of Work: This project includes those endeavors in which the mass spectrometry workgroup collaborates with other groups, both inside and outside the Institute to solve problems of mutual interest. The major focuses of these projects are: 1) structure determination of unknown compounds; 2) identification and/or confirmation of biological pathways; 3) quantitation; and 4) development of strategies for the analysis of biologically important compounds. We continue to collaborate with D. Zeldin, LPP, on the quantitation of arachidonic acid metabolites in physiological samples by selected ion monitoring (SIM) under negative ion chemical ionization (NICI) mass spectrometry. As part of this work, we have characterized the epoxyeicosatrienoic acids formed by new P450s, CYP2Ns, found in Fundulus heteroclitus. The expression of these P450s was found to be regulated by environmental factors such as starvation or treatment with phorbol esters.We have developed a capillary electrophoretic method for the separation of apolipoprotein I and II and their isoforms in which 10-20% acetonitrile is added to the buffer. These conditions are compatible with MS detection in contrast to alternative buffer conditions we identified in which high concentrations of inorganic salts are used. Our results from the on-line separation of the isoforms coupled with mass spectrometric detection suggest that the multiple components observed in the CE separation appear to be oxidized forms of the proteins in addition to the native proteins. These results are in agreement with previous reports that the methionine residues of the HDL proteins are sensitive to oxidation, which in turn alters their lipid binding characteristics and secondary structure. Capillary electrophoresis using nonaqueous modifiers in combination with mass spectrometry, therefore, is a promising technique for screening and characterizing isomers of the plasma apolipoproteins.
