In the Laboratory
Using an NMR Spectrometer To Do Magnetic Resonance Imaging An Undergraduate Physical Chemistry Laboratory Experiment
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Wayne E. Steinmetz* and M. Cyrus Maher Department of Chemistry, Pomona College, Claremont, CA 91711; *
[email protected] NMR spectroscopy, a routine tool in the chemical laboratory, has become an invaluable aid to medical diagnosis through the development of magnetic resonance imaging (MRI). Several factors motivated our development of an MRI experiment for Pomona’s course in experimental physical chemistry. Although NMR was first discovered by physicists, chemists such as Richard Ernst and Paul Lauterbur have played a prominent role in its development. NMR spectrometers equipped with many of the components required for imaging are frequently found in chemistry departments. Students learn best by doing and MRI illustrates important topics such as gradients, multidimensional NMR, and relaxation. It clearly demonstrates the relevance of chemical principles to students of biology and medicine who contribute heavily to enrollments in chemistry courses. Imaging is achieved in MRI through the fundamental dependence of the Larmor frequency on the magnetic field B: ω = γB where γ, the gyromagnetic ratio, is 26753 radian Gauss᎑1s᎑1 in the case of protons (1). In chemical spectroscopy, great care is taken to maintain a constant, homogeneous magnetic field. However, in the imaging of biological specimens where water makes the dominant contribution to the signal, the magnetic field is varied linearly across the sample. As a result, there is a one-to-one relationship between the NMR frequency and position. Gradients are now routinely used in chemical spectroscopy in the execution of two-dimensional experiments (2). Most NMR spectrometers are now shipped with hardware and probes designed for one-dimensional gradients so imaging in one dimension is possible without additional hardware. Imaging in three dimensions requires the purchase of a probe with gradient coils in three orthogonal directions and a threechannel gradient amplifier. One major limitation on the execution of MRI procedures needs to be addressed: the biological specimen must fit inside a 5-mm o.d. NMR tube. However, in comparison with imagers in hospitals, conventional spectrometers have magnets with much higher homogeneity and produce images with higher resolution. The experiment employs the multi-slice-multi-echo (MSME) procedure, which is a two-dimensional experiment with gradients. We use the terminology of Ernst in the following simplified outline of the method (3). Consult the classic monographs by Callaghan and Liang and Lauterbur for the details (4, 5). Smith has written an instructive review that is accessible to undergraduates (6). The preparation period consists of a long and therefore selective RF pulse and a gradient applied along the z direction with the result that coherence is generated within a narrow slice at a location determined by the frequency of the pulse. A 180⬚ pulse is inserted in the center of the preparation period so the method functions as a variation of the classic Carr–Purcell spin–echo pulse sequence, 1830
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a feature of many pulse sequences (1, 2). The length of the preparation period functions as the echo time. During the evolution period, a second gradient is applied along the y direction. The pulse sequence ends with the detection period while a third gradient is applied along the x direction. Experimental Following a detailed protocol (available in the Supplemental MaterialW), the student performs all the steps normally conducted by a technician or radiologist in a hospital setting. A one-on-one pedagogy involving a series of tutorials has the result that the students are able to operate sophisticated instruments on their own and obtain excellent results. The student selects the biological specimen. We have obtained optimal results from plant materials with axial symmetry that nearly fill the volume of the NMR tube. A piece of dental floss is attached as a leash to the specimen, which is gently inserted into the active region of the sample tube. After tuning and matching the probe, the student repetitively executes a gradient-free pulse sequence while adjusting the shims to minimize the line width of the water peak. This method of shimming on the spectrum is the best means of optimizing the magnet homogeneity. The final preparatory steps of loading the pulse sequence and setting the imaging parameters are performed using ParaVision, Bruker’s imaging software. A long duty cycle, achieved with a repetition time of 2 s, is required since the instrument’s gradient coils are air cooled rather than water cooled. A field of view of 0.5 cm and a matrix size of 128 in both dimensions yield a resolution of 0.039 mm pixel᎑1 and define gradient currents that do not exceed the power limitations on the gradient coils. The full pulse sequence is designed to generate a train of echoes. The signal decays exponentially over the echo train with a decay time given by the transverse relaxation time, T2. With plant materials we observe a T2 of ca. 25 ms and are able to obtain acceptable images from seven echoes. A typical run with acceptable signal-to-noise requires one hour. In a transparent manner, ParaVision handles the processing of the 2D data set, the examination of the images, and the quantitative measurement of the signal in selected regions of interest (ROI). The students perform an anatomical analysis of their specimen from the images and extract values of T2 from the decay of the signal with the echo time. For example, those who choose plant specimens are able to identify anatomical elements such as the phloem and xylem in their pictures. They find that the experimental values of T2 are orders of magnitude shorter than the value expected for the conventional dipole–dipole model involving rotational diffusion (7). An alternate model developed by Brownstein and Tarr that is based on translational diffusion in a confined space—the cell—is proposed as an acceptable model (8).
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www.JCE.DivCHED.org
In the Laboratory
Bruker’s medical imaging software. XWINNMR controls the spectrometer and ParaVision is used to set up the experiment and process the data. ParaVision is designed for medical MRI units and installation of this software on an NMR spectrometer presents special challenges. The crucial configuration files are provided in the Supplemental Material.W Hazards No one with a pacemaker can enter the room with the spectrometer. The instrument employs high magnetic fields so ferromagnetic materials—for example, tools and gas cylinders—are also not allowed in the room. Students are advised to keep magnetic media such as credit cards outside the 5 Gauss line surrounding the magnet. Acknowledgments Figure 1. Two-dimensional image of the stem from a leaf of a tulip tree acquired by the MSME method. The 1-mm slice displayed was selected by the application of a selective, shaped pulse. The black regions contain no protons.
Results
A generous grant from the Howard Hughes Medical Institute covered the purchase of the triple-axis gradient probe, the three-channel Acustar amplifier, and two licenses for ParaVision. Walter Knoeller of Bruker vastly improved the quality of our images by refining the preemphasis file. W
Our students have enthusiastically embraced the new experiment and their lab reports have shown that the experiment is an excellent vehicle for learning challenging topics, such as relaxation. They have consistently obtained excellent images that compare favorably with those generated at laboratories devoted to MRI. A representative example is given in Figure 1. Implementation of Magnetic Resonance Imagining If a department already has a Fourier-transform spectrometer with a single-axis gradient probe, MRI is possible with the purchase of a triple-axis gradient probe and replacement of the single-axis gradient amplifier with a triple-axis amplifier. Careful adjustment of the probe’s preemphasis parameters is essential for obtaining good images. These accessories cost ca. $40,000. Pulse shaping and variable-temperature control are desirable yet not necessary options. A license for MRI software is also required unless the user wishes to write code for all the procedures. The instrument used for this experiment is a Bruker 400 MHz spectrometer that runs under Version 2.6 of XWINNMR and Version 2.1.1 of ParaVision,
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Supplemental Material
The experimental protocol, the ParaVision configuration file, and the gradient preemphasis file are available in this issue of JCE Online. Literature Cited 1. Friebolin, H. Basic One- and Two-Dimensional NMR Spectroscopy, 4th ed.; Wiley-VCH: Weinheim, Germany, 2005. 2. Berger, S.; Braun, S. 200 and More NMR Experiments; WileyVCH: Weinheim, Germany, 2004. 3. Ernst, R. R.; Bodenhausen, G.; Wokaun, A. Principles of Nuclear Magnetic Resonance in One and Two Dimensions; Oxford University Press: Oxford, UK, 1987. 4. Callaghan, P. T. Principles of Nuclear Magnetic Resonance Microscopy; Oxford University Press: Oxford, UK, 2001. 5. Liang, Z.-P.; Lauterbur, P. C. Principles of Magnetic Resonance Imaging; IEEE Press: New York, 2000. 6. Smith, S. L. Anal. Chem. 1985, 57, 595A–608A. 7. Carrington, A. L.; McLachlan, A. D. Introduction to Magnetic Resonance; Harper and Row: New York, 1967; Chapter 11. 8. Brownstein, K. R.; Tarr, C. E. Phys. Rev. A 1979, 19, 2446– 2453.
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