I am new to the field of oligonucleotide(up to 30mer), nucleosides and nucleotide base analysis.
At present we use a range of C18 columns from Phenomenex,Waters, and Jones for the analyses.
The polar embedded phases seem to offer some useful selectivity differences from the straight C18 columns. I have noticed that the price differentials between manafacturers is very high, are some columns much better than others? Has anyone experience of these columns for our applications? Are their any other columns we would find useful? Advice would be appreciated.

Try the new ATLANTIS column from Waters. There is nothing comparable on the market for separation of highly polar compounds. Do not try to compare this column with those "Aqua" or so called AQ or other polar embedded columns. However, you can run those columns with 100% water or buffer, but you won 't get increased retention of your polar analytes. This is different with the ATLANTIS. You will get retention even for Uracil (3 or 4 minutes or more). In addition, the ATLANTIS is not more expensive than other polar embedded phases. I think this would be an option for your separation problem. Just contact your local Waters sales rep. Or just visit www.waters.com. They have some applications available, I think your analytes were among them. Hope this can help a bit.

F. Poetsch, Ph.D. PharmD.

Do not confuse the presence of "embedded polar groups" with "polar compound retention." That is a fallacy. Highly polar compounds (like nucleosides, nucleotides and nucleic acids) do not retain longer on columns that contain stationary phases with embedded polar groups. A properly designed C18 phase will retain highly polar compounds much longer than an embedded polar group phase.

I agree with Dr. Poetsch. Go to the manufacturers' websites and look for separations of the types of compounds you are interested in. Your potential column should also be compatible to highly aqueous phases since the analytes you mentioned require aqueous or nearly aqueous mobile phases. Lastly, what type of detection are you using? All LC columns bleed to some extent. Embedded polar group columns' bleed is detectable by MS.

Good luck

To Anonymous, 2002/10/07

First of all I have to emphasise that the ATLANTIS packing has both advantages: no dewetting when running with 100% water or buffer and good retention of highly polar compounds. In addition this material does not show any bleeding in MS detection and does not show infinite retention of apolar compounds.
On the other hand, have you ever tried to get retention of about 9 min. with excellent peak symmetry of, let 's say, Adenin on "normal" C18 phases with MS compatible eluents?
Then perhaps you will know what I mean...
To my knowledge, you can call for an application booklet from Waters.
By the way...i am not a Waters Rep.

To: Anonymous II (October 8, 2002).

From Anonymous I (October 7, 200)

Thank you for your reply. I actually went to the Atlantis portion of the Waters website to see what was there. I was able to download some useful information. I also looked at the Atlantis columns separations that are shown.

You are correct, adenine is retained for 9 minutes with very good peak shape. In the past I've tried to analyze Adenine and it was difficult due to its sensitivity to unreacted silanols (and poor peak shape). Other chromatograms show that uracil can be retained as well!!

I think I'll try one of these columns. Thanks again.

Does somebody have have information about the chemistry of Atlantis columns?

I believe the Waters website has information on these columns. Have you checked it?

Perhaps Uwe can shed some light on this question?

On the Waters web site are only applications, nothing on chemistry. Not even the position in the Waters/Neue selectivity chart. From some of the applications I know it looks like a usual ODS column with good endcapping. But that wouldnt explain the high retention for polar analytes.

Call Waters and ask for the applications team, they would be more than happy to fax you the information you need. 800-252-4752. No, I don't work for waters.

Here is the story:
We were looking for a packing that can be run in 100% water and give maximum retention for vary polar compounds. One of the avenues that we explored was to get over the hydrophobic collapse hurdle that prevents the successful use of modern RP columns in 100% water, but without loosing out on the retention. We studied the retention and hydrophobic collapse
in water as a function of the packing properties. It turns out that a low C18 coating prevents hydrophobic collapse, but at the same time provides more retention than other alternatives. We also managed to do this with complete endcapping with standard trimethylsilane endcap instead of funny things. The end result of this is that you have standard hydrophobic retention with no strange side business working in a completely aqueous mobile phase.
Being an old advocate of packings with embedded polar groups, I was surprised myself that this approach works better - gives more retention - than the stuff with the embedded polar groups.

Now this is all very confusing. Having tried several polar embedded, etc., columns it was found that they indeed retained the polar compounds slightly more than normal ODS columns, with slightly better peak symmetry. While experimenting with one compound I got non-chromatography (multi-component peaks, spread over 10+ min) with MeOH from a different vendor (nothing else changed). I have not digested this completely yet and have to study this further, but: the polar embedded columns gave a much more severe failure (with the new MeOH) of the chromatography than normal ODS. The peak spread mostly toward the front (to lower retention time). This is almost not printable: Some peaks even spread ahead of the water dip (presumably the dead time). Maybe one should just do an ostrich head dip here.

Anyway, does anybody make a column with only trimethylsilane endcapping, no C4 to Cn?

Hans,
It is very confusing what you describe and what you are trying to do. Why do you want a C1 column? Too much retention?
Uwe

Where are all the messages that have been added since 14-Oct-02?

Anonymous,

Thanks for pointing this out. The Forum is missing a number of messages from the past nine days. We are checking into this. Please report any further disturbances in message strings.

Moderator
ChromForum

Moderator,

No problem. I posted a response (after Oct 14) and I know that there were other posts during this time as well.

Thanks for keeping on top of things.