NIR vs. Raman: Spectroscopy Showdown

Raman and near-infrared (NIR) spectroscopy are complementary methods, both probing vibrational transitions in molecules. In general the strong bands in the (N)IR spectrum of a compound corresponds to weak bands in the Raman and vice versa. This blog will look at some of the differences between Raman (light-scattering) and NIR (light absorption) methods.

Here just for the meat? Skip down to the “Cutting to the Chase” for some practical similarities and differences further below.

Quick Refresh: What is Vibrational Spectroscopy

Figure shows some typical (normalized) Raman spectra of two pharmaceutical powders, HPMC-AS (green) and PVP-VA (blue), as well as crystalline drug Indomethacin (red).
A time series of normalized NIR spectra of pharmaceutical drug Indomethacin undergoing a phase transition.

Atoms aren’t wallflowers, they dance to the good vibrations!

When energy of the frequency range between 10,000 cm-1 and 100 cm-1 is absorbed by a molecule, it is converted into energy of molecular vibration.  A stretching vibration occurs when the interatomic distance between two atoms increases or decreases along the bond axis, while bending vibrations are characterized by a change in the angle between two bonds with a common atom or the movement of a group of atoms with respect to the remainder of the molecule.  The frequency or wavelength of absorption at which a vibration occurs depends on the relative mass of the atoms, the force constants of the bonds (i.e., bond strength), and the geometry of the atoms.  Therefore, certain groups of atoms, or functional groups, give rise to characteristic peaks at or near the same frequency regardless of molecular structure; this is the premise for compound identification using vibrational spectroscopy.  You can read more about this in a previous blog.

Raman: Digging into the Principles

Raman spectra are acquired by irradiating a sample with a monochromatic source (think: lasers) at a frequency (or energy) which is higher than the resonant vibrational frequencies of a molecule.   Raman activity requires that the incident photons cause a transient distortion of the electron distribution of molecular bonds—a change in polarizability—which results in photon absorption.  The absorption process promotes electrons to a virtual, non-quantized energy level, from which photons are reemitted (i.e., scattered) and detected. In general, atoms in which electrons are far from the sphere of influence of the positively charged nucleus are more polarizable (i.e. homonuclear bonds which experience symmetric vibrations). 

Raman radiation may be elastically or inelastically scattered.  When excitation occurs from the ground vibration energy level (v = 0), the inelastically scattered radiation is of a lower frequency than the excitation frequency, h(vex  – vv), where vv is the vibrational frequency of the bond.  This is called Stokes scattering.  When molecules in the first excited vibrational state (v = 1) scatter radiation inelastically, the Raman signal produced is of energy h(vex  + vv);  the scattering occurs at a higher frequency than the radiation source and is called anti-Stokes scattering.    Elastic scattering occurs when a photon is emitted with the same energy as the excitation source (hvex), called Rayleigh scattering.  Only inelastically scattered photons provide information regarding molecular vibrations, although Stokes emission is generally favored for data analysis due to the higher signal intensities that are achieved relative to anti-Stokes scattering, a consequence of a greater population of electrons in the ground state at any given time relative to the excited state.

Raman spectra are recorded according to the wavenumber shift from the excitation frequency which is equivalent to the vibrational frequency.  Therefore, the change in energy produced by the vibration would be equivalent to that observed in absorption if the bond was also IR-active, making the Raman frequency shift and the IR absorption frequency identical.  Raman shift is independent of the excitation wavelength.  The intensity of a Raman band depends on the polarizability of the molecule, the source intensity, and the concentration of the active group.  Raman intensity is directly proportional to the concentration of the analyte. 

Cutting to the Chase:

What are notable practical similarities between NIR and Raman?

  • Both are used in chemistry to provide a structural fingerprint by which molecules can be identified
  • Multivariate spectroscopic data analysis of Raman or NIR signal can be used to accomplish quantitative (compositional) analysis of organic samples
  • They are both non-destructive, enabling direct analysis of intact samples (i.e. no dissolution, trituration or dilution of samples)
  • Fast measurements (on the order of seconds to a few minutes) make these methods practical for higher frequency sampling or process monitoring
  • Both spectrometer types may be interfaced with probes to improve sample accessibility, which is useful for process analytical technology applications

What are the practical differences between NIR and Raman

  • If a molecule has a center of symmetry, Raman active vibrations would not be visible in the infrared.  Asymmetric stretches, which induce a dipole, are infrared active.  This difference is why Raman and Infrared Spectra are considered complementary
  • Vibrations involving polar bonds (e.g. C-O, N-O, O-H) are weak Raman scatterers but strong NIR absorbers, while bonds such as C-C, C-H and C=C are observed well using Raman and not using NIR
  • NIR spectra is more broad and overlapping than Raman, which is characterized by sharper peaks (signal). While multivariate analysis (e.g. PLS, PCA) often overcomes this challenge, Raman may offer better selectivity in some cases and is more appropriate for analysis of unknowns
  • Whereas water is a strong absorber in NIR, it is inactive in Raman.  Therefore, Raman analysis may provide an advantage when water is a major interference and not an analyte of interest (e.g. monitoring crystallization in aqueous solutions)
  • When water is an analyte of interest (e.g. when monitoring a drying process), NIR is appropriate
  • NIR is preferred for non-crystalline, hydrophilic molecules (like pharmaceutical excipients) which are often poor Raman-scatterers relative to the generally small aromatic, heterocyclic drug molecules
  • Some Raman lasers may cause sample heating which may result in chemical degradation. Laser excitation may also cause sample fluorescence or photobleaching, which imparts baseline effects. This may be observed most in noncrystalline (amorphous) samples

You can learn more about BUCHI NIRSolutions, including: the flexible R&D laboratory NIRFlex N-500, the rugged at-line NIRMaster and ProxiMate, and the in-line or on-line NIR-Online using the links.

Want to dig in deeper into the NIR vs Raman comparison? Check out an article published in Spectroscopy Magazine on the subject.

NIR: a Spring-y subject

Winter felt brutal and eternal, as it always does for someone who doesn’t ski or care for hot chocolate, I suppose. What a relief it is to see signs of Spring emerging from my brownish-colored yard and hear birds chirping outside once again.

Did you know NIR is quite Spring-y as well. This blog will explore some spring-themed theory.

beautiful bird bloom blossom
Photo by Pixabay on Pexels.com

At temperatures above absolute zero (i.e. even in the dead of a Northeast USA snowmageddon), all of the atoms in a molecule are in continuous vibration with respect to each other.

The behavior of molecular vibration is analogous to a mechanical model in which two masses connected to the ends of a spring! A disturbance of one of these masses along the axis of the spring results in a vibration called the simple harmonic oscillation.

close up of metal
Photo by Pixabay on Pexels.com

Vibration, or the displacement of an atom relative to its equilibrium position, produces potential energy proportional to the work required to displace the mass.  This energy is at its maximum when the spring is stretched or compressed to its maximum amplitude. It is at its minimum (i.e., zero) at the equilibrium position.

The fundamental requirement for infrared (Far-IR, mid-IR, near-IR) activity, leading to the absorption of infrared radiation, is that the energy of incident radiation matches the vibrational energy levels exactly, and that the vibration itself causes a change in dipole moment.  The frequency of radiation that will bring about this change can be calculated by Hooke’s Law:

Hookes Law

where c is the speed of light (3×103 cm/s), f is the force constant of the bond (dyne/cm), and Mx and My are the masses of  the atom x and atom y involved in the bond, respectively.  The force constant is positively correlated to properties such as bond order or bond strength (i.e. the “springiness” of the bond).  In accordance with the Boltzmann distribution, frequencies which correspond to fundamental transitions between the ground state and first vibrational level (n = 1) dominate the vibrational absorption spectrum.  Because the majority of absorption bands of chemical compounds correspond to fundamental vibrations at infrared frequencies, it is a common tool for structural elucidation.

How does thee vibrate? Let me count the ways.

The number of possible or theoretical fundamental vibrations is determined by the total degrees of freedom of the molecule.  Each atom requires three degrees of freedom in order to describe its position relative to other atoms in the molecule.  Therefore, a molecule of N atoms has 3N degrees of freedom.  For nonlinear molecules, six degrees of freedom are used to describe translation and rotation; the remaining 3N – 6 degrees of freedom are vibrational degrees of freedom (i.e., fundamental vibrational modes).  For linear molecules, only two degrees of freedom are required to describe rotation, resulting in 3N-5 normal modes.

Disclaimer: Sometimes 1+1 doesn’t equal 2

The number of theoretical bands will not necessarily equate to the number observed experimentally.  The number of theoretical bands observed may be reduced by: lack of a change in the molecule’s dipole as it vibrates or rotates, fundamental frequencies that fall outside of the infrared region or are too weak to be observed, vibrations that coalesce, or the occurrence of a degenerate band from several absorptions of the same frequency in highly symmetrical molecules.  On the other hand, vibrations at integer-multiples of a given frequency and combination tones will increase the actual number of bands observed.  It is from the combination and overtones transitions that NIR spectra arise.

The NIR region of the electromagnetic spectrum covers the range of approximately 14,000 to 4,000 cm-1, or about 700 to 2,500 nm.   The most prominent absorption bands occurring in the NIR region include overtones and combinations of fundamental vibrations of the IR-active –CH, –OH, -CO, –NH and –SH functional groups present in most pharmaceutical drug molecules.  Due to the relatively weak molar absorptivities of the transitions responsible for the peaks observed, sample dilution is not required.  This characteristic also provides for relatively deep sample penetration up to several millimeters thick, especially at shorter wavelengths (e.g., 700-1500 nm).  Even though NIR spectroscopy is characterized by spectra which are typically broad, overlapping and of low intensity relative to the fundamental mid-IR absorption bands, it has some practical advantages.  The richness and utility of NIR spectra is a consequence of anharmonic oscillation.

Anharmonicity: where the simple rules start to break down and things start to get interesting

Bonds which share a common atom seldom behave as independent oscillators.  As the interatomic distance separating two atoms decreases, coulombic repulsion between the nuclei results in an additional force which acts in the same direction as the force restoring the system toward equilibrium.  Thus, the potential energy of the system increases more rapidly than predicted by the harmonic oscillator.  On the other hand, as the interatomic distance approaches that at which dissociation of the atoms takes place, a decrease in the restoring force and potential energy of the system occurs.  The intramolecular interactions produce non-symmetric vibrations about the equilibrium position.  The anharmonicity results in non-equivalent energy changes between vibrational states, where ΔE becomes smaller at higher quantum numbers.   Moreover, the selection rule is not rigorously followed (because rules are made to be broken), thus allowing the overtones responsible for much of the NIR spectra, where Δn = ±2, ±3 and ±4 represent the first, second and third overtones, respectively.

The degree of anharmonicity determines the extent of the displacement from an integer multiple of the fundamental frequency, as well as the intensity of the overtones.   Vibrations stemming from intramolecular hydrogen-bonding vibrations have the highest anharmonicity constants, leading to their prevalence and high intensity in the NIR region. That’s why NIR is so great at measuring low levels of water in samples!

More interesting stuff in the spectra:

NIR spectra are further enriched when vibrational modes interact to give absorptions at frequencies that are the approximate sums or differences of their fundamental frequencies.  These combination bands, which generally occur between 1900 and 2400 nm, are a consequence of energy absorption by two bonds rather than one, allowing the photon to excite two vibrational modes simultaneously.  As with overtones, the intensities of combination bands are weaker than their fundamental frequencies.

A special type of interaction called Fermi resonance occurs as a consequence of accidental degeneracy of different vibrational modes have the same symmetry and approximately the same frequency as a fundamental vibration.  This results in two relatively strong absorbance bands which are displaced at slightly higher and lower frequencies than expected, respectively.

Darling and Dennison resonance affects vibrations which have identical symmetry species and similar energies, leading to several pairs of absorption bands.

Coupling between oscillators results in slight to moderate shifts in the absorption frequency of the molecules involved.  In general, coupling requires that vibrations be of the same symmetry species and a common atom or bond between the two vibrations or vibrating groups, respectively.  The interaction is greatest when the coupled groups have nearly equivalent energies; little to no interaction is observed by groups separated by two or more bonds.  Despite the fact that coupling leads to uncertainties in functional group identification, it is this phenomenon that provides the unique features of a spectrum enabling compound identification.

 

For more reading on this spring-y subject:

Skoog D.A. and Leary J.J. Principles of Instrumental Analysis. 4th Edition. 1992.

Silverstein R.M, Spectrometric Identification of Organic Compounds, 5th Edition. 1991.

D. Burns, and E. Ciurczak,Handbook of Near-Infrared Analysis 2nd Edition, Marcel-Dekker, Inc. New York, 2001.

E. Ciurczak and J. Drennen, Near-Infrared Spectroscopy in Pharmaceutical and Medical Applications, Marcel-Dekker, Inc. New York, 2002.

L. Weyer and S.-C. Lo, “Spectra-Structure Correlations in the Near-infrared,” In Handbook of Vibrational Spectroscopy, Volume 3, Wiley, U.K., 2002.

 

 

What is NIR?

More alphabet soup

Near-infrared spectroscopy. “N-I-R.”  Let’s illuminate the subject a bit, shall we?

Spectroscopy is a branch of science interested in the interaction of light with matter. Near-infrared (NIR) spectroscopy happens when the light used to do the measurements falls within a certain energy or frequency range; typically, 12000 – 4000 cm-1 (or about 700 – 2500 nm in terms of wavelength).

This idea isn’t new. The first observations of NIR light were made by Herschel in 1800, and Coblentz was considered its pioneer in the early 1900’s. However, this small but mighty portion of the electromagnetic spectrum didn’t debut commercially until the 1970’s, coinciding with advancements in PC computing power that radically simplified it’s application.

Why do people use NIR?

Everything you’ve come to love in your life: people, places, baked goods… they are all made up of molecules. Those molecules are made of atoms, and those atoms are moving and grooving (i.e. the bond lengths and bond angles aren’t static, but rather wagging and scissoring and bending and stretching). We can use NIR to measure that molecular dance party, or more technically, molecular vibrations. Those vibrational modes can tell us stuff about the sample that most QC departments like, think: sample identity or composition. 

When the molecules of a sample are hit with NIR light, the light is either absorbed or scattered. When the light is absorbed, we see a peak in our NIR spectrum. When light scatters due to the physical properties of the sample (e.g. particle size, particle morphology, bulk density), the overall slope of the spectrum is impacted. Chemical bonds that absorb NIR well are: oxygen-hydrogen (O-H), carbon-hydrogen (C-H), carbon-oxygen (C-O), nitrogen-hydrogen (N-H), and sulfur-hydrogen (S-H). While NIR isn’t the magic bullet for every analysis, we see samples that are dominated by these types of bonds in many industries, from pharmaceuticals to pet food.

The series of peaks and valleys that appear in an NIR spectrum is the summation of molecular vibrations of the sample. Consider the spectra of 5 different solutions in the figure below, where Absorbance is shown as a function of wavelength (nm). The red spectrum is pure methanol, the green spectrum is pure water. Peaks resulting from the -OH vibrations of both the alcohol and water, as well as the -CH vibration of the alcohol, are labeled.

water-alcohol

Since the energy of the -OH (and -CH) bonds of water and methanol (or donuts) differ, they produce unique spectra, as illustrated in the figure above. Combine that with the fact that spectra (of water, methanol or donuts) is repeatable, and we’re going somewhere. That means, if I scan water with my NIR analyzer 10x, I will get (more or less) the same spectrum, and that spectrum is unique from the other stuff I want to analyze (here, methanol). For those reasons, NIR would be a good tool for identification purposes.

Take another look at the figure above. The peak intensities corresponding to each molecular vibration reflect the relative composition of the molecules (water and alcohol) contributing those bonds to the solution. More specifically, you can see the peak corresponding to the water -OH vibration around 1450 nm increase as the relative proportion of water increases in solution. In the same way, the peak corresponding to the -CH combination vibration around 2250 nm increase as the proportion of methanol increases in the solution. That’s Beer’s Law working for us, where Absorbance at a given wavelength is proportional to the pathlength * molar absorptivity * concentration of the absorbing analyte. Note to the academics: if you dig into the theoretical research, you will see that Beer’s Law applies strictly to ideal solutions. Oh, and if you hadn’t heard, almost nothing is ideal in real life. However, we have mathematical ways to address non-linearity, and in the end, most methods work well. And by work well, we mean produce satisfactory standard errors of prediction. 

But before you get too excited about the infinite possibilities of NIR, let me give you some fine print. Our mortal bodies typically can’t run marathons without training first… and an NIR can’t run a qualitative or quantification application without being trained, either.

Additional resources:

For a good historical, theoretical and applications overview, see the Handbook of Near-Infrared Analysis, edited by Donald A. Burns and Emil W. Ciurczak