Chromatography Forum

I am trying to diagnose a very strange problem that started yesterday on my 1100. For no apparent reason I started to have intermittent pressure loss during gradient runs. Pressure would be normal from 0% B to 55%B and then it would be lost during a 100% B buffer column wash. The pressure returned to normal at 0% B. This happened twice before I began trying to 'fix' it. This does not appear to be a degassing problem or air in the lines. I do not see any air bubbles going into the AIV valve.

A Buffer is 40 mM Phosphate pH 7.8,
B buffer is MeOH/ACN/water 45:45:10.
Normal flow is 0.5 mL/min

I removed the AIV and examined the cartridge. I'm not sure what it should look like but the body was about 2/3 white and 1/3 clear (empty?). I sonicated it in MeOH for 20 min and returned it to the pump. This did not solve the problem.

I then tried sonicating the outlet check valve in MeOH and that seemed to work for a while with normal pressure attained at 100% B, but when I tried to change mobile phase I lost pressure again. If I pump at 100% B, pressure will be lost for a while, then come up to normal, then alternate every 5-10 minutes between normal and no pressure. Eventually it will run steady at normal pressure for 30 min. The moment I change solvent composition I see a complete loss of pressure. Again there are no air bubbles going into the AIV. And solvent degasser seems to be fine.

Here are two observations that are confusing to me: When there is no pressure and I open the purge valve I can see solvent go forward and then retreat from the waste line. It's about 2 steps forward and 1 step back. I also ran the pump with the outlet check valve removed. The empty female joint gradually filled with B buffer but also did so in a 2 steps forward, 1 step back mode.

I'm guessing that for there to be reverse flow with no load (either purge valve open or outlet valve removed) that the problem must be the AIV. If I disconect the solvent line at the AIV I get decent solvent flow by gravity.

I have ordered a new AIV, AIV cartridge, outlet valve assembly, as well as pump seals. I'm hoping that these do the trick but I wondering what is the most likely source of the problem? Also, if it's the AIV, does replacing just the AIV cartridge sometimes solve the problem?

I am having other problems, perhaps due to bacterial growth in the phosphate buffer, that cause me to need to change the purge valve frit about every two months. The symptom for that problem is high back pressure, but I'm wondering if now if it could be causing valve problems. We are now trying to deal with that problem by not leaving the buffer on line for more than a week and by rinsing the degasser channels with 70% IPA/water about twice a month.

Has the instrument worked well with this method before? It seems to me there is a mixing problem with the buffer A and a 90% organic solution B. The buffer is precipitating out and thus no flow, ie no pressure.

Is this a quaternary pump?

I have not heard of sonicating an AIV cartridge before and believe it likely will not help. You may move the contamination around but it likely will not be purged/removed. From your description of the cartridge it sounds normal.

The highest wear part in the AIV unit is the cartridge so I just replace the cartridge until that no longer works, then replace the electronic actuator. The actuator lasts much longer than the cartridge.

One way to check the operation of the pump is to pull up ~5mm air gap in the tubing and watch it move. It should move forward, stop and move forward again. If the gap moves back toward the reservoir then the pump is malfunctioning. Do this test with 100% of one channel at 1-2mL/min.

For the outlet check valve I prefer to sonicate first with 10/90 MeOH/water to remove inorganic deposits then follow with 100% MeOH.

Flushing the degasser periodically with IPA is a good practice but adding 5% MeOH or ACN to your buffer will essentially eliminate bacterial growth for up to 2 weeks. Of course you'll have to account for the difference in % organic but this preventative measure will help reduce maintenance and headaches.

Thanks for the responses.

Yes this method has worked before and it's a quad pump.

I'm pretty sure we have had sodium phosphate crash in there. While going over things with my coworker it turns out their was a miscalculation and the A buffer he made was actually 80 mM, not 40 mM, so I think we did get salts to crash when we ramped from 50%B to 100%B. It certainly explains a lot. We now have a high degree of leaking from inside the pump head.

I'm going to take it apart, check the pistons and replace the seals, the outlet valve and the cartridge for the inlet valve.