HPLC Passivation with Nitric Acid & any unwanted side effect

[warlock4761]

Hi,
I recently started using a passivation technique on Agilent 1100 and 1200 series hplcs. I have checked with my Agilent rep and they gave me a recommended procedure also stressing the importance of removing the degasser and detectors from the flow path (and obviously using a column blank rather than a column). The procedure I am using goes something like this:
1. Purge at 5mL/min with water for 5 mins on both channels (binary pump)
2. flush system with water at .5mL/min for 30 mins (so far just to clean the system)
3. purge both lines at 5mL/min for 5 mins using 20% nitric acid
4. Flush system with 20% Nitric acid and .5mL/min for 30 mins
5. Repeat water rinse (steps 1 & 2)
6. Purge with 12% ammonia, flush system with 12% ammonia
7. Repeat water rinse (steps 1 & 2)
8. Purge with 1% formic acid, flush system with 1% formic acid
9. Purge with water, leave system flushing with water at 0.2mL/min for around 48 hours, until pH of effluent water is neutral.

I've carried out this procedure several times with no hiccups however recently a colleague ran a method with a triethylamine mobile phase (specifically pH 2.5 triethylamine buffer as the aqueous mobile phase and acetonitrile as organic mobile phase). On injecting a blank/diluent a large peak is eluting just after the injection peak. One theory is that the triethylamine is reacting with the oxide film provided by the passivation technique and is being released on injection.
I'm not quite sure how to troubleshoot this, so far I've flushed the system with organic solvent and cleaned the injector both with buffer and organic. Have also changed the PTFE frit at the purge valve and the glass in-line filters were not used during the passivation.

[tom jupille]

Isocratic or gradient?

And does the peak get smaller with successive injections?

[warlock4761]

Been troubleshooting the issue. Peak is present even when injecting water while running water as mobile phase, therefore it is a contaminant in the system and not related to the method. I started by injecting large volumes of water (100uL) and the peak size reduced, which lead me to suspect that the contaminant was somewhere in the injector/rheodyne valve.
I washed with water for a long period of time with the flow set to "by pass" so as to rinse the flow path which i hadn't done previously and this seemed to reduce the peak size quite significantly.

However the peak is still present! Next step is to inject water repeatedly at 100uL volume in the hopes that whatever is lodged in there will finally be purged out.

[tom jupille]

Tedious, but that *does* sound like the best plan.

[LCbob]

Hi
In these instances I put on a bottle of 50:50 IPA/Water and make 100 injections of 50ul on a 0.75min run time.
Come back in the morning and check. If you still have problems change the seat, stator, needle and rotor seal, and retry.
If this doesn't work repost and I will explain how to start removing some of the injector tubing and metering pump to try and identify what is contaminated.

[warlock4761]

After many injections with 50% IPA the peak has diminished to a negligible absorbance. I'm hoping that over the weekend, left cleaning at low flow with water, and with some use it will disappear.

Still not sure what it could be, best guess is that it was so gunk that got washed out with the passivation procedure and with held within the injector.

If anybody has experience with the passivation procedure I mentioned above and some helpful tips (or experienced unwanted side effects) please let me know!

thanks,
Warlock

[rhaefe]

I have done similar passivation procedures in the past (you can look in the archives of this forum).
One thing I always did: I passivated the injector as well. Just make sure that all components can handle the nitric acid. In my case I had PEEK rotor seals, Teflon needle seals and so on.
It was important to get the nitric acid completely flushed out. Since the flow cell of our UV detector was able to handle nitric acid without any problems I monitored the rinsing/flushing process online with UV detection. Using a water soluble buffer at pH 7 (like phosphate) speeds up the whole thing.

I still have the original passivation procedures from Agilent (for the HP1090), Waters (WISP autosampler) and Merck (=Hitachi system) somewhere stashed away...

Thanks,

Robert

[Chromatographer2010]

I have seen in the past that PEEK rotor seals will "express" a plasticizer contaminant when exposed to a very strong acid. I would probably suspect any PEEK component would have a similar issue. The strong organic wash you used is similiar to what I used when getting rid of it. Upchurch, sorry IDEX, and I think most manufacturers recommend that you avoid concentrated nitric acid when using PEEK.

[rhaefe]

I've had no problems with PEEK and nitric acid (at room temperature that is). Some manufacturers of PEEK state that it can safely be used with nitric acid up to 30, 50 and even 60% (the concentrations I used for my passivation procedures ranged from 30-50%). See here:
http://www.vici-jour.com/support/resistance.php
and here:
http://www.zeusinc.com/technicalservice ... tpeek.aspx
The chemical resistance chart on the Nalge-Nunc website states that PEEK tubing can handle 9.52M nitric acid (at 20°C) for 30 days constant exposure without any change. That is about 60% nitric acid.

But as always: Heed the advise of the instrument manufacturer!

cheers