Agilent 1260 Purging Lines

[rfazily]

I've got a question regarding the procedure for purging lines in an Agilent HP system before washing column. This is how I would do it. Advise on changes you would make.
I have a run where I have to use only lines A & B. Flow rate is 1.0ml/min.

1. Open the purge valve
2. Put flow rate to 4ml/min and put % flow as 25% for each line A,B,C,D
3. Let it run for 15 minutes
4. Turn flow to zero
5. Close the purge valve
6. Individually wash the column at 1.0ml/min with Mobile Phases in A and B for 20 minutes each

Any comments or suggestions that I can do to make the process better for my run?

[Consumer Products Guy]

Early on, when the 1050 series was introduced, we had some issues with the quaternary multi-channel mixing valve when only using two solvents. HP technical (before it became Agilent) recommended purging each channel separately for 5 minutes at 5ml/min to avoid any issues.

Even though this was before these had purge valves or vacuum degassers, our company continued this process to avoid problems. When I retired, we had like 7 Agilent HPLC systems in R&D, still did that each day of use.

When we only used one channel (like for RI or conductivity), we bypassed the multi-channel mixing valve completely.

[uzman]

When purging all channels together , miscibility of the solvents must be taken into consideration.

[James_Ball]

As long as the solvent in each channel is miscible then you should be ok. Just make sure one channel doesn't have a high buffer concentration otherwise when it mixes with 100% organic from another channel it could cause the salt to fall out of solution.

Either way, the one thing I would add would be to run at 100% for a few minutes on the first solvent you are going to put through the column to flush the line from the mixer through to the purge valve, if you don't then the column is going to be hit with a mixture of all four channels as there will be some of each left in the lines and pump. Letting it flush with what you want going through the column will remove any of the mixed solvents from the system dead volume.

[Multidimensional]

Always prime one line at a time (100% A, 100% B and so on).

Be sure to account for the contributing volume of any low-pressure tubing PLUS any vacuum degasser channel. So many people are unaware of this and never flush out enough solvent when starting up the system (or changing mobile phase bottles). Extremely common problem as we get a few calls each week related to this. Then they wonder why their results keep changing over the next day or two until the old solvent is finally flushed out!

The minimum flush / prime volume needed will be different for different HPLC systems. It may even be different for what appears to be identical HPLC systems (because they may use different degasser models or designs). Some systems may only need 12 mLs per channel to flush one channel, while others may need 50 mLs. You need to look at the specification of your instrument and how it is connected up.

Here is a link to a free article with more tips and help:

"Tips and Advice for Priming your HPLC PUMP (or similar pumps, FPLC or UHPLC Pump)", https://hplctips.blogspot.com/2020/09/tips-and-advice-for-priming-your-hplc.html

[Multidimensional]

Another trick that my boss taught me, that we teach in our HPLC training classes, is that for AGILENT HPLC pumps, specifically the QUAT pumps which have a flow range up to 10 mL/min, you want to use that extra flow range to flush the lines 2x faster than on a binary pump with a 5 mL/min max. Open the prime-purge valve. Just set the max pressure limit on the QUAT pump to 200 bars and then the system will allow you to enter FLOW = 9.999 (which the system will understand to mean you want 10.000) and the flow will be 10 mL/min ! Flush down one line in half the time. We use it all of the time in the lab (every day when we start up the systems) to quickly blow out the lines with fresh mobile phase.

[Consumer Products Guy]

We set up a "Method" to run each channel for 5 minutes at 5ml/min so this was just done automatically.

This needed to be extended for 1050 degassers because of the larger dead volumes with their degassers.

[DavidHPLC]

Another trick that my boss taught me, that we teach in our HPLC training classes, is that for AGILENT HPLC pumps, specifically the QUAT pumps which have a flow range up to 10 mL/min, you want to use that extra flow range to flush the lines 2x faster than on a binary pump with a 5 mL/min max. Open the prime-purge valve. Just set the max pressure limit on the QUAT pump to 200 bars and then the system will allow you to enter FLOW = 9.999 (which the system will understand to mean you want 10.000) and the flow will be 10 mL/min ! Flush down one line in half the time. We use it all of the time in the lab (every day when we start up the systems) to quickly blow out the lines with fresh mobile phase.

So I can just connected the "Game boy", set max pressure and enter "9.999"? I remember setting the pressure as suggested and then putting "10" and it didn't let me.

[Multidimensional]

David wrote: "I remember setting the pressure as suggested and then putting "10" and it didn't let me."

That is correct because you entered the wrong flow value number. As explained, you enter 9.999 and it will change to 10.000 mL/min and allow you to run at the higher flow.

[rycar]

When thinking of purging, what would be considered "old" solvent? In our systems we run on a daily basis and they often stop sometime during the night. When we come in the next morning, would it be necessary to purge the system of the mobile phase that is still in the lines before starting them up again?

[James_Ball]

When thinking of purging, what would be considered "old" solvent? In our systems we run on a daily basis and they often stop sometime during the night. When we come in the next morning, would it be necessary to purge the system of the mobile phase that is still in the lines before starting them up again?

It would depend on the mobile phase composition. If it includes volatile buffers like Ammonium Formate then probably so, if you replace it every day since it can lose the volatile component each night. Also if it is something them will degrade quickly. For normal Acetonitrile, Water or Methanol then maybe not.

If PFOA/PFOS compounds could interfere with the analysis then it is a good idea to flush the lines since those could leach out over night into the mobile phase from the teflon lines in the degasser. Also if water sits still in the lines for a long period of time it can allow algae to grow, but overnight you should be ok.