HPLC contamination

[wenchenluo]

Recently, we observed one HPLC peak shifted retention time due to chelation to copper(II)(confirmed by LC-MS). After washing LC with 0.5% phosphoric acid, the copper(II) complex peak was gone. But after several injections, the chelation peak reappear. We washed system with 6N nitric acid, but doesn't seem to work. The mobile phase we are using are 30 mM ammonium acetate (pH 5.3) and acetronitrile. I wonder if anyone had similar problem before. Thanks.

Wenchen

[Judy-Vancouver]

yes, this problem usually will have when you use a column with metal sift. You could use a column with peek sift to avoid this problem

[wenchenluo]

Thank you very much, Judy. The "sift" you mentioned is the frit on both end of the column, right? Do you think the tubing inside HPLC could be the problem too? We tested the same method on new column, but still have the copper contamination. I wonder if we should change the tubing as well.

Wenchen

[JA]

Aren't all the metallic components of a typical column 316 stainless steel? (If so, there's no copper in it).

[irene_wu]

hi:
one article I read is about to clean and regeneration of hplc columns:
it says some column silicas can interact with metal chelating or scavenging compunds. and these undesired interferences might be observed by a detector and appear as chromatographic peaks , retention times shift and tailing .... ....sometimes , washing with organic solvents can fail to remove the column contaminants. this situation is particularly true if metallic ions are sorbed to the silica or bonded phase. A chelating reagent such as 0.05M ethylenediaminetetraacetic acid (EDTA) can be flushed through a column. The EDTA complexes with many metallic specis and solubilizeds. them . after treatment with and EDTA solution, analysts can wash the column thoroughly with water...

if you feel this helpful. you could find them from January 2003 LCGC NORth America voume21 number 1