Our research is focused on the application and development of mass spectrometry techniques and instruments for clinical research. I have a specific interest in developing approaches to small molecule quantitation or characterization using mass spectrometry which would better understanding diseases, drug treatments, and therefore health. This interest involves the use and development of both imaging mass spectrometry (IMS) to directly probe excised tissue sections for chemical distributions and traditional liquid chromatography tandem mass spectrometry of (UPLC/MS/MS) homogenized tissue and plasma or serum. We have been at the fore-front of developing instrumentation and applications for IMS studies and plan to continue developing this technique to provide high-throughput analyses and better identification of unknowns.
Critical to many small molecules studies in clinical research is the development of quantitative multi-analyte assays that are focused on biochemical pathways (Targeted metabolomics). These multi-analyte assays require both innovation in UPLC and mass spectrometry to separate and identify low level entities in clinical specimens as well as statistical correlations necessary to show what may be changing. A key component of this research objective is the integration into global metabolomics studies with a proposed Metabolomics Center at UF. The targeted metabolomic aspect will develop assays based on biomarkers disocovered from large scale global assessment of metabolism with the final goal of developing new assays that would be translated to a clinical mass spectrometry lab in Pathology.
One example of a multi-analyte assay we have developed is for metabolites of tryptophan. This method analyzes for tryptophan, kynurenine, kynurenic acid, and anthranilic acid and relies on the use of selected reaction monitoring (SRM) on a triple quadrupole mass spectrometer. The assay was applied to a clinical study group in pediatric orgran transplatation, which comprised 25 consecutive subjects with time points taken every month for 1 year post-transplant. Serum kyn/trp ratios were significantly elevated in the group that experienced acute rejection within the following 30 days (mean ratio 8.66+ SE 1.93) compared to the other two groups (ratio 5.69 + 0.29 in stable group and 6.84 + 1.19 in major infection event group, p value = 0.02). In contrast, urine kyn/trp ratios were not significantly different between groups (mean 15.1 + 0.9 stable group versus 17.7 + 2.6 rejection group versus 15.1 + 1.3 infection group, p value = 0.64). Because the method measured additional analytes of the tryptophan pathway (kynurenic acid and anthranilic acid) in urine as well as plasma, we identified an unknown marker appearing only in the urine of patients who exhibited a cytomegalovirus (CMV) infection. In addition, since we were monitoring the patients on a monthly basis, the clearing of the injection was also assessed, showing a removal of this unknown peak after clearing of infection. This shows the potential of targeted metabolomics to uncover new biomarkers and associations in a clinical study group.
In moving forward with high-throughput applications in UPLC/MS/MS, it is also critical to assess the potential of green alternatives to traditional higher flow approach. Instead of typical high flow rates (>200 uL/min), we evaluate nanoflow (<1 uL/min) and microflow (>1 uL/min) studies in connection to nanospray ionization for their use in clinical chemistry. In addition, the lower flow rates provide for the use of gas-phase separation modules such as ion mobility or high-field asymmetric waveform ion mobility (FAIMS) that offer the potential to remove the UPLC component for some analyses.