NMR Spectroscopy in the Undergraduate Curriculum - ACS Publications

Chapter 11

Downloaded by PURDUE UNIV on November 29, 2016 | http://pubs.acs.org Publication Date (Web): September 15, 2016 | doi: 10.1021/bk-2016-1225.ch011

Creating Scholarship Opportunities for Undergraduate Students through Use of High Field NMR Susan M. Schelble,*,1 Kelly M. Elkins,2 Ethan Tsai,1 Milton Wieder,1 and Rosemarie DePoy Walker1 1Department

of Chemistry, Metropolitan State University of Denver, P. O. Box 173362, CB 52, Denver, Colorado 80217, United States 2Department of Chemistry, Towson University, 8000 York Road, Towson, Maryland 21252, United States *E-mail: [email protected]

In August 2013, the Chemistry Department at Metropolitan State University of Denver was awarded a TUES grant. These funds made an immediate difference in the educational experiences of undergraduates enrolled in organic chemistry laboratory courses and involved in undergraduate research. The title of the grant, NMR Spectroscopy: Introducing the Modern Chemist’s Toolkit to Undergraduates aptly describes the success that the installation and broad use of a high-field NMR spectrometer has had on the pedagogy at this PUI. Curricular changes include a transition to inquiry-based labs and project assignments that introduce research. This publication describes the uses of the instrument in organic and forensic chemistry laboratory courses. The intellectual merits for enhanced learning, broader impacts, and assessments thereof are summarized.

© 2016 American Chemical Society Soulsby et al.; NMR Spectroscopy in the Undergraduate Curriculum: Upper-Level Courses and Across the Curriculum Volume 3 ... ACS Symposium Series; American Chemical Society: Washington, DC, 2016.


Introduction The need to develop a modern, inquiry-based organic curriculum has been critical in propelling a paradigm shift in the overall chemistry program at Metropolitan State University of Denver (Metro), a 4-year undergraduate urban institution serving 425 undergraduate chemistry majors. The majority of these students are first-generation post-secondary students, who often have no built-in mentorship for navigating the paths to modern careers in science. The TUES (1) grant and the Denver Metro Chem Scholars grant (2) have combined support for this student population, and have been successful in preparing our chemistry majors to compete with their peers across the state and the country. Inclusion of modern spectroscopy into the undergraduate chemistry curriculum immediately provided students with skills that will be used in their careers. This chapter will describe the learning opportunities experienced by students at four institutions and the evaluation of goals that have been emerging since the inception of these two grants from NSF (Fall 2013). It will also compare and contrast using fixed magnet spectrometers (under 90 MHz) and high-field multinuclear probe spectrometers.

Obtaining Funding for Instrumentation The TUES proposal emphasized that: modern spectroscopic technique is a fundamental component of daily tasks for any organic chemist. It is imperative that basic spectroscopic technique is not only taught in organic chemistry laboratories, but also employed throughout laboratory practice—particularly with regard to one of the most powerful tools of the synthetic chemist: NMR spectroscopy. The award funded the purchase of a 300 MHz Bruker Avance instrument, which was installed in the fall of 2013. This award was key for developing a modern undergraduate research program and supporting the enhancement of instruction in organic and upper division chemistry laboratory courses. Key instrumental enhancement purchases were MNovaTM processing software (MestReNova v9.0.0-12821), as well as a 16-channel auto-sampler. The software was installed on multiple computers where students can retrieve file data from samples they submit for NMR analysis. Following the NSF award, the high field NMR spectrometer was quickly brought on-line to serve the approximately 420 organic laboratory course students and 20-35 research students at Metro. By the second year of the grant, users from other institutions also were sharing time on the instrument. These users include students at the Community College of Denver, CCD, the University of ColoradoDenver, UCD (both co-located on the same campus) and Towson University.

How the Instrument Was Used The 300 MHz NMR spectrometer was installed in the basement of the 145,000 ft2 science building (2011 construction) shared by three institutions: Metro, UCD, and CCD. Access to the co-located research centers in the basement is limited to trained users. Students generally cannot use the instrument directly, but can 184 Soulsby et al.; NMR Spectroscopy in the Undergraduate Curriculum: Upper-Level Courses and Across the Curriculum Volume 3 ... ACS Symposium Series; American Chemical Society: Washington, DC, 2016.

prepare, submit, and work-up data from samples they send to be analyzed by highfield NMR. All chemistry majors have the option of learning to work directly with the instrument. Students in the organic chemistry laboratory courses are provided hands-on spectrometer use of the 60 MHz Anasazi NMR spectrometer located in their laboratory space. Along with the new equipment, the Anasazi was also upgraded to include MNovaTM processing software.


Pre-Instrument Purchase Before installing the 300 MHz NMR spectrometer, organic lab courses were only able to provide 10-15% of ~650 organic lab students with access to collecting NMR spectra each year. Challenges came from out-dated processing software, limitations of the lower field magnet, as well as user time. Other users (students in UCD courses) were limited by older spectrometers (ca. 1995) and increasing instrument failures and slow shimming/processing software. We investigated numerous options for providing NMR access to more users, including benchtop NMR spectrometers. Because the current Anasazi instrument was already functioning at the low field magnetic strength, this option was not taken. Instead joint efforts were made to acquire and support the acquisition of a high-field 300 MHz NMR spectrometer.

Changes after 2013 Once the instrument came on line, students were able to submit NMR samples. Funding also supported the training of Teaching Assistants (TAs). These advanced students were trained to use the 300 MHz NMR spectrometer, worked with faculty on team-teaching in the laboratory courses, and were educated through the Denver Metro Scholars Grant (2) with additional support coming from the Dean of College of Letters, Arts and Sciences at Metro. The team of faculty and TAs prepared interactive on-line laboratory manuals (3, 4) and incorporated High Order Cognitive Skill Development (5–7) into the labs. The undergraduate TAs had opportunities to help their peers construct (6) their knowledge. On a typical day, TAs would take sixteen samples from a given section; run each sample with appropriate protocols (solvent, experiment, etc.) and email the FID data to each student user. This provides a turn-around of 24 hours, where student users can then process data on computers equipped with MNovaTM software.

Collaborations with Metro One of the NSF grant (1) Co-PI’s, Kelly Elkins moved to Towson University before the grant was awarded. This transition resulted in a very positive impact on the project. Because Dr. Elkins is located at an institute with graduate studies in forensic science, we were able to establish collaborations between the two institutions. Dr. Elkins collected FID spectral data from forensic science course laboratory work at Towson University which was sent to Metro, where three 185 Soulsby et al.; NMR Spectroscopy in the Undergraduate Curriculum: Upper-Level Courses and Across the Curriculum Volume 3 ... ACS Symposium Series; American Chemical Society: Washington, DC, 2016.

of the PI’s (Elkins, Tsai, and Schelble) created figures through MNovaTM for a publication (8). Details about this and other instrument uses follow.

Results Several interesting examples of learning opportunities have been provided to students using the grant funds. Examples illustrate skill development in laboratory courses and advanced learning acquired by students doing research.


Identification of an Unknown Organic Molecule One of the skills we strive to develop in organic students is the ability to use data to identify the structure of an unknown compound (Table 1).

Table 1. Possible Alcohol Identity

Students collected physical and FTIR data on the clear, colorless liquid and predicted the likely identity to be one of the alcohols in Table 1. The physical data is too close for students to be 100% confident in the identity of the compound. When students collected a 1H NMR spectrum on the 60 MHz instrument, the data in Figure 1 was obtained.

Figure 1. 1H NMR spectrum of an unknown alcohol at 60 MHz. 186 Soulsby et al.; NMR Spectroscopy in the Undergraduate Curriculum: Upper-Level Courses and Across the Curriculum Volume 3 ... ACS Symposium Series; American Chemical Society: Washington, DC, 2016.


Whereas an experienced organic chemist might find it easy to determine which compound produced this spectrum, the novice student has difficulty. This is partly because this spectrum does not look like examples from current textbooks. When the same sample was run at 300 MHz (Figure 2), enough information was evident to identify the compound.

Figure 2. 1H NMR spectrum of unknown alcohol at 300 MHz. There is a pentet overlapped by the alcohol proton at δ=1.5 ppm. Six of the most upfield methylene groups are a broad multiplet at δ=1.25 ppm. The high-field instrument was able to display the methylene triplet at δ=3.68 ppm and methyl triplet at δ=0.9 ppm.

The experienced organic chemist using the 60 MHz instrument will recognize that the triplet at 3.6 ppm indicates 1-hexanol. Students are not as adept at seeing this likelihood. The eight aliphatic proton signals overlap too much to make a definitive structural assignment. The student who processed the data in Figure 2 was new to using MNovaTM. The integration defaulted to assigning 1.00 for the proton environment at 3.68 ppm. However, when led to think about proton proportions, the typical student was able to see that all the integrals needed to be doubled, which made the data consistent with 1-hexanol. This exercise, where students looked at spectra run at two different magnetic strengths also provided them with insights about the varying location of the alcohol proton. It is at 1.7 ppm and 1.5 ppm in Figures 1 and 2, respectively. Identification of a Mono-Substituted Benzene The 1H NMR spectra of benzylacetone at 60 MHz and 300 MHz are shown in Figures 3 and 4. 187

Soulsby et al.; NMR Spectroscopy in the Undergraduate Curriculum: Upper-Level Courses and Across the Curriculum Volume 3 ... ACS Symposium Series; American Chemical Society: Washington, DC, 2016.


Figure 3. 1H NMR spectrum of benzylacetone at 60 MHz.

Figure 3 shows a pseudo-singlet signal for the five benzene protons and a nondescript multiplet for the four methylene protons. At 300 MHz (Figure 4) the spectrum is more resolved. The methylene protons alpha to the carbonyl group and the benzene ring, have an indeterminate “signal” at 2.7 ppm at 60 MHz. At 300 MHz, the “signal” resolves into two clear triplet signals (inset in Figure 4).

Figure 4. 1H NMR spectrum of benzylacetone at 300 MHz. Also defined at 300 MHz are the aromatic protons into two multiplets of 2H and 3H.

Analysis of a para-Disubstituted Benzene When the two substituents on the benzene have significantly different deshielding properties, such as in p-hydroxybenzylacetone, students expect to see two doublets in the aromatic region, integrating to 4 protons (Figure 5). At 60 MHz, the four protons appear as a pseudo quartet. At 300 MHz, these are clearly resolved into two doublets, as illustrated in Figure 5. 188 Soulsby et al.; NMR Spectroscopy in the Undergraduate Curriculum: Upper-Level Courses and Across the Curriculum Volume 3 ... ACS Symposium Series; American Chemical Society: Washington, DC, 2016.


Figure 5. The aromatic regions on the 60 MHz (left) and 300 MHz (right) for 4-hydroxybenzylacetone. The Ha protons appear as a doublet at 7.05 ppm; the Hb protons as a doublet at 6.75 ppm. The integration on the latter indicates that the phenolic hydrogen is overlapping with the Hb doublet.

Analysis of 3-Heptanone The 1H NMR spectrum of 3-heptanone acquired at 60 MHz is difficult to interpret to multiple overlapping signals (Figure 6), but the spectrum is nicely resolved at 300 MHz (Figure 7).

Figure 6. 1H NMR spectrum of 3-heptanone at 60MHz. The quartet from a overlaps with the triplet from b at 2.4 ppm. Multiplets from c and d overlap at 1.4 ppm. The triplets at 1.0 ppm are from overlapping signals from e and f.

At 300 MHz (Figure 7), the four alpha methylene hydrogens exhibit a “signal” that consists of a quartet overlapping a triplet. All remaining proton signals are clearly resolved and are consistent with the structure of 3-heptanone. 189 Soulsby et al.; NMR Spectroscopy in the Undergraduate Curriculum: Upper-Level Courses and Across the Curriculum Volume 3 ... ACS Symposium Series; American Chemical Society: Washington, DC, 2016.


Figure 7. 1H NMR spectrum of 3-heptanone at 300 MHz. The spectrum exhibits a quartet (methylene a) qQQq overlapping with a triplet (methylene b) tTt. Going from most downfield it appears as qQtQTqt. Multiplets from c and d are nicely resolved at this magnetic field (appearing at 1.55 and 1.35, respectively). The triplets at 1.1 and 0.9 ppm, respectively are from e and f. Analysis of an SN2 Reaction Once structural identification skills of pure compounds have been developed, students are ready to apply NMR analysis to a classic reaction for a first semester organic lab (Figure 8). This presents multiple opportunities for students to advance their skills in using NMR spectroscopy to characterize structures, evaluate success of techniques, and introduce 13C NMR and C−H 2-D NMR.

Figure 8. Planned conversion of 1-butanol to 1-bromobutane. This reaction, as typically performed in a first semester organic laboratory course, presents challenges to the student. The bimolecular nature of the mechanism is hindered by taking place in water. Hydrogen bromide is safer to handle when generated in-situ, however this means working with concentrated acid and excess NaBr salt. At the neutralization step, poor techniques and/or choices of base can lead to significant reversal of the reaction to starting materials. Prior to the reaction, students acquired a 1H NMR spectrum of the 1-butanol starting material at both 60 and 300 MHz (Figures 9 and 10). These spectra were collected at two magnetic strengths for the specific purposes of using them to 190 Soulsby et al.; NMR Spectroscopy in the Undergraduate Curriculum: Upper-Level Courses and Across the Curriculum Volume 3 ... ACS Symposium Series; American Chemical Society: Washington, DC, 2016.


monitor success of reactions by comparing effectiveness at various magnetic fields and to learn about aspects of J-coupling.

Figure 9. 1H NMR spectrum of 1-butanol at 60 MHz. At this point in the course, students calculate the J-coupling for the triplet at 3.625 ppm on the 60 MHz Anasazi instrument. For the data here, with two decimal places, the triplet has Δ ppm of 0.11 and 0.10 or 7 Hz and 6 Hz, respectively. The same calculation on the triplet (at 3.596 ppm) from the 300 MHz spectrum the triplet has Δ ppm of 0.020 and 0.022 or 6 Hz and 7 Hz, respectively. These subtle differences offer the perfect opportunity to explain how coupling constants are calculated and how magnetic field impacts Δ ppm but not Hz.

Figure 10. 1H NMR spectrum of 1-butanol at 300 MHz. The inset corresponds to the second and third methylene groups (integrated as 1:1). The chemical shifts of the triplet from the alcohol methylene are 3.616, 3.596, 3.574 ppm. A final insight can be gleaned from Figure 10. The methylene groups do not appear as the expected pentet and sextet that students are taught to predict. This is a chance to introduce impacts of constitutional and diastereotopic protons. The autosampler and programmable queue for samples allows users to request analysis of other nuclei besides protons and 2-dimensional spectral analysis. The total time for acquiring and processing the data for Figures 10-12 was about 30 minutes. Figures 11 and 12 illustrate the 13C NMR spectrum and 2D HMBC analysis of 1-butanol. 191 Soulsby et al.; NMR Spectroscopy in the Undergraduate Curriculum: Upper-Level Courses and Across the Curriculum Volume 3 ... ACS Symposium Series; American Chemical Society: Washington, DC, 2016.


Figure 11.

13C

NMR spectrum of 1-butanol at 300 MHz.

Figure 12. 2D NMR (1H 13C HMBC) of 1-butanol at 300 MHz. The challenges in synthesizing 1-bromobutane under less than perfect SN2 conditions have an upside for learning. A typical 1H NMR spectrum of a student product from this lab is shown in Figure 13. Students analyzing the spectrum in Figure 13 are challenged to explain the number of multiplets. This result offers numerous insights about experimental outcomes, improving yields, interpreting quantitative data from the respective integrations of the triplets (product versus starting material), and why those multiplets are all that are needed to draw conclusions about the success of the synthesis. This experiment prepares students for the second semester organic chemistry laboratory course and subsequent research opportunities involving spectral interpretation of results. It also serves as an introduction to the “scholarship” 192



we are trying to develop in our undergraduate population. Students in the introductory organic chemistry laboratory course are required to present work in oral, poster and written form (in the style of the Journal of Organic Chemistry). Several examples of laboratory course projects and introduction to research applications follow. These are used to extend the scholarship of undergraduate students. At every juncture we encourage communication of the interpretation of results. This could not be done at a modern level until we incorporated the 300 MHz instrument into the course work.

Figure 13. 1H NMR spectrum of product of 1-bromobutane and starting material. The starting material methylene (Hs) at ~3.6 ppm and product methylene (Hp) at ~3.4 ppm are clearly separated triplets. The inset of these two multiplets can be used to determine approximate amounts of each in the product mixture. Other multiplets in the 2-component mixture overlap.

Analysis of an Atypical Aldol Condensation Reaction The incorporation of high-field NMR spectroscopy has allowed the faculty to modify the curriculum to include investigations based on primary literature. An on-going project in this endeavor is the aldol condensation reaction. A classic paper (9) for a mixed aldol procedure is based on the reaction of aromatic aldehydes and symmetrical ketones under base catalysis in ethanol. The publication calls for performing the reaction in a 10% solution of ethanol with 2 equivalents of aldehyde (20-50 mmol), one equivalent of ketone (10-25 mmol) and 20-40 mol of base (as a 15% solution in ethanol). The addition takes place at room temperature, and the stirred mixture is heated for 30 minutes at 70 °C. Cooling yields crystals which can be isolated using by vacuum filtration. A newer paper (10) notes a novel variation on the outcome when using p-chlorobenzaldehyde and 3-pentanone. The reactions are summarized in Figure 14. 193



Figure 14. Reactions that take place under aldol condensation conditions. The top reaction was described in reference (9) and the bottom in reference (10). Before installing the high-field NMR spectrometer, we challenged students to perform these aldol condensations and investigate the conditions that led to the divergent outcomes. They carried out the reaction from (10) and recovered a white solid with physical properties matching the second reaction in Figure 14. At 60 MHz, the aromatic protons, Ha and Hb appear as a broad singlet (Figure 15). The methyl groups at 0.9 ppm are clearly a doublet from Hd, and Hc appear as a doublet from Hd at a typical chemical shift of 4.4 ppm.

Figure 15. 1H NMR spectrum of aldol product at 60 MHz where the aromatic protons, Ha and Hb appear as a broad singlet. The methyl groups at 0.9 ppm are clearly a doublet from Hd, and Hc appears as a doublet from Hd at a typical chemical shift of 4.4 ppm. The Hd protons are a doublet of quartets at 2.9 ppm, but at this field strength, it appears as a broad multiplet. The Hd protons are a doublet of quartets at 2.9 ppm, but at 60 MHz, it appears as a broad multiplet. The integrations match the compound shown, however, the expected pseudo quartet for the para-disubstituted aromatic regions is not evident. 194 Soulsby et al.; NMR Spectroscopy in the Undergraduate Curriculum: Upper-Level Courses and Across the Curriculum Volume 3 ... ACS Symposium Series; American Chemical Society: Washington, DC, 2016.


Once the high-field instrument was brought into service, this compound was analyzed at 300 MHz. Figure 16 illustrates the aromatic region of this spectrum. Students in undergraduate teaching laboratories are expanding this investigation beyond the published experiments. It is a perfect opportunity to see if other substituted aldehydes (e.g. p-bromobenzaldehyde) or other symmetrical ketones (e.g. 4-heptanone) will lead to atypical outcomes for the aldol condensation reactions.

Figure 16. 1H NMR spectrum of aldol product at 300 MHz where the aromatic protons, are a pseudo quartet (doublet of doublets from Ha and Hb). Some additional details can be seen for the multiplets derived from Hd protons at 3.04 ppm. These ring protons however, do not exhibit a typical quartet. Analysis of Cathinones One of our co-PIs moved to a new institution, Towson University, after the grant proposal was submitted to NSF for review. The goal and responsibilities of bringing forensic applications of NMR to undergraduate students and students in forensic science courses was carried out through collaboration with Towson University. Initially conceived as a research project with two undergraduates during spring 2014, the methods were later implemented in a regular forensic chemistry course during the fall 2014 semester. A JEOL 400SS MHz NMR, also acquired with NSF funding, was used in the project and course. Most forensic laboratories began to encounter designer cathinones in casework in 2009 with a significant increase by 2011-2012 according to the National Drug Intelligence Center. Synthetic cathinones are small organic molecules that have been presented in the literature for almost a hundred years. 195



(See Figure 17). Forensic scientists often rely upon GC-MS and FT-IR analysis, among other methods, to identify the compounds present in seized materials. However, the drawback of using these techniques for identification is that previously recorded spectra for each substance must be present in a library for pattern matching. Although most crime laboratories do not have NMR spectrometers, students are taught that NMR is one tool that can be used for total structure determination of new designer drugs (11). Additionally, drug dealers increase their product and profits by extending the quantity of “drug” by adding other cheap, white powders—called cutting agents, or adulterants—to the drug. The cutting agents are often widely available sugars or drug-like substances such as caffeine or lidocaine that impart little to no drug-like effects. Designer cathinones often contain wide-ranging concentrations (

NMR Spectroscopy in the Undergraduate Curriculum - ACS Publications

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