Chromatography Forum

I have an application which needs to run a gradient at 6ml/min. I have an option of a low pressure or a high pressure mixing system on a pump. What are the advantages/disadvantages of either system?.

The primary advantage of the high-pressure mixing design (HPG) is the reduced gradient delay volume compared to the low-pressure mixing design (LPG). This advantage is mostly helpful at low flowrates. The other advantage is that it does not need any degassing. The disadvantage of the HPG is cost; it has two expensive pump mechanisms. In your situation, the flow rate is high enough that the HPG advantage is probably not worth the extra expense.

The disadvantage to HPG mixing is that baselines are generally noisier at the extremes (95:5 or worse) than they are w/ LPG mixing.

Either way, degas.

I've been a chromatographer for a quarter-century. Some things don't change with time, do they? This high-pressure/lo pressure battle is as old as HPLC, and never seems to end!

I've sold HPLC's and I've bought HPLC's, so I guess I've seen both sides of this particular issue.

First - You always need to degas your mobile phase, irrespective of high-press or lo-press. High-press vendors claim 'there's no need to degas'. Truth is, if you don't degas, outgassing can and will occur in the worst possible place - your detector's flow cell. The flow cell, remember, is at atmospheric pressure.

'Back-pressure regulators will eliminate outgassing'. Back-pressure regulators do just that - they put a significant back-pressure on the flow cell, thus suppressing outgassing. Unfortunately, quartz flow cells can crack at around 350 psi. Makes more sense to degas your solvents in the first place.

So, you degas. High-press or lo-press, you degas.

'Lo-press systems cost less' - they don't.

'Lo-press systems have lower dead-volume.' True, but not really an issue if you're using a good dynamic mixer for gradient mixing.

In fact, gradient homogeneity with lo-press systems can be a problem. This is because of the proportioning valves that lo-press systems use.

Bottom-line: It doesn't matter which system you prefer. What matters is the results and separation you finally get. Give or take a few theoretical plates, you'll get the same results eventually, provided your separation chemistry is OK to start with.

I would not ask the question about advantages or disadvantages of either system. Today's technology permits either system to function equally well, without disadvantages. In both a high-pressure system and a low-pressure system, you need some mixing volume to eliminate ripples. This mixing volume is of no relevance in isocratic chromatography, but it does make a difference in gradient chromatography. In this case, its influence can be minimized by a delayed injection, if such a feature is available on the system. I could argue the case of mixing with a very low or very high percentage of organic modifier either way. So I do not see from this standpoint a significant advantage of one system over another.

There is a little kicker that occasionally may throw you off when you try to transfer a method from one type of system to another. In a low-pressure gradient system, the mixing is happening in front of the pump, and the pump will deliver the flow. Therefore you will always have a flow as accurate as the pump can deliver. In a high-pressure system, you mix the solvents behind the pump, and mixing solvents can created expansion or contraction. In RP it is usually some contraction. Therefore, your actual flow rate is lower than what you programmed it to be.

The consequence is that there may be some unavoidable differences between the retention times on a low-pressure-mixing system and a high-pressure-mixing system. For isocratic separations, they are nothing else but a percent or two in retention, and nothing in selectivity (the solvent composition is identical in high-pressure mixing and low-pressure mixing).

As long as one is aware of this little idiosyncracy, either system is fine.