Transformative effects of higher magnetic field in Fourier transform ion cyclotron resonance mass spectrometry

N. Murat Karabacak 0 1 4 Michael L. Easterling 0 1 2 Nathalie Y. R. Agar 0 1 3 Jeffrey N. Agar 0 1 4 0 Address reprint requests to Dr. J. N. Agar, Department of Chemistry, Brandeis University , MS015 , 415 South Street, Waltham, MA 02454, USA 1 Published online March 31, 2010 Received November 16, 2009 Revised March 4, 2010 Accepted March 4, 2010 2 Bruker Daltonics, Billerica, MA, USA 3 Department of Neurosurgery, Brigham and Women's Hospital, Harvard Medical School , Boston, MA, USA 4 Department of Chemistry and Volen Center for Complex Systems, Brandeis University , Waltham, MA, USA The relationship of magnetic field strength and Fourier transform ion cyclotron resonance mass spectrometry performance was tested using three instruments with the same design but different fields of 4.7, 7, and 9.4 tesla. We found that the theoretically predicted transformative effects of magnetic field are indeed observed experimentally. The most striking effects were that mass accuracy demonstrated second to third order improvement with the magnetic field, depending upon the charge state of the analyte, and that peak splitting, which prohibited automated data analysis at 4.7 T, was not observed at 9.4 T. (J Am Soc Mass Spectrom 2010, 21, 1218 -1222) 2010 American Society for Mass Spectrometry - important instrument performance metrics can be affected by factors with no- or convoluted field dependences. For example, the relationship between mass accuracy and resolution is blurred by anything that affects ideal peak shapes [1517]. There are isolated literature examples of what Marshall terms the transformative effects of higher magnetic fields. For example, post processing of 14.7 T FTMS spectrum to simulate a 7 T FTMS spectrum revealed that three compounds that were identifiable at 14.7 T appeared to be a single compound at 7 T [18]. In another example, the instruments sensitivity for a complex and highly charged sample (29 kDa protein dissociation products) significantly increased when going from 6 T to 9.4 T FT-ICR [19]. On the other hand, despite the availability of 15 T instruments, the highest published resolving powers of 8,000,000 (m/z 1148) [20], 17,000,000 (m/z 1084.5) [21], 1,000,000 (m/z 12,360) [22] were acquired (in one case 11 years ago [20]) at 9.4 T or 7 T, illustrating the importance of factors other than magnetic field. While the effects of field homogeneity upon instrument performance have been characterized [8], we are aware of no publication that systematically explores the relationship of FT-ICR MS performance and gross magnetic field. Here, we make an empirical determination of the merit of higher magnetic field, using the same instrument type, the same acquisition and processing software and methods, the same methods for optimization, the same user, and employing magnetic fields from 4.7 to 9.4 T. Moreover, we perform a meta-analysis of reported mass accuracies of FTMS instruments ranging from 1 to 15 T, which is consistent with our experimental findings. F spectrometry [1] (FT-ICR MS) is currently the ourier transform ion cyclotron resonance mass highest resolution mass spectrometry method. The cyclotron frequency of ions in a given magnetic field (eq 1) gives a very accurate measure of the mass to charge ratio of ions. Numerous performance parameters in FT-ICR MS are predicted to improve with magnetic field, including linear improvements in mass resolving power and acquisition speed, and higher order improvements in mass accuracy, dynamic range, kinetic energy, and peak coalescence [27]. All of these parameters combine to determine the figure of merit of a mass analyzer, but not in a manner that is easily predicable. Improving FT-ICR MS performance is critical in many fields such as proteomics, petroleomics, and MALDI imaging, and in many cases enables, rather than improves, analytical capabilities. In practice, however, a component with no a priori field dependence, for example field homogeneity [8] vacuum strength, or acquisition speed, which set fundamental limits on resolution [2], or a phenomenon with a convoluted electrical and magnetic field dependence such as peak coalescence [9] or phase locking [7, 10 14], can become the limiting factor and the determinant of performance. In addition, the relationship of All chemicals except ubiquitin (Boston Biochem, Cambridge, MA, USA) were purchased from Sigma-Aldrich (St. Louis, MO, USA). This study involved three separate commercial hybrid q-FT-ICR mass spectrometers (Bruker Apex-qe) with the same cell infinity ion trap geometry [23], electrospray source (Apollo 2), ion transfer optics, vacuum elements (with readings of below 4 10 10 mbar in the ICR cell during this study) but with different magnetic fields. A sodium formate (Sigma) 0.01 mg/mL solution was prepared in 50% acetonitrile 0.1% formic acid. A complex peptide and protein mixture with a concentration ladder spanning four orders of magnitude was prepared in 50% acetonitrile 0.1% formic acid and introduced by a syringe pump at 2 L/min. This mixture consisted of Substance P (1 M), insulin (0.1 M), orexin B (0.01 M), ubiquitin (0.1 M), angiotensin (0.1 nM), myoglobin (1 M), human Cu/Zn superoxide dismutase (0.05 M), ribonuclease A (0.5 M) and lysozyme (0.01 M). One M Substance P was used for ultra-high resolution narrowband experiment. Instruments were tuned and operated using our routine techniques and parameters, which involved the following steps. Initially, electrospray source and transfer parameters were optimized for maximum signal magnitude. Static trapping [24] at relatively low trapping potentials (1.0 V/0.8 V for 4.7 T, 0.9 V/0.8 V for 7.0 T and 1.2 V/1.2 V for 9.4 T for front/rear trapping plates) was used. These trapping potentials were 30% greater than potential where no signal was observed (and presumably little trapping occurred). Next, excitation amplitude was tuned for each instrument by determining the excitation amplitude that yielded maximum mass accuracy when internal calibration was performed using sodium formate clusters. Resulting amplitudes were 1.25 dB for 9.4 T, 4.5 dB for 7.0 T and 9.75 dB for 4.7 T. Using these optimized parameters, spectra of the peptide and protein mixture were acquired. Ions were directly infused into an electrospray source at 2 L/min flow rate, externally accumulated in the source and collision cell hexapole for 1 s each, and transferred to the ICR cell using ion transfer optics that had been previously optimized for signal intensity. Chirp excitation and image charge detection was performed. Three, 2-M-word datasets in the range of m/z 274 3000 were acquired and 4, 16, or 64 scans were averaged for each dataset. The FID was multiplied by a sine bell apodization function and was Fourier transformed. The narrowband experiment (Figure 3) involved a 32-k-word dataset covering 3.0 Thomson (Th) and was averaged 325 times. We tried to eliminate bias during data analysis in the following way. Only the p (...truncated)