I knew that THF is selectivity wiser in HPLC system and THF rather dissolves many things (right?). This is my first time to involve with THF as a mobile phase, so please advise me about it.

I want to elute and analyse compounds that strongly absorbed by C18 column.

I want to ask rather in theory. Does any know that if the compound can not be eluted with 95% acetronitrile : 5% water (or even 100%acetronitrile) from C18 column, can the 50% acetronitrile : 45% THF : 5% water (or 50%acetronitrile : 50% THF) ?

I want to know more theory about THF because I had some problem with no peak of the analyte came out, but I know that the structure of the compound has many benzene rings, so I think it should be stronly absorbed in column. I don't want to change to other column or method because I have not much experience (any advices will be good). So I decide to change mobile phase. Thanks

Sorry for the above question, I just tried to find out more information and have already known that polarity index of methanol , acetronitrile and THF is 5.1, 5.8 and 4, respectively, and also found in HPLC textbook which compares about mobile-phase strength of these 3 solvents. And the textbook also advise that if 100% acetronitrile cannot elute compounds from column, higher %THF should be required.

Anyway, any suggestions will be welcomed. Sorry again.

Some experience from practice:
A compound with lots of aromatic rings is an ideal example for the use of a phenyl phase instead of a c18 phase (more pi to pi interactions and other theoretic stuff I can not explain correctly).
A reversed phase seperation normally needs some conc of water within the solvents. Too low conc of waters results in loss of selectivity and nearly all peaks will eluate at the same time. Sure, it could be different with your kind of samples, but I guess it won't. Start your seperation with the the solvent you have in mind but also try some runs with increasing conc. of water.
Have you tried to solve the sample with some ml of acetic acid first? Acetic acid can be a stornger solvent than ACN or even THF.

Hi Anon-07:42:

THF can be a good solvent for HPLC mobile phase. But its cut-off is ~225 nm (it is more important) and the baseline could be ugly!!! If the analyte is aromatic rings type the response can be lost because there interference between the mobile phase and analyte absortion. These two points have to be consider.
THF can be increse the pressure of your system, too.
Good elutions!!!

In general, the only types of compounds that are difficult to elute from a C18 column at high concentrations of organic in the mobile phase are things like proteins, peptides (which can be challengeling at most conditions).

The suggestion of using a phenyl column is a good one. This or any other less retentive column (like a C8, C4, C2, or cyano) would greatly reduce the retention.

As far as solvents are concerned, you could use a mixture of acetontrile/THF, or acetonitrile/ethyl acetate, or even acetonitrile/acetone. The UV cutoff of these solvents are higher (330nm for acetone), but if your compound is a poly aromatic hydrocarbon, the UV absorbance maximum should also be at a higher wavelength. You might also consider 1,4-Dioxane. It is miscible with water, low UV cutoff (215nm), and strong eluent for a C18 column. The only drawback is the nastyness of dioxanes.

I haven't any experience with the idea that it is necessary to include a small amount of water to maintain some selectivity.

Good luck

Thanks for all comments.

First of all, sorry for coming to this forum again so late.

If anyone is still here, please tell me why adding acetic acid to mobile phase will help us to elute compound from column (as Anon 8:42 commented "Acetic acid can be a stornger solvent than ACN or even THF. ") or what is the purpose of adding acetic acid in mobile phase - as I saw many mobile phases often have acetic acid.

Hello Anon,

in low concentrations (< 1%) acetic acid is used as a weak acid. Differences in retention can be explaned most times with the acidic strenght compared with other acids.
When the conc of acetic acid is much more higher than usual (around 5% or more) one can assume that the influence of acetic acid within the seperation is not only that of an acid but also that of an organic solvent. Under such circumstances it is very likely that a normal RP column won't last for a longer time without been damaged from this mobile phase.

It is more common to use acetic acid in combination with other solvents to solve the sample. The mobile phase to seperate the sample on the column don't have to contain acetic acid. When either THF or acetic acid were necessary to solve the sample I tend to use acetic acid because it dissolves faster in the mobile phase.