Chromatography Forum

I am running a method with a mobile phase of 94% Acetonitrile and 6% aqueous (aqueous is 1% Phosphoric acid and 1% Triethylamine). Flow rate is 1ml/min with a C18 150 x 2.0mm column. The typical back pressure is around 110 bar but we have observed that when running on 2 different systems a gradual loss of pressure was observed until eventually the pump hit the pressure minimum (10 bar). Both were run on Shimadzu LC10AS pumps and the mobile phases were degassed by filtration. As soon as the pumps were purged the pressure returned to normal but would again drift over several hours.

Any ideas on this?

Do you premix mobile phase or do you mix online? Your problem sounds like online mixing on a low pressure gradient pump (though I am not familiar with the pump you mentioned) without online degasser. Degassing by filtration might be not sufficient in that case. I reached good results by putting the mob. phase in a ultrasonic bath and evacuating 2-3 times for about 10 seconds with a vacuum pump.

Thanks. The mobile phase is mixed and then degassed. Pump is single head isocratic.

OK, I might have been on the wrong track.
But it still sounds like a air-in-the-pump problem. The mentioned way of degassing or He-sparging (the safest and most expensive way ) might be worth a try, since ACN/water mixtures contain a lot of dissolved air. Does the pump "sound" different when the pressure drops? Do you observe bubbles in the waste line when purging the pump? Does the flow rate decrease with the pressure?

It would be a good idea if you measure the flow rate by collecting the column effluent for a given period of time at the end of the column in a measuring cylinder. This would give you a starting point for the diagnosis.

I agree. It does sound like a problem with outgassing of the mobile phase in the pump. Look for any evidence of bubbles entering the pump head. I once saw a system where the solvent tubing was routed next to the fan on the HPLC; the warm air in combination with low flow caused outgassing.

Thanks. The flow rate startes off ok but decreases with the drop in pressure. Unfortunatley the system does not have an on-line degasser or helium sparge. I did helium sparge the mobile phase on another system for 30 minutes prior to the last runand it has been fine for the past 12 hours. Roughly how long will a mobile phase stay degassed for?

Is your system a high pressure mixing system or a low pressure mixing system- you did not answer Klaus' question?

Personally I would not recommend running either type of system without some form of on line degassing but I think people do this with high pressure mixing systems. You obviously have a helium cylinder in your lab. Why not just move it next to the instrument and solve the problem?

..rather leave the helium cylinder where it is and splice the line and provide yourself with on-line degassing on both systems. It should not cost more than a few dollars to do this.

The system is capable of running high pressure gradient but we use a single pump to run isocratic for this method. The helium cylinder is specifically linked to another old HPLC system in another lab that is not capable of running this method.

If the planger stroke is ok then I think gradually your systems check valves ruby ball becoming trap by thinfilm of solvent. Resulting at last no movement of the ball.

Thanks

Placing the mp reservoir in an elevated position can help keep a pump primed.

J

Chris- I'm sorry I missed your previous line about single head isocratic.

Actually, now I'm not sure about this degassing business. I wonder why it should cause such problems with a pre-mixed solvent and acetonitrile as the modifier (methanol is much more problematic). Maybe John is right. You can also introduce air into your system if the frit on the solvent inlet is partially blocked-might be worth replacing it and see what happens.

Otherwise you just need a few bits of tubing and Swagelock fittings to supply your system with helium. We used to run 4 or 5 systems from a single cylinder.