Analytical Separations, Capillary Electrophoresis, Capillary Liquid Chromatography
Ph.D.,Indiana Univeristy (1979); B.S., Northern Illinois University(1974); Fellow of Academy of Arts & Sciences (2007), American Chemical Society Award in Analytical Chemistry, 2007
Research in our lab concerns harnessing the power of techniques such as capillary electrophoresis (CE), capillary liquid chromatography (LC), and mass spectroscopy (MS), to solve problems in complex mixture analysis. Samples of biological and environmental origin are usually complex in character, containing many hundreds to thousands of detectable compounds. Mixtures of such great complexity are a challenge to the resolving power of existing analytical methods. For an analytical technique to successfully handle such samples, the technique must have either extreme selectivity for a particular compound of interest, or the technique must provide a great excess of resolution space (peak capacity) for the compounds to be resolved into. We are currently pursuing techniques of great peak capacity, such as combined liquid chromatography-capillary electrophoresis (LC-CE), as well as techniques with extreme resolving power, such as flow counterbalanced capillary electrophoresis (FCCE).
Combinations of chromatography with mass spectrometry have long been recognized as having the requisite selectivity to permit effective complex mixture analysis. We are exploring the possibilities of combining two separation columns to provide much greater peak capacity. Combinations such as LC-CE and LC-LC are being developed for this purpose. These combinations of two separation techniques are done in what we refer to as a "comprehensive" manner; all components of the sample are subjected to full two-dimensional analysis. Comprehensive LC-CE has generated peak capacities in excess of 20,000 peaks. Such systems permit effective analysis of mixtures containing thousands of detectable compounds. We are also initiating work on LC-LC-MS and LC-CE-MS as systems of extraordinary power for analysis of complex mixtures. The combination of these two-dimensional separation systems with mass spectrometry should permit the further resolution of compounds, and aid greatly in the identification of compounds.
Flow-counterbalanced capillary electrophoresis is a technique that takes advantage of a unique property of electric field driven separations; that the migration of species of interest may be balanced against a counterflow of solvent, thus allowing the separation to continue to develop and improve for an unlimited time period. In general, the resolution of a particular compound from its neighbors improves in proportion to the square root of the time the compound has undergone separation; i.e., to double the resolution requires a four-fold greater amount of time. With the flow counterbalanced approach we can subject the compound of interest to electric field driven separation for an indefinitely long period of time. We have demonstrated separations with tens of millions of theoretical plates. If conventional CE had been used, it would have required a capillary 30 meters in length, and a driving potential of over 3 million volts.
An additional unique capability of CE and capillary LC is their ability to handle sample volumes in the nanoliter to picoliter range. This coincides well with the volume of single cells. We have been using capillary LC to investigate the composition of individual neurons and individual cells from cell cultures. In these studies, we have been able to determine that cells from the adrenal gland are clearly of two types; those which store and secrete adrenaline, and those which store and secrete noradrenaline.