Analytical Applications

A Technical Forum Moderated by Patricia Van Arnum, featuring contributions from PerkinElmer, BioTools, Chiral Technologies, Shimadzu Scientific Instruments, GE Analytical Instruments, and Waters. This article is part of a special issue on analytical technology.
Nov 01, 2011
By Pharmaceutical Technology Editors

Developing methods to analyze drug substances and finished drug products is crucial for ensuring the quality of pharmceutical products. Several industry experts discuss applications in pharmaceutical analysis. Jerry Sellors, IR business manager at PerkinElmer, examines attenuated total internal reflectance sampling in Fourier tranform–infrared spectroscopy. Bo Wang, research scientist, and Laurence A. Nafie, chief technology officer, both with BioTools, and Elena Eksteen, senior business manager, business development and planning at Chiral Technologies, discuss vibrational circular dichroism technology in determining the absolute configuration of enantiomers. Robert H. Clifford, PhD, industrial business unit manager at Shimadzu Scientific Instruments, discusses ultraviolet–visible spectrophotometry in determining the quantitation limit of residual samples in cleaning validation. Richard Godec, new product development manager, Jon Yourkin, pharmaceutical market manager, and Kevin Aumiller, product manager, at GE Analytical Instruments, examine on-line total organic carbon analysis for pharmaceutical water. St. John Skilton, PhD, senior marketing manager, business operations, LSD, pharmaceutical life sciences at Waters, explains the role of hydrogen/deterium exchange with mass spectrometry in biopharmaceutical analysis.

Attenuated internal reflectance sampling in FT–IR

Jerry Sellors, IR business manager at PerkinElmer

Most pharmaceutical laboratories use a Fourier transform–infrared (FT–IR) spectrometer for testing materials. Mid-IR spectroscopy provides rapid and highly specific pharmaceutical ingredient identity testing. IR spectroscopy is used for quantitative and qualitative analysis of solids and liquids, but the most common use for pharmaceuticals analysis is identity confirmation of powdered ingredients. Three common sampling techniques are used: two involving transmission sampling and a third involving attenuated total internal reflectance (ATR).

One approach in transmission sampling is the alkali halide disk method (often referred to as the KBr method), which involves dispersing the sample in a nonabsorbing matrix (e.g., potassium bromide [KBr]) and pressing the mixture into a semitransparent disk to be measured in transmission by passing the IR beam through the disc. Alternatively, in another transmission method, the sample is mixed with a mineral oil and ground into a paste that is pressed into a thin layer between two IR-transparent windows and measured. Both techniques require manual sample preparation, are time-consuming, and may be prone to error. Among all the pharmacopeias, however, these two methods for IR solids sampling are most commonly used. Despite advances in FT–IR technology to provide increased sensitivity, reproducibility, and reliability, developments in sampling apparatus for these techniques have been unremarkable.

Figure 1 (FT–IR): Schematic for attenuated total internal reflectance for infrared (IR) measurement of solids. (FIGURES 1–3(FT–IR) ARE COURTESY OF THE AUTHOR)
Advances in transmission sampling have been slow to develop in part because of the use of another approach, attenuated total internal reflectance (ATR) techniques for IR measurement of solids. With ATR, the sample is pressed into contact with a high refractive index crystal, such as germanium or diamond, and the sample is measured by reflection (see Figure 1). ATR systems often are designed as a central component of the overall FT–IR system. Under ATR, users can more easily acquire good quality IR data from a wide range of samples with minimal sample preparation. ATR technology has improved with respect to crystal design and hardware coupling with FT–IR systems as well as with respect to the software performing automated system performance, suitability, and contamination checks. ATR crystals also can be coupled with mid-IR fiber optics and light-guide systems, where improved sample accessibility is required.

The level of acceptance of the ATR technique among the different pharmacopeias is somewhat mixed. For example, the US Pharmacopiea recognizes the technique as an acceptable alternative, but many other pharmacopeias do not mention ATR. The responsibility is with the user to demonstrate equivalence when this method is chosen to replace existing transmission methods.

Figure 2. (FT-IR): Attenuated total internal reflectance spectra of kaolin at different sample pressures. A is absorbance.
The acceptance of ATR in regulated environments requires a general understanding of ATR because new sources of variation not encountered with conventional transmission measurements can affect the reliability of results. ATR is a reflection technique and can show distinct surface and optical geometry effects that may need to be characterized (1). For example with ATR, IR spectral-band intensities generally change with increasing force applied, and the effect is confounded by an increase in sample penetration depth across the wavelength range measured. This variability can affect quantitative and qualitative IR measurements, which rely on relative band intensities. An example of this can be seen in the ATR spectra of kaolin (see Figure 2) recorded at two different contact pressures, where a silicon–oxygen band in the spectrum of the mineral has a significant shift attributed to deformation of the crystal lattice. Assessing the effects of such variability is important for some materials, where altering pressure can change the degree of sample crystallinity and polymorphic form. Consequently, some ATR devices are designed to provide real-time display of force and spectral intensities before recording the IR spectra so that both can be checked before measurement. Pharmaceutical packaging materials also are frequently analyzed using ATR.

Figure 3: Attenuated total internal reflectance spectra of polylactic acid with rotation of sample. A is absorbance.
In cases where oriented samples are presented to the system, care should be taken to understand the effect or orientation on the FT–IR spectra used in the analysis. This orientation effect is particularly true with molded or extruded polymers. Figure 3 shows the spectra of the surface of a polylactic acid where the only difference between the two measurements is a rotation of the sample on the ATR device. This difference is large enough to cause problems with routine measurements.

As IR use increases for routine measurements of pharmaceuticals, the acceptance of ATR among the pharmacopeias is likely to increase. Using ATR as an alternative to existing transmission methods requires showing evidence of suitability. Controlling and understanding the sources of measurement variation are important. IR instrument packages are being expanded with more sophisticated software routines to improve confidence in ATR results and knowledge-based offerings to assist the user in method development and validation.

FT–IR reference

1. R. Spragg, "Contact and Orientation Effects in FT–IR ATR Spectra, Spectrosc., (Aug. 1, 2011).