Ahura Scientific, Inc.

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Technology Comparison:
FTIR, NIR and Raman Spectroscopy for
Pharmaceutical Raw Materials Inspection
The collection of analytical techniques known as “vibrational spectroscopy” consists of three
main tools, namely Mid-infrared or more commonly Fourier Transform Infrared (FTIR); Near-
Infrared (NIR), and Raman spectroscopy. Each has strengths and limitations, making them
complementary tools for the analysis of pharmaceutical materials. FTIR, the historical work
horse of analytical laboratories, is well-known for its molecular selectivity and resulting
performance in raw materials analysis. However, due to the fragile nature of presently
available instruments and the complications associated with contact sampling and
preparation, FTIR raw materials analysis is currently restricted to the laboratory. The desire
to monitor the physical form of some raw materials and take analysis from the lab to the
sample has spurred the growing adoption of cartable NIR systems. However, NIR’s poorer
selectivity and the impact of physical attributes such as particle size, packing density and
moisture on results pose a significant practical challenge for cost-effective implementation of
the technique. Emerging from recent advances in hardware miniaturization and embedded
analysis is handheld Raman spectroscopy, a technique that combines many of the favorable
attributes of FTIR and NIR. Excellent selectivity, rapid analysis, and unparalleled portability of
the TruScan™ Raman instrument, for instance, now make handheld identification of raw
materials at the loading dock a reality.

These vibrational spectroscopic technologies each have unique strengths and weaknesses
and must be used at the appropriate time and place. A careful selection of technique will
enable necessary analyses to be carried out where needed, and in the most cost-effective
manner. This white paper compares the three techniques, discusses their similarities and
differences, and perhaps more importantly indicates where they should be utilized in order to
maximize their effectiveness in the raw material inspection process.

FTIR Theory
Mid-infrared spectroscopy (mid-IR, commonly configured as FTIR) has for the last 60 years
been heavily used for material identification and authentication.1 When infrared light is
passed through a compound, some wavelengths of the light may be absorbed, while others
merely pass through the sample unaffected. The frequencies of the light which are absorbed
correspond to the vibrational frequencies of the chemical bonds within the molecules of the
sample. The absorption frequencies in FTIR spectroscopy (typically expressed as
wavenumbers (cm-1)) are from 400 cm-1 – 4000 cm-1. A plot of absorbance (or transmittance)
against wavelength is called an “infrared spectrum.”

As absorptions correspond to the vibrational frequencies of different bonds within the

molecule, they can be used to identify a particular functional group, e.g. C-O, C=O, O-H, N-H,
since the vibrational frequencies of these bonds in the mid-infrared range are well known.
Because each bond may have several vibrational frequencies, and a molecule may
incorporate many different bonds, the infrared spectrum of a material may be extremely
complex. Fortunately, it is not necessary to identify, or “assign” every absorption frequency
since it is possible to match an infrared spectrum against others providing they were all

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obtained under the same conditions. Especially interesting for this purpose is the so-called
fingerprint region (900 cm-1 – 2000 cm-1), which usually contains the greatest number of
absorptions.

FTIR Sampling
FTIR absorptions are strong, which offers benefits and poses challenges. Glass absorbs FTIR
wavelengths very strongly, and thus cannot be used to contain materials during FTIR
measurements. Materials must be measured directly, and when using traditional transmission
sampling, it is necessary to dilute the sample by preparing a KBr pellet or Nujol (oil) mull.
More recently, Attenuated Total Reflectance (ATR) sampling techniques have been improved,
and these are now very popular sampling methods for FTIR; they remove the need for much
tedious sample preparation.

Despite recent improvements, current sampling techniques introduce uncertainties into the
analysis. For ATR (the most common sampling method today) the sample of interest must be
in direct contact with the ATR crystal, and only a few microns (a thousandth of a millimeter)
of sample are actually interrogated. This raises significant issues when analyzing
heterogeneous materials. ATR accessories can be difficult to use, somewhat operator-
dependent and breakable. They also introduce characteristic distortions to the FTIR
spectrum and it is necessary to correct for this distortion using a software algorithm before
searching against libraries that have generally been created from transmission measurements.
This correction procedure introduces additional uncertainties to the interpretation.

Traditional FTIR instruments are large, have moving parts and generally require operation via
a computer, making them impractical for use outside the laboratory. While fiber-optics for the
mid-infrared spectral region do exist, they are delicate, have extremely poor transmission
(limiting use to a maximum length of about 1 meter) and are very expensive. For these
reasons, FTIR fiber-optics are not routinely used and it is necessary for the operator to bring
the sample to the instrument for measurement. Regardless of practical difficulties, FTIR is still
extremely popular for identification of raw materials, final products and trouble-shooting for
contamination.

FTIR Interpretation
Since FTIR has exquisite molecular selectivity, it is relatively simple to interpret, making it
useful in sample identification. Compared to NIR and Raman, there are many more reference
texts, correlation charts and electronic libraries containing mid-IR spectra of compounds. A
number of automatic search methodologies can be employed to compare the spectrum of an
unknown sample against many different spectra in a library, including simple peak matching
and discrimination analysis using Euclidean distance or other algorithms. However, analysts
must use caution when interpreting the results. Searching techniques can easily generate
spurious results as the closest known match may actually be significantly different from the
unknown material; the search simply reports the best known match from the spectral library,
even though the unknown may be from a very different class of materials.

Sample FTIR Spectrum

Fingerprint region is useful for ID

Water band interference

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FTIR Application
FTIR instruments are common and can be found in most pharmaceutical companies’ quality
control (QC) laboratories. Its output will most often be used with a peak picking routine or
full-spectrum search to confirm the identity of incoming raw materials. As mentioned
previously, currently available FTIR instrumentation is still relatively large and the limitations
of sample preparation relegate it to continued laboratory use. However, the costs associated
with lab-based FTIR analysis, including sample collection, health and safety issues, quarantine
requirements, and training of highly-skilled lab personnel, have encouraged the investigation
of both NIR and Raman as alternative and complementary techniques.

The strength of FTIR absorptions (compared to NIR) is certainly an advantage when trying to
identify a low level contaminant; and troubleshooting production failures using FTIR
microscopy is common. On the other hand, water’s extremely strong and broad absorbance
in FTIR can pose a problem. Significant amounts of water in a solid sample will likely prevent
some useful information being measured. In addition, measurement of the solute in aqueous
solutions is not possible with FTIR spectroscopy.

NIR Theory
Near-Infrared (NIR) spectroscopy gained favor in recent years largely because of its
convenient and relatively large volume sampling.2 NIR characterizes the material based on its
absorption in the ~ 4000 – 12,500 cm-1 wavelength region, corresponding to vibrational
overtone (harmonic) and combination modes which are much weaker than the fundamental
modes measured in the mid-infrared. Since each fundamental absorption in the mid-infrared
region has several corresponding overtone and combination bands, many of which overlap
and are broadened, NIR spectra are characterized by much broader features than FTIR
spectra and it is often not possible to make direct bond assignments to particular
frequencies. The bands observed in NIR predominantly arise from stretching of O-H, C-H,
and N-H bonds; as these substructures are very common across organic molecules, the
differences between NIR spectra of different compounds are often very subtle, resulting in a
much lower inherent molecular selectivity than FTIR.

Because the sampling method is generally diffuse reflectance (as described below) the
physical nature of the sample is extremely important in NIR analyses as NIR spectra contain
both chemical information (peak shape and position) and physical state information (baseline
slope).

NIR Sampling
The weakness of NIR bands can often be used as an advantage as there is no requirement to
specially prepare a sample as there is in FTIR. Additionally, samples can be analyzed through
some translucent packaging. This has lead to an interest in the use of NIR to identify raw
materials in pharmaceutical quality control.

The most commonly used mode of sampling for pharmaceutical solids with NIR spectroscopy
is diffuse reflectance. Trigger-operated fiber probes (usually a multi-fiber bundle for solids)
and integrating spheres (made of PTFE or gold-coated) are both popular diffuse reflectance
sampling configurations.

However, probes and spheres pose challenges for solids interrogation. Probes can produce
operator-dependent results as the pressure on a powder sample causes baseline movement
in the NIR spectrum. The baseline of the spectrum also rises and falls if there is any
movement in the fibers during sample collection. Integrating spheres necessitate a sample
being collected and placed directly onto the window of the accessory or put into a suitable
container first (albeit an inexpensive one), negating one of NIR’s benefits and exposing the
operator to the material.

NIR Interpretation
NIR bands are generally broad and ill-resolved, lacking the specificity of FTIR so prized by
those seeking to identify chemical spectra. As such, peak picking algorithms are not used to
identify a material. Chemometric techniques such as multivariate discriminant analysis are
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required for sample qualification with NIR data. There are many different methodologies
available, but they can be thought of as methods to reduce the dimensionality of the data;
model the inter- and intra-class dispersion of the data; and thereby classify new samples that
are in accordance with historical training/calibration data.

Extensive and comprehensive library construction is critical to accurate interpretation of NIR

spectra. Generally, a NIR library can be produced by training the system to recognize what is
representative of a sample. However, because NIR spectra are affected by moisture content,
as well as particle size and density, all methods require great care when compiling a library to
ensure that representative materials are used and a full validation procedure takes place.
Otherwise the system may fail quite soon after commissioning, as an acceptable sample may
display some subtle physical change from the samples used to prepare the library. The new
sample may then have to be included as a representative sample and the library updated. The
cost of preparing and maintaining an NIR library is often recognized as one of the most
significant costs of operation.

Sample NIR Spectrum

Broad peaks make interpretation

and match difficult

NIR Application
NIR gained favor during the past 20 years largely because of its convenient and relatively
large volume sampling. Additionally, if information on the particle size or moisture content of
the sample is required, NIR is the preferred technique. In FTIR, sample preparation destroys
the original particle size distribution and samples with a high level of moisture cannot be
measured. A Raman spectrum is largely unaffected by particle size and moisture.

A popular purchase of NIR instrumentation in pharmaceuticals is for the quality control (QC)
laboratory or a similar protected environment. Few systems are truly portable or suitable for
a hostile production environment, and they are most often found in a specially prepared area
within the warehouse, rather than in the loading bay itself.

As with FTIR and Raman, NIR systems must be validated prior to implementation for sample
identity determination. Validation is not an easy task nor a fast one, and it may take many
months before a system is fully prepared for use in the QC environment for one or two raw
materials. In one study, for instance, six compounds tested required a combined 300
reference standards.3 Sites have taken, in some cases, in excess of two years gathering data
sets to validate NIR use. As noted, NIR spectra contain both chemical and physical
information which makes simple and direct interpretation difficult.4 As such, continual library
maintenance is required as subtle differences in particle size and other sample characteristics
have to be captured in the method, leading to the requirement of multiple calibration
reference samples. Along this dimension, NIR compares poorly to Raman and FTIR’s typical
single standard requirement.

Often, a dedicated instrument will be purchased as a “method development” system where

libraries are built and validated before being transferred to other instruments. Transfer to
other instruments is not always straightforward due to NIR’s sensitivity to instrument
variability, and may require library augmentation.

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Raman Theory
In contrast to FTIR and NIR, which are absorption techniques, Raman is a scattering
technique. The sample of interest is illuminated with an intense single wavelength light
source. The light scatters from the sample, most of which is scattered without any change in
wavelength; this is the elastic scatter or Rayleigh-scattered light. A very small proportion of
the light is inelastically (Raman) scattered. The frequency of the Raman-scattered light has
shifted from the original wavelength, with the difference in frequency corresponding to the
vibration frequency of bonds within the molecules of the sample. Typically one photon in 106
– 108 is Raman-scattered; the rest are Rayleigh-scattered or absorbed.

The mechanism causing Raman scatter is different from that which causes FTIR or NIR
absorption. FTIR and NIR require a change in dipole movement in the vibrating bond for
absorption to occur; Raman requires a change in polarizability in the vibrating bond for
Raman scatter to occur. A molecule showing a change in polarizability required to make it
Raman active may or may not show the FTIR/NIR activity (i.e. a change in the dipole). Many
absorptions which are weak in FTIR are strong in Raman; mid-IR and Raman are therefore
said to be complementary. Symmetric vibrations give rise to intense Raman lines; non-
symmetric ones are often weak and sometimes unobservable. In Raman, as in FTIR,
fundamental vibrational modes are interrogated resulting in outstanding molecular selectivity
with little dependence on physical properties such as particle size.

In the past, Raman was not as popular as FTIR and was often relegated to the research and
development lab. Laser sources were large, unreliable, expensive and difficult to maintain,
and sensitivity was very poor requiring many hours for each measurement. The development
of FT-Raman in the late 1980s and early 1990s improved matters significantly, and the
development of high performance optical blocking filters and CCD detectors in the early-mid
1990s was a further significant improvement. The instruments that emerged from these
advances were still large, costly, and suited to only controlled laboratory environments.
Recently, advances in semiconductor lasers, optical miniaturization techniques, compact data
processing capabilities, and probability-based statistical analysis algorithms have combined
to allow the development of compact, rugged, self-contained Raman instruments that can be
reliably used in harsh outside-the-laboratory environments. Ahura Scientific’s TruScan is such
an instrument.

Raman Sampling
One of the greatest strengths of Raman over other technologies is the ease of sampling.
Glass, plastic film and water are very weak Raman scatterers, enabling sampling through
containers and packaging not permitted in FTIR such as a UV cuvette, NMR tube, capillary
tube, vial, plastic bag or bottle. Raman sampling is non-contact and non-destructive and can
be made through the double-bagged internal containment in pharmaceutical drums. Raman
measurement of aqueous solutions is possible with the water being analytically ignored and
the dissolved analytes being interrogated. The sensitivity of contemporary Raman
instrumentation is such that acquisition times of seconds are now the norm; this is
comparable to FTIR and NIR. TruScan’s rugged and compact design coupled with rapid
speed of analysis make handheld operation a reality.

Raman Interpretation
A major benefit of Raman over NIR is its insensitivity to physical form of the sample. This
results in a much simpler approach to data interpretation. Peak picking routines can be used
with Raman libraries because the peaks are distinct and sharp and do not move unless the
chemical moiety is affected. The TruScan approach, based on a patented probabilistic
statistical analysis, is more robust, greatly simplifying method development and allowing fast
implementation of new methods without sacrificing speed or accuracy in routine use.5

Raman measurements may be limited by the phenomenon of fluorescence. Fluorescence

occurs when electrons in an excited molecule return to a relaxed state, emitting excess
energy as photons. Fluorescence appears in the Raman spectrum as a very broad baseline
effect, and may be intense enough to overwhelm the Raman signal in some cases.
Fortunately, the effect is not frequently observed with today’s 785 nm (or longer) Raman
excitation lasers.
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Sample Raman Spectrum

Characteristic sharp peaks make

interpretation and match more robust

Raman Application
The majority of Raman systems found in the pharmaceutical industry are bench-top systems
requiring a high level of skill in their operation. They are still more likely to be found in R&D,
used for troubleshooting, rather than in routine QC.

Ahura Scientific has recognized the need in QC for robust instrumentation which is easy to
use and requires minimal interpretation. Recent advances in light source and detector
technology and major developments in interface design, sampling and algorithms have
enabled newly-designed instruments which no longer require the protection of a laboratory,
configuration by skilled chemometricians, or operation by highly-trained staff. Miniaturization
and improved speed of analysis, for instance, have created the opportunity for
instrumentation to be employed in warehouse and loading dock locations. TruScan
addresses this need with rugged hardware designed to withstand field use, an easy to use
interface, and fast, cost-effective validation and analysis.

Conclusion
All three vibrational spectroscopy techniques have their place in the pharmaceutical industry.
FTIR is still the most widely implemented for ID testing. Its sharp peaks make it ideal for
qualitative analysis, yet its sampling requirements and current size requirements limit it
primarily to lab use. NIR combines both chemical and physical form information in the
spectra and this may be useful for some compounds where physical form affects efficacy.
Although NIR offers sampling advantages as it is non-contact and can analyze through some
container materials, it suffers from lengthy and continual calibration set maintenance making
validation difficult and expensive. The TruScan Raman instrument is sample-container
independent with easily interpreted results and small calibration sets. Raman combines the
selectivity of FTIR and the portability of NIR to make an ideal raw material inspection tool
that will likely see widespread implementation in the pharmaceutical industry.

Ahura Scientific, Inc.

46 Jonspin Road Contact Ahura Scientific for more information on TruScan, the
Wilmington, MA 01887 company’s rugged, handheld Raman system for raw material
+1 978 657 5555 voice verification.
+1 978 657 5821 fax
TruScan@ahurascientific.com

www.ahurascientific.com TSWP01 Copyright 2007, Ahura Scientific, Inc.

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Comparison Summary

FTIR
FTIR Raman
Raman NIR

selectivity High; directly interpretable High; directly interpretable Low; extensive calibration and
spectra spectra chemometrics often required to
distinguish materials; difficult to
interpret directly

interference Measurements strongly influenced Fluorescence baseline can result in Measurements influenced by
by the presence of water longer measurement times; new water; signal impacted by
lasers such as that used in TruScan physical attributes such as
greatly reduce sample fluorescence particle hardness, size and shape

sampling KBr. Nujol mulls, or ATR crystals
with direct sample contact; fiber
optics not readily available
Stand-off sampling through glass
and plastics possible; fiber optics
common; TruScan has relatively
large volume sampling (~2mm spot
at the sample plane)
Can measure through some
materials such as glass; fiber
optics common; large sampling
volumes common

portability Typically large laboratory devices, Typically large, but TruScan is a Most NIR systems are large and
designed for static environment miniaturized and robust option designed for static environment;
some cartable options are
becoming available

method Visual examination often used (no
development development but highly subjective
and irreproducible); similarity scores
common; chemometric
methodologies can also be used
TruScan uses a managed process
for method setup and execution;
automated data management and
user independent statistical analysis
Analysis typically based on
chemometric methods; low
inherent selectivity and sampling
effects makes for laborious and
expensive method development

References

1
J.A. Ryan, S.V. Compton, M.A. Brooks, D.A.C. Compton, Rapid verification of identity and content of drug formulations using mid-
infrared spectroscopy, J. Pharm. Biomed. Anal. 1991, 9, 303-310 and references therein.
2
M. Blanco, J. Coello, H. Iturriaga, S. Maspoch, and C. de la Pezuela, Near-infrared spectroscopy in the pharmaceutical industry,
Analyst 1998, 123, 135R-150R.
3
P. Gemperline et al., Raw materials testing using soft independent modeling of class analogy analysis of near-infrared reflectance
spectra; Anal. Chem. 1989, 61, 138-144
4
M. Ulmschneider, A. Wunenburger, E. Penigualt, Using Near-Infrared Spectrsoscopy for the noninvasive identification of five
pharmaceutical active substances in sealed vials, Analysis 1999, 27, 854-856.
5
C,D. Brown, R.L. Green, Performance Characterization of Material Identification Systems, SPIE 2006, 6378, 637809-1 – 637809-
11.

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