A major challenge posed in the assessment of risk at Superfund sites is our ability to detect, quantify and determine the structure of environmental compounds and their metabolites. Precise structural identification of chemicals is crucial, because chemical structure dictates biological and toxicological activity. Moreover, accurate quantification is important to assess hazard and estimate risk. To this end, we are developing and applying a new, highly specific and sensitive mass spectrometry technique to detect and quantify environmental compounds and their metabolites, including those of tetrachloroethene, 1,1,2-trichloroethene, cis-1,2-dichloroethene, vinyl chloride, chloracetaldehyde and chloroform.

We have successfully developed and demonstrated a new instrument for resonance electron-capture mass spectrometry, the gas chromatograph electron monochromator time-of-flight mass spectrometer (GC-EM-rTOF-MS). Many of the technical problems common to standard instrumentation are avoided with this new instrument, including the generation and moderation of electron energies and all the attendant problems of irreproducibility and spurious ion production that result from ion-molecule reactions. With the EM-rTOF-MS, three-dimensional negative ion electron capture spectra are recorded in real time in an interval of approximately 1 second, as opposed to the several days that are required to record a complete spectrum with an EM-quadrupole or EM-magnetic sector mass spectrometer.

We have also advanced our work elucidating the molecular structure of neurofilament proteins (NFP), which are structural elements in the neuronal cytoskeleton. NFP are highly phosphorylated, a condition that is believed to be responsible for their assembly and stability. We have used mass spectrometric sequencing to determine the N?terminal sequence of bovine neurofilament-M (NFM), which was previously unknown. Moreover, we have shown for the first time that another bovine neurofilament, NFL, has three phosphorylation sites, while NFM has twenty-two defined and two tentative phosphorylation sites. Most of these sites are homologous to those previously identified in other mammalian NFM. We also found other structural modifications as well, including deamidation, oxidation, N?terminal acetylation, and pyroglutamic acid formation.

Environmental chemical pollutants can produce toxic or other undesirable effects and are, therefore, a major public health concern. These chemicals, which consist mainly of chlorinated compounds, nitro compounds, polyaromatic hydrocarbons and organophosphates, are strongly electrophilic, rendering standard methods for their detection and quantification relatively inaccurate. The GC-EM-rTOF-MS instrument will thus provide greater sensitivity, specificity and reproducibility than do standard methods. Furthermore, the design of the analyzer and source are compact so that the instrument can be engineered for portability and act as a “chemical sniffer” for dump sites and other polluted environments.

Studies in experimental animals show that hexane and related compounds are metabolized to 2,5-hexanedione by the P-450 system, causing neurological problems, cell damage and death. The side chains of lysines in NFPs are thought to react with these neurotoxins, which causes cross-linking of the proteins. Sites of phosphorylation on NFPs must be identified before any progress can be made on mapping the 2,5-hexanedione-adducted peptides. This has now been achieved for NF-M and NF-L.