Advances in Technology
The Metabolic Profiler combines NMR, MS, and LC into one platform for metabolic profiling. Developed by Bruker BioSpin (www.brukerbiospin.com), the system provides complementary information when searching for a new drug metabolite or a low-level metabolite.
“A significant amount of the work is done by NMR alone,“ says Werner Maas, Ph.D., vp of R&D. He explains that a single sample can have an unknown amount of low-level metabolites. “Once you get to lower levels, around the 10-microMolar level, there are many metabolites that result in signatures that all overlap and add up as the NMR spectrum. Unraveling this information is a bit tricky.“
To do that, the signatures of individual metabolites need to be known. The company has made this information available through its recently launched NMR Metabolic Profiler Database. “Since the NMR signature changes as a function of pH of the sample, we characterize those shifts as a function of the compound. Once we know the pH of the sample, we can go back into the database and say, we have a pH of 6.3, so this is the spectrum we have to use to unravel the complete spectrum,“ Dr. Maas states.
The database contains endogenous metabolites, as well as metabolites from aspirin, Tylenol, and dietary components. Pharmaceutical customers can customize the database and add their own metabolites.
Waters (www.waters.com) also provides technology for metabolic profiling. Their UPLC (Ultra Performance Liquid Chromatograph) systems are designed to provide maximum sensitivity, resolution, and speed.
“We recommend using our UPLC and then using a time-of-flight mass spectrometer to provide accurate mass information about the samples. Then the data goes into our informatics system where it is extracted from the raw chromatograms. Various found compounds are put into a format where we can perform statistical analysis and see which ones could potentially be biomarkers,“ says John Shockcor, Ph.D., business development manager, metabolic profiling.
He stresses that the system does not find biomarkers. “People misunderstand that this is some type of blackbox technique for finding biomarkers.“
Dr. Shockcor says the company is attempting to tune its UPLC systems to tackle the challenge of lipids since they are involved in many metabolic disorders, such as heart disease and diabetes. “Were trying to make separation of lipids before they go through mass spectroscopy as simple as possible.“
Thermo Electron (www.thermo.com) is also advancing its MS technology. The LTQ Linear Ion Trap MS has three unique tools. One is its segmented ion trap where the center segment provides the purest field. “A lot of analyzers produce a fringe field, which isnt good,“ says Diane Cho, ion trap marketing manager. “You want to create the most homogeneous field to trap and analyze your ions.“ In addition, the ions are radially ejected into a dual detector. “This allows us to detect all the ions with no loss.“
The LTQ can be combined to high-resolution mass hybrids, the LTQ FT, and the LTQ OrbiTrap. “These provide high-resolution and accurate mass assignment on a chromatographic-scale without having to use internal standards,“ says Cho.
The system also provides cycle time to achieve high-throughput and high-sensitivity. “We have the highest MSn spectral quality,“ she states. “We have customers using this for MS to the tenth. Its important because you need to look at the fragmentation pathway of unknowns to build back to your metabolite.“ The system can also handle simultaneous quantitative and qualitative screening in one run.