Robert S. Brown
Associate Professor, Analytical Chemistry
B.S., 1978, University
of Massachusetts at Lowell.
Ph.D., 1983, Virginia
Polytechnic Institute and State University.
Postdoctoral Research Associate, 1983-1987, University of California at Riverside.
Our research is primarily concerned with studies aimed at improving the analysis of large, nonvolatile molecules by mass spectrometry. This involves research into fundamental ionization processes of large molecules and development of new instrumentation for mass spectrometry. Our principle ionization technique involves the use of high powered pulsed lasers (laser desorption) to effect ionization of a wide range of molecule classes. These include biologically relevant molecules such as peptides, proteins and glycoproteins, oligonucleotides, DNA fragments and synthetic commercial polymers. Mass analysis is carried out via time-of-flight (TOF) mass spectrometry. TOF mass spectrometers are well suited for pulsed laser ionization sources and provide both good mass resolution and mass accuracies. TOF mass spectrometers also combine very high sensitivity with an extended mass range for analyte analysis. Intact biomolecules, such as antibodies with molecular weights in excess of 150,000 are easily analyzed via TOF mass spectrometry with sensitivities typically in the femtomole range.
In order to both volatilize and ionize such large
molecules, a small quantity (typically 1-10 picomoles) of analyte (for example
a protein) is mixed with a large excess of a small
molecule "matrix" (10,000:1) and the two materials coprecipitate as a
solid. The matrix is chosen so that it can provide a source for proton transfer
to the analyte for ionization as well as being strongly absorbing at the laser
wavelength. For our work, cinnamic acid derivatives are one class of compounds
utilized. The mechanism for this matrix assisted laser
desorption/ionization (MALDI) process is still not completely understood,
but appears to involve the formation of a plume of matrix molecules which are
desorbed from the sample surface upon laser irradiation. The nonabsorbing
analyte molecules are carried along into the gas phase with the matrix molecules
as "free" ions which can then be analyzed by mass spectrometry.
Recent work in our laboratory has involved the development (see reference 7
under recent publications) of delayed extraction
techniques for MALDI time-of-flight mass spectrometry. This allows significant
improvements in the mass resolution obtainable with time-of-flight mass
spectrometers. A typical mass spectrum produced by this
technique from a peptide mixture can be found at this link. In addition to
the significant mass resolution improvements that on-axis delayed ion
extraction (first demonstrated by our research group in 1993 for MALDI) offers
for time-of-flight mass spectrometry, it also allows for the study of MALDI
metastable ions. In the case of peptides (B chain of
insulin example) and even some small proteins, metastable ions produced by
MALDI fragment along the backbone of the amino acid chain and provide a wealth
of sequence specific information (see reference 10 under recent
publications).
In addition to improving the instrumental aspects of
this technique and its applicability to various chemical and biochemical
problems, our research is actively engaged in more fundamental studies of the
underlying mechanisms involved. In particular, we are interested in determining
the exact role which the matrix plays in the ionization processes. This should
facilitate development of improved matrices for current applications and expand
the applicability of the technique to additional compound classes. Additional
fundamental questions which our research is addressing include: the possible
role of excited state species in the ionization process; the nature and source
of the ion activation processes that induce MALDI metastable ion decay; the
effect of the laser wavelength and pulse width employed on the MALDI process;
and the proton transfer processes involved in analyte ionization. This combined
approach of both fundamental and applied studies allows us to better understand
a very complex process, which, in turn, leads to tangible experimental
improvements to one of the most important new analysis method for large
biomolecules.
Some good mass spectrometry links:
The American
Society For Mass Spectrometry Home Page
Base-Peak
- Mass Spectrometry Resource by Wiley
Protein
Prospector - Protein Database Search Site
(Held in San Maleo, France in September 2000).
(Held in Estes Park, Colorado in September 2002).
(Held in St. Petersburg, Russia in September 2004).
(Held in Kifissia, Greece in September 2006).
Dr. Brown can be reached at:
(435) 797-0545 (Voice)
(435) 797-3390 (FAX)
