Chromatography Forum

how good/success rate to run NPLC with gradient? like 15 min gradient?
especially how good it is to do the scale up? like if i get a good analytical NP gradient method, how much probability i can scale it up on prep column by direct way, not changing it to isocartic mode?

by my limited experience with chiral analysis with chiral tech columns with Hexane/alcohol, which is NP mode, the short column (15cm) behaves pretty well with gradient, but i still have to switch it to isocratic when i do the prep.
(i never try the same gradient on prep).
so please shine some light on this.
the samples are small molecules.

Hi,
I am not quite sure what is the question exactly.
NP prep gradients are possible; The scaleup process is similar to RP scaleup. Flow should increase with the square of the column diameter.
Different "DwellTimes" (dwell volume / flow) might change your real (at column) gradient profile and said that also the separation. This becomes less important the smaller the dwell volume of the analytical system is.
Then there is an effect in np gradients you should be aware of: the more polar solvent is at the beginning adsorbed at the support, so the blank gradient profile doesnt look as nice and linear as the rp gradient profile.
hope that helps,


Alex

this is my understanding so far:

you are saying that NP scale-up is as straightforward as RP scale up, except the factor for polar adsorbents at the beginning, which means the gradient should start at lower percentage for scaleup?

my question is doubt about how effective to run gradient on prep NP column, since i haven't run that way.

normal phase in liquid chromatography is a very interesting subject.
I think your problem is about detection. Try TLC!!!

If you're getting good results on an analytical scale, there is no reason you should not be able to go to prep scale with NP. There are a couple of possible pitfalls:

- NP gradients are not truly "linear solvent strength" (NP solvent strength is closer to a logarithmic function of composition), so be careful about making adjustments to column length. If you scale flow rate to the square of the column diameter, as suggested above, you should be OK.

- If you're using and adsorbent (e.g., bare silica), mobile phase trace water content can have a significant effect on selectivity, and equilibration time can be a problem. If you're using a bonded phase column (e.g., cyano), then ignore the previous sentence.

any mathematical model or empirical formulas/guidelines for solvent strength prediction on NP?
thanks,