HPLC, positive/negative isomers

Direct analysis with the on-line ORD detector linked to the HPLC was hampered by peak broadening, which led to incomplete resolution of the two peptides. Diffusion in the large flow cell may be a major factor in this difficulty. Because detector response can be either positive or negative, it is important to have an excellent signal-to-noise ratio and baseline HPLC resolution of the peptide isomers to be able to quantitate the degree of racemization by this technique. While the on-line ORD detector is promising, further improvements in resolution as well as sensitivity are required for it to be useful for routine peptide analysis. 

HPLC with ESI produces mostly molecular ions and few to no molecular fragments. This limits the use of LC-MS in the differentiation of isomers. Two atmospheric pressure ionization (API) interfaces allow for the formation of molecular ion, [PAH]" , yet in general, derivatization and additives are required to induce fragmentation. ESI is an interface that transfers ions from the mobile phase into the gaseous phase for introduction into the mass spectrometer so that atmospheric pressure chemical ionization (APCI) can cause ionization of chemical species in the gaseous phase. Both techniques can be operated in the positive- or negative-ion mode. Reports exist for the ionization of PAHs by both techniques, although there are few actual applications to the analysis of real environmental samples. 

Fenthion, temephos and their degradation products were determined by HPLC-DAD and APCl-LC-MS in rice field water samples after SPE (Empore Cjg). Four fenthion transformation products could be identified in the positive or negative modes whereas the degradation of temephos resulted in six products. For both pesticides the oxo-analogue products predominate [366]. After rice fields had been treated with fenitrothion, crop waters were sampled and analysed by ELISA, LC-DAD and MS coupled by APCI interface. Eenitrothion, fenitrooxon, 3-methyl-4-nitrophenol and the s-Me isomer of fenitrothion were identified by LC-MS [367]. 

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