Hi, everyone:

There has been discussion here on molecules that could chelate with residual metal ions in the HPLC system or the column. Assuming that the analyte of interest does form chelators with residual metal ions, what will show up in the chromatogram (tailing peak? Very broad peak? split peak?)? Thank you very much for your input.

It is extremely difficult to generalize with regard to your question. I have seen all three of the symptoms you described above as consequences of residual metal ions on the chromatography of chelating agents. Probably, tailing is the most common symptom but the deciding factor is the retention characteristics of the complexed form of the molecule. In some cases, the complexed form of the molecule will be significantly more polar than the chelating agent itself. This happens most often when the complex is 1:1. Assuming you're retention mechanism is based on hydrophobic interaction, you will observe a fronting peak in this case. However, often the complexing agent will form 2:1 or 3:1 complexes with metal contaminants. In this case, the complex will usually have a longer retention time (in reversed phase mode) than the complexed form of the same molecule. This will result in either a tailing peak, a broad peak or a split peak depending upon the relative retention time of the two forms. In addition to this, you may also observe multiple peaks with "bridges" if the formation kinetics are slow. Of course, if you retention mechanism is ion exchange or ion pair the deciding factor is the relative charge of the chelating agent and the complex. If the complex has a lower charge than the chelating agent, a fronting peak will result. If the net charge increases the result will be tailing, broad or split peaks depending on the relative increase in retention.

In my experience, the best strategies for preventing this problem are somewhat dependent upon the chelating agent in question. The simplest solution is to perform the separation at a pH which precludes formation of stable complexes. Another strategy which I have used successfully is to place a column packed with chelating resin immediately ahead of the injection valve in order to strip any metals out of the eluent. After placing such a column in line, it's generally necessary to inject the chelating agent at high concentrations repetitively to remove any residual metal contamination from the separation column. Dionex sells a column (MetPac CC1) specifically for this purpose. Also, it's best to minimize any metallic components in the column hardware, especially the column inlet and outlet frits.

Chris:

Thank you very much for your very thoughtful input. When posting the question, I had only RP in mind.

I still have two questions:

1. Is someone aware of any RP columns that have plastic column inlet and outlet frits? If yes, where to buy (It can be Dionex)?

2. I heard that Resteck has columns, the inside of which is specifically passivated to reduce metal ions and minimize chelatoing effects. Are there other supplies for us to get such columns?

Thank you in advance.

Jimmy,
There are many columns now available in PEEK format with various non-metallic or titanium end frits. You might start with Alltech, I know they offer a variety of prepacked and custom packed columns. You can reach them at 800-225-8324 or on-line at www.alltecchweb.com.

Regards,
Mark

There are also these SGE (Australia) columns which are classlined on the inside with non-metal frits.

Jimmy,

As far as sources of reversed phase columns with PEEK frits are concerned, in addition to the sources listed above I located a few others:
http://www.higanalyt.com/basepage1.htm http://www.sisweb.com/lc/agilent/column_system.html (apparently Agilent has a line of cartridge columns which use PEEK frits). In addition, you can get PEEK replacement frits from Upchurch(although these might not fit all column hardware): http://www.upchurch.com/
By the way, although titanium is widely touted as "bio-compatible" I don't consider it to be a great alternative. It may be more inert than stainless steel but it may still present problems in work with chelating agents. I've seen a number of rather unpleasant side effects of this so-called inert material.

Waters has PEEK columns, and any of their materials can be custom packed.