How MRR Works

Key MRR Features and Advantages

Extraordinary chemical specificity

  • Unequivocally discriminates between all types of isomers including regioisomers, diastereomers, enantiomers, and isotopic species.
  • Each compound has a unique and absolute frequency pattern – no matrix effects or frequency shifts.

Ultra-high resolution

  • Reliably analyzes components in complex mixtures directly, without analyte isolation.

Quantitative

  • MRR intensities are directly and precisely proportional to concentrations.
  • Built-in response factors for isomer and isotope applications – no pure standards needed.

Online capable

  • Can bring stereoselectivity and other capabilities of high-end instrumentation directly to the process line to bridge analytical gaps of conventional online techniques.

MRR Background and History

Molecular rotational spectroscopy has been used for decades by research groups around the world for characterizing the structures of small molecules. One area where rotational spectroscopy is especially useful is for studying molecules in space – including in the interstellar medium and the protoplanetary disks around young stars. Radio telescopes are used to measure the microwave and millimeter wave spectra of distant regions, which include sharp lines due to molecules. Of the nearly 200 detected astromolecules (an up-to-date list is maintained at http://astrochymist.org/), the vast majority have been identified through their pure rotational spectrum. Due to increasing laboratory data and more sensitive radiotelescopes such as the Atacama Large Millimeter/Submillimeter Array (ALMA), more is being understood about our universe each year due to these rotational lines.

Beyond astronomy, rotational spectroscopy, has been more recently emerging as a powerful new tool for diverse chemical laboratory and process monitoring applications. This emergence is a result of incredible progress made recently in the rotational spectroscopy instrumentation, enabling a range of new measurement capabilities. Perhaps the most important of these developments is the chirped-pulse Fourier transform microwave spectroscopy, which increased the speed with which broad-bandwidth rotational spectra could be measured by many orders of magnitude (approx. 1,000X) and reducing the amount of sample required for an analysis. Now, BrightSpec offers commercial instrumentation for the measurement of MRR spectra for both industrial and research applications.

Molecular Rotational Resonance Spectroscopy – Fundamental Principles and Advantages

In the low-pressure gas phase, a molecule’s rotational angular momentum is quantized, leading into discrete frequencies at which it can absorb or emit radiation and hence increase or decrease its rotational speed. These energy levels are incredibly sensitive to that molecule’s three-dimensional geometry: the moments of inertia. Even molecules with the same mass (isomers) will rotate differently because of their subtle differences in structure. The classical analog to this is a figure skater: a figure skater will rotate at a different speed with her arms tucked in than with arms extended, for the same energy input.

At the molecular level, we can use the information the rotational spectrum provides to learn about the 3- D geometry of the molecule (or molecules) present in the sample under study. No two compounds have the same MRR spectrum, with the exception of enantiomers (and these can be resolved by a special method as discussed later). Through the combination of this incredible specificity and the ultrahigh resolution of the technique (typically better than one part in 10 5 ), MRR can identify compounds without ambiguity or overlap in a mixture. The frequency measurement is also highly accurate, through precise frequency referencing to atomic standards, and so, as the renowned chemist E. Bright Wilson put it in a 1968 paper in Science, “Once a molecule has been caught and finger-printed by this method, it is forever recognizable.” Recent work has also demonstrated that quantitation by MRR is also possible as the dynamic range of the measurement is accurate over several orders of magnitude, and the line intensity is proportional to the concentration of that analyte in the gas-phase sample.

For the anayltical chemist, the advantages of MRR are:

  • Separation is not needed: samples can be injected directly into MRR spectrometers and analyzed. In the case of multi-component mixtures, the spectra almost never overlap and so each component in the mixture can be identified and quantified. If MRR is combined with a separation technique (such as with a gas chromatograph), co-eluting compounds can be easily distinguished through MRR.
  • No chemometrics: Components don’t overlap spectra in a mixture. This means that identification and quantification is simple, and unexpected impurities don’t complicate or invalidate the analysis.
  • Pure reference standards are not needed: Compounds can be identified directly in crude mixtures by comparing computed and measured moments of inertia. This can save synthetic chemists lots of time and cost required to synthesize pure standards.
  • Measurements can be fully automated and brought on-line, with cycle times of only about 10-15 minutes.

Another Breakthrough: Chiral Analysis

As noted above, the one set of compounds that couldn’t be measured by MRR until recently are enantiomers, which have the same moments of inertia. Just as for other analytical techniques, either a chirally sensitive detection method or a chiral resolving agent is needed. In recent years, both of these have been developed for MRR. A chiral resolution method called chiral tagging, invented at the University of Virginia, has been incorporated into BrightSpec instruments for measurement of enantiomeric excess and absolute configuration. As for all other MRR analyses, a standard is not required. A small chiral molecule called a tag is mixed into the analyte mixture, and the combination is injected through a pulsed supersonic nozzle into the vacuum chamber. This interaction allows weakly bound complexes to form. When the two enantiomers form complexes with the chiral tag, diastereomers result with distinct moments of inertia and the capability to be resolved just as for all other analytes in MRR. Recent studies have shown that chiral tagging works across a wide array of compounds, and produces enantiomeric excess values with comparable accuracy to chiral chromatography.

Comparison to Other Techniques

MRR vs Mass Spectrometry

Mass spectrometry (MS) identifies species based on their mass and fragmentation pattern, whereas MRR spectroscopy identifies molecules based on their three-dimensional mass distributions. As such, MRR chemical specificity exceeds that of mass spectrometry. For example, MRR can easily discriminate between all types of isomers (see Figure below), isotopomers of identical mass, and, with the chiral tagging technique, can also perform direct chiral analysis.

Compared to the direct quantitative mixture analysis capability of MRR, MS often needs to be combined with chromatography to improve the analysis accuracy. As such, the key MRR implementation benefit is the elimination of the chromatography step. This results in faster analyses, reduced consumable costs (no columns), and eliminating time-consuming chromatography method development and validation steps. Please see our Natural Products and Deuterated Drug Analysis pages for MRR analysis examples.

Alternatively, MRR can be hyphenated with gas chromatography (GC) to result in an exceptionally selective and powerful GC-MRR combination. As we demonstrate in the Angewandte Chemie Article, GC-MRR can successfully execute analyses that are impossible with any other today’s analyticaltechniques or combination of techniques including GC-MS. As a result, GC-MRR can likely not only serve as a powerful add-on or alternative to GC-MS but also open a new era in several fields of analytical chemistry including deuterated pharmaceuticals, natural products, metabolomics, compound-specific isotope analysis (CSIA), biodegradation and environmental fate studies, and many others.

Summary of MRR advantages over mass spectrometry:

  • Resolves all types of isomers including the regioisomers, diastereomers, and enantiomers.
  • Enables a superior position-specific and/or compound-specific isotope analysis: resolves not only isotopologues of different masses but also isotopomers of identical mass.
  • Easily discriminates between deliberately deuterated and natural abundance isotopologues.
  • Does not suffer from ion suppression.

MRR vs NMR

NMR technique is a current gold standard for de-novo structure analyses and impurity structure elucidation. Sharing the extraordinary sensitivity to molecular structure with NMR, MRR can serve as an orthogonal technique to confirm or complement small-molecule NMR structure elucidation studies. In addition, MRR does not require deuterated solvents and can simplify sample workflow in your lab.

MRR analyses can be of an especially high value when NMR produces ambiguous results. For example, resolving stereoisomers or isotopically labelled species can be easily performed by MRR on minute quantities of samples, and without reference standards.

In addition, due to its high resolution and lack of frequency drifts, MRR is better suited for determining the structure of components directly in a crude mixture, without the need for isolating individual components.

Furthermore, MRR is online-capable and can perform routine process monitoring without a loss in resolutions. As a result, MRR can achieve a laboratory-grade quality of chiral and achiral analyses at process setting.

MRR vs FTIR/Raman

MRR spectra typically contain many more sharply resolved features than IR or Raman spectra. On top of that, MRR has a very high resolution so the likelihood of overlaps in mixtures is very low. As such, MRR is typically better suited for analysis of complex multicomponent mixtures, as this technique can resolve not only main components but also trace level impurities. In addition, MRR is stereoselective and easily discriminates between all types of isomers including diastereomers and enantiomers that cannot be resolved by IR or Raman.

Furthermore, as MRR shares online capability with IR and Raman, this technique can bridge the analytical gaps of IR or Raman and bring the stereoselectivity, ultra-high resolution, and other capabilities of high-end instrumentation directly to the process line. In addition, MRR implementation can result in significant time and labor savings, as time-consuming multivariate chemometric model development and maintenance are not required for MRR (in contrast to IR and Raman). As a side benefit, quantitative process monitoring with MRR is likely to be exceptionally reliable, as chemometrics-free MRR methods are not expected to suffer from unavoidable short-term and long-term process variations.

Figure below directly compares MRR and Raman capabilities for online monitoring of Ru/C catalyzed stereoselective hydrogenation of artemisinic acid (AA) to dihydroartemisinic acid (DHAA). Specifically, Raman can monitor the reaction progress but cannot resolve the side products, such as DHAA’s epimer and the tetrahydroartemisinic acid (THHA – the overreduction byproduct). In contrast to Raman, MRR enables direct interference-free quantitation of all four crude reaction mixture constituents including starting material, product, and both side-products.

Summary of MRR advantages over IR and Raman spectroscopy:

  • Superior specificity due to several fold greater number of and much sharper spectral features.
  • Stereo and enantioselectivity: diastereomers or enantiomers are not a problem.
  • No matrix interferences, absolute frequency accuracy, background-free.
  • No multivariate chemometric models are required: no time spent on model refining and / or maintenance.
  • No pure standards are required for a wide range of applications.
  • Easy method transfer between instruments and/or lab-scale, pilot-scale, and full-scale processes.
  • For process monitoring, MRR results do not suffer from unavoidable process variations.