Chromatography Forum

Hello, Is it possible that packing methods which work very well with analytical size 4.6 mm i.d. tend to give much lower performance with microbore columns such as 2.1 mm i.d? What is the phenomenological basis of this apparent discrepancy?

Thanks.

Hello, the problem is the different linear flow in the column, so you will get a different packing density.
How to overcome the problem? Training, training, training.

Farooq, are you evaluating the 2.1-mm i.d. columns on an HPLC system that's been set up for 4.6-mm i.d. columns? If so, then peaks from the 2.1-mm i.d. column will exhibit severe fronting, due to diffusion into the excessive extracolumn volume. The column itself may be fine. The solution is to increase the flow rate - thereby reducing the time available for diffusion - or to use shorter connections with narrower i.d.'s. Also, use a detector cell with a volume of 1-2 µl instead of the more usual 8-9 µl.

Farooq, are you evaluating the 2.1-mm i.d. columns on an HPLC system that's been set up for 4.6-mm i.d. columns? If so, then peaks from the 2.1-mm i.d. column will exhibit severe fronting, due to diffusion into the excessive extracolumn volume. The column itself may be fine. The solution is to increase the flow rate - thereby reducing the time available for diffusion - or to use shorter connections with narrower i.d.'s. Also, use a detector cell with a volume of 1-2 µl instead of the more usual 8-9 µl.

You are correct. An ordinary instrument can halve the plate count e.g. an ordinary HPLC gave 6000 and an optimized system gave 14,000 plates for a 100x2.1 mm i.d (home packed column). Still the reduced plate height is not near the perfect one (~ 2). The instrument is in a perfect condition for minimum extra-column effects (< 5% contribution to peak variance).

Have you observed the same thing that 4.6 mm id column can be packed very well but same approach does not work with 2.1 mm i.d? Is it just the linear velocity problem as Gerhard just said?

Thanks.

Gerhard is correct.

Gerhard is correct.

Thanks. I noticed that there is another school of thought which just thinks the opposite. They use ultrahigh pressures to pack microbore and capillary columns (e.g. 25,000 psi to 60,000 psi). With normal packing pressures it seems that the wall area is too tightly packed and the central core is loose in narrow i.d. columns. This gives rise to severe tails since the wall is so close to the injected band in a narrow column.

There are alternative ways to accomplish that. There's a lot of trade secrets in packing columns, Farooq.

I might note that narrower columns are never as well-packed as wider ones. The narrower the column gets, the greater the percentage of the total volume that's near the walls. Wall effects are real.