Combining Raman Spectroscopy and Microscopy to Support Pharmaceutical Development
Gary McGeorge, Ph.D.
Bristol-Myers Squibb Company
Spectroscopic imaging has seen a significant increase in attention
as a result of two independent processes. Firstly, the introduction
of faster spectroscopic imaging technology has resulted
in very high quality systems that can generate data on a timeframe
of minutes, not hours. Secondly, the FDA has an increased interest in
process analytical technology (PAT) as a tool to augment and support
drug product manufacturing. Consequently, fast, accurate spectroscopic
imaging systems that can analyze materials in a non-destructive
fashion can clearly offer a distinct advantage to the pharmaceutical
manufacturer. In this paper, a brief discussion of the spectroscopic
imaging technique is provided, followed by the analysis of a
drug candidate that crystallized as a mixture of polymorphs. FTRaman
spectra were collected to provide the benchmark spectra for
analysis followed by a restricted wavelength spectroscopic image.
Raman mapping of a whole intact tablet with a FT-Raman system will
serve as an example where the concentration dependence across the
surface of the tablet was established. Both of these examples will
hopefully demonstrate the utility of Raman imaging (mapping) to
obtain additional information that can ensure the quality of a drug
substance or oral dosage form.
Raman mapping and imaging technologies have developed significantly
to the point that more commercial instruments are available on
the market. These microscope-based tools allow for very high-resolution
imaging. Hyperspectral imaging (also known as direct-imaging)
is a technique whereby a monochromator that preserves the spatial
integrity of the image is placed directly in the optical path of a
microscope. These monochromators can be acousto-optically
(AOTF) [1] or liquid crystal (LCTF) tunable filters. Tuning the
monochromator through various wavelengths sequentially allows for
selectivity of the appropriate spectral regions to be probed. These
spectrometers typically use green (514 nm) through to red (765 nm)
laser irradiation. The resultant monochromatically filtered photons
are collected on high sensitivity CCD or CID camera that can further
enhance the utility of such an imaging technology. The power density
compared to high-resolution confocal mapping Raman microscopy
is significantly lower since the laser is distributed over the whole field
of view. As a result, the lower power density is less destructive to
samples.
Mapping technology is more common at the moment for the collection
of spectroscopic images. Mapping systems, on the other hand,
require one to aperture down tightly on the sample and collection of
an entire spectrum at discrete sample positions. Sequentially moving
the sample under the microscope and piecing the points together creates
the image of choice. The spectral resolution can be very high
with this methodology, and it is limited mostly by the spectrograph
used for dispersive systems. However, the spatial resolution is related
to both the aperture used to confine the active region of interest
and the spatial accuracy of the microscope stage movement. Small
apertures reduce the signal, therefore requiring high sensitivity detectors
and more discrete steps to generate the same final image size. As
such, acquiring a Raman spectral image can take many hours. For
hyperspectral imaging, assuming one knows the spectral regions of
interest, only a handful of wavelengths (images) are required for a full
analysis. For the analysis of a moderate Raman scatterer, (where 4
wavelengths might be collected) it may take just a few minutes to collect
the data.
Many Raman spectrometers operate by the Fourier transform principal
(FT) and use near infra-red excitation (1064 nm). By the very
nature of these spectrometers, high-resolution images are not attainable,
and correspondingly would appear to be the perfect candidate
for a macro-imaging system. Macro systems would provide lower
resolution of detail, but at an increased speed. This approach is very
useful if one needs to map an entire tablet, but is unsure of what components
(peaks) to look for. An excellent discussion of the instrumentation
and applications of Raman microscopy is given by Turrell
and Corset [2].
Many of the regulatory requirements during drug substance manufacturing
focus on the chemical identity, purity, impurity profile and
residual solvents from the synthetic process. An increasing number
of examples have demonstrated that, additional to the composition,
one needs to understand the physical nature (polymorphic [3] and
morphologic) of a drug entity. In fact, one main source of morphologic
variety arises from the crystallization of a different polymorph
[4]. The FDA’s perspective and patent protection issues are of considerable
interest to the pharmaceutical manufacturer. The FDA
expects that the crystallization process is in control and that the polymorph
generated will provide predictable properties in-vivo. Likewise
for patent rights, it is important to understand and characterize allknown polymorphs of a particular NCE (new chemical entity). This
requires knowledge of what the polymorph is, preferably via a single
crystal structure or by inference via a combination of physical characterization
techniques involving spectroscopy, powder XRD and
thermal analyses [5]. Polymorphism, by definition, is the ability of a
material to adopt multiple, independent, packing
arrangements, which generates a crystalline lattice. To demonstrate
the importance of polymorphism, the ROY compound, investigated
by Byrn et. al. [6], has six known polymorphs. Each of these polymorphs
has different colors.
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