Is there any trick for the adjustment of the probe tip? And how is it possible to get less peak areas even though tuning with another solution gave increased signals?

Answers

  • Hi Johannes,

    A problem with loss of sensitivity is usually caused by a dirty cone, very rarely it is the extraction lens and ion block unless buffers with poor volatility ,ion pair reagents or high concentrations solution infused directly into the cone.

    You are right to test the sensitivity before and after cleaning with something simple like the setup solution.

    It is probably best to do this with direct infusion with a syringe rather than an injection as this will only bring the injector into the equation. Do the infusion of the setup solution at 5ul/min (source temp 80C desolvation temp 150c desolvation gas flow 350 and no cone gas) and bring up an ion at a similar MW to what you are analyzing. (The solution contains PEG so a number of ions will be available over a range of 175-1080) confirm the sensitivity at a span of about 5 and print the tunepage. You should be able to return to this test any time in the future to access how the system is performing.

    The most common cause of sensitivity loss after cleaning the ion block is poor vacuum caused by badly seated "O" rings or failure to tighten the large screw with the teflon washer on the ion block. Also it is often good practice to replace all "O" rings when cleaning as they often lose their elasticity from sitting in the hot source and will not seal well as second time. The valve cock for the vacuum on the ion block can also leak if incorrectly re-build and adjusted.

    A good tip for adjusting the probe position is to place a 8mm allen key under the edge of the block and turn the adjuster till the allen key will just slide out. This will place the end of the cap about 5-6 mm out from a straight line down the cone entrance. Newer systems a fitted with a micrometer adjustment, one day I will add a tutorial on how to set and read a micrometer.

    PS every time you remove the probe inspect the capillary tip with an eye loope to make sure it isn't damaged or burnt, it will protrude about half the diameter of the tube it runs thru if correctly set.

    Hope this helps.

  • First I want to thank you for the quick reply. A Waters-Engineer came to us for the routine MS (Quattro Micro API). He supposed contamination to be the cause of the problem. He had taken a look to our source compounds and told us what we have to clean again. He also gave advice how to clean every part. So we carfully cleaned all compounds with Aluminium oxide as a scouring agent. After extensive rinsing with water all parts were placed in the Ultrasonicator with 100% MeOH and than 100% ACN. After replacement there was no improvement in performance, but the set-up solution had much better intensity since last time infusing it. So were checking the UPLC to be the cause of the Problem. We are flushing the column with water, MeOH and ACN seperatly. We also want to collect the fractions of the chromatography and compare the unseperated sample with the fractions by direct infusion.

    Maybe anyone has an idea how the UPLC can cause a loss of intensity in the MS?

    PS the O-rings were replaced and the ESI-Probe was adjusted with the micrometer adjustment on the day of installation and never varied since then.

    I am going to post results of column cleaning and fraction comparison today afternoon.

  • You must very careful what you use to clean source parts, the use of abrasives is very bad unless the source parts are already a "write off".

    The surfaces of some of the components have a satin like finish (cone and extraction lens) that has been done on purpose and impacts on performance.

    The cone gas cone can be cleaned with the yellow lapping paper that is supplied in the source start-up kit but it should not really be used on anything else.

    I have used 0.05micron polish on a very dirty hexapole once but I repeat most cleaning can be completed with normal solvents and an ultrasonic bath.

    In extreme cases use the "magic mix" we use to clean the UPLC ( IPA/MeOH/CH3CN/1%Formic in equal parts or in some cases 85% formic).

    I assume that this method was running ok before cleaning? i.e. All parameters for injection mode, volumes etc for the UPLC were OK.

    Loss of sensitivity (poor signal to noise) is caused by poor signal or high noise!

    I assume that with an infusion of start-up solution your signal is good?

    Then then cause must be too much chemical noise.

    Try removing the column and look at the background on the tune page (MS1) , if you open a window about 300 amu wide centered around the mass of you compound. The signals should be less than about 10% of scale at 128X. If the background is higher identify the masses and try to identify the source. (If you can't find it I wlll send a common contamination ions spreadsheet that Waters have)

    A system wash can be performed without the column using procedures found elsewhere on this forum, this may reduce your background. In all cleaning use the best quality solvents and water as I have often seen contamination introduced by the cleaning solvent. (Using solvent in a wash bottle to rinse off a cone after cleaning or using oil contaminated aIr to blow it dry are common causes.)

    Good Luck!

  • We had a maintenance by a service engineer. Probably it was the wear of all the scource spare parts (i.e. capillary, cone...).

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