QUALITATIVE PHYSIOCHEMICAL ANALYSIS
4.3 X-ray Powder Diffraction
4.3.1 Introduction
The X-ray powder diffraction (XRPD) experiment is the foundation of a truly basic material characterisation technique called diffraction analysis, and has been used for many decades with exceptional success to provide accurate information about the structure of materials (Pecharsky and Zavalij, 2005).
Much of what is known about the arrangement and the spacing of atoms in crystalline materials has been determined directly from diffraction studies. XRPD provides a convenient and practical means for the qualitative identification of crystalline
54.80°C
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compounds. It is the only analytical method that is capable of providing qualitative and quantitative information about the compounds present in a solid sample (Skoog et al., 2004).
XRPD materials analysis involves characterisation of the sample by means of atomic arrangements in the crystal lattice. XRPD uses single or multiphase specimens, comprising of a random orientation of small crystallites, each of the order of 1-50 µm in diameter. Each crystallite, in turn, is made up of a regular, ordered array of atoms. An ordered arrangement of atoms (the crystal lattice) contains planes of high atomic density which in turn means planes of high electron density. A monochromatic beam of X-ray photons will be scattered by these atomic electrons and if the scattered photons interfere with each other, diffraction maxima may occur. A diffraction pattern is typically in the form of a graph of diffraction angle (or inter planar spacing) against diffracted line intensity. The pattern is made up of a series of super-imposed diffractograms, one for each unique phase in the specimen. Each of the unique patterns can act as an empirical “fingerprint” for the identification of various phases (Meyers, 2000).
Although powder data usually lack the three dimensionality of a diffraction image, the fundamental nature of the method is easily appreciated from the fact that each powder diffraction pattern represents a one-dimensional snapshot of the three-dimensional reciprocal lattice. The quality of the powder diffraction pattern is usually limited by the nature and the energy of the available radiation, by the resolution of the instrument and by the physical and chemical conditions of the specimen (Pecharsky and Zavalij, 2005).
Of all the methods available to the analytical chemist, only X-ray diffraction is capable of providing general purpose qualitative and quantitative information on the presence of phases in an unknown mixture. A diffraction pattern contains a good deal of information of which three parameters are of special interest:
The position of the diffraction maxima.
The peak intensities.
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The intensity distribution as a function of diffraction angle.
These three species of information can, in principle, be used to identify and quantify the contents of the sample, as well as to calculate the material‟s crystallite size and distribution, crystallinity, stress and strain (Meyers, 2000).
The role of XRPD for identification of unknown chemicals is limited in the pharmaceutical industry, since there are numerous techniques available for determining molecular structure of organic molecules. The techniques of mass spectrometry and nuclear magnetic resonance are predominant in this role. On the other hand, due to its speed of data acquisition and sensitivity XRPD exists as the primarily tool for phase identification (Stephenson, 2005; Beckers, 2004). While it does serve as a means for identification of primarily inorganic unknowns, by non-destructively determining the crystallographic constitution of samples, XRPD is used to:
Determine crystal structures.
Screen for polymorphs or hydrates.
Detect changes in morphology or crystalline state of active ingredients (e.g., during processing or at non-ambient conditions).
Detect and quantify crystalline impurities (in some cases down to 0.05%).
Determine the crystallinity or the crystallite size of a compound.
Analyse and optimise final dosage forms.
XRPD data on a compound or drug is also required for new product registration and patent application/protection (Litteer and Beckers, 2005).
4.3.2 Methodology
XRPD patterns for cyclo(Phe-4Cl-Pro) and cyclo(D-Phe-4Cl-Pro) were obtained by using an Bruker Advanced Solutions XRD Commander (Diffracplus Version 2.3) diffractometer (Bruker Advanced Solutions, Germany), and the resultant diffraction
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patterns were interpreted using Bruker Advanced X-ray Solutions Eva Software (Diffracplus Version 10.0 Rev.1). n-Hexane (BD Laboratory Supplies, England) was used as a wetting agent, which enabled small samples of the cyclic dipeptide to adhere to silicon plates, without causing dissolution of the compounds. The silicone plates were then placed within the X-ray powder diffractometer and scanned for 1 hour.
4.3.3 Results and discussion
The diffraction patterns obtained from powdered samples can only be identified when compared alongside the diagrams of known substances until a match is obtained (Willard et al., 1998).
The XRPD patterns of cyclo(Phe-4Cl-Pro) and cyclo(D-Phe-4Cl-Pro) are illustrated in Figures 4.7 and Figures 4.8, respectively. Determining the phase composition of the cyclic dipeptides was not possible, since none of the powder diffraction databases available included experimental diffraction patterns to which the compounds could be compared. Since the obtained Bragg angles and intensities of the cyclo(Phe-4Cl-Pro) and cyclo(D-Phe-4Cl-Pro) have no known match, the collected data can potentially be used as a “fingerprint” to identify new crystalline substances. The identity of the cyclic dipeptides should, however, firstly be confirmed through structural elucidation to ensure that it is not a mixture of compounds.
From the diffraction patterns of both cyclic dipeptides it can be seen that they are crystalline solids, since the curve of scattered intensity versus 2θ for crystalline solids is zero everywhere except at certain angles where sharp maxima occur.
Both morphous solids and liquids have structures characterised by an almost complete lack of periodicity and tendency to order. The atoms in amorphous structures also show preference for a particular interatomic distance, resulting in x-ray diffractograms exhibiting only one or two broad maxima.
66 Figure 4.7 X-ray powder diffraction pattern of cyclo(Phe-4Cl-Pro)
Cyclo(phe-4Cl-pro)
67 Figure 4.8 X-ray powder diffraction pattern of cyclo(D-Phe-4Cl-Pro)
cyclo(D-phe-4Cl-pro) 10-08-10
Operations: Import
cyclo(D-phe-4Cl-pro) 10-08-10 - File: cyclo(D-phe-4Cl-pro) 10-08-10.raw - Type: 2Th/Th locked - Start: 5.000 ° - End: 50.000 ° - Step: 0.020 ° - Step time: 2. s - Temp.: 25 °C (Room) - Time Started: 11 s - 2-Theta: 5
Lin (Counts)
0 100 200 300 400 500 600 700 800 900 1000 1100 1200 1300
2-Theta - Scale
5 10 20 30 40 50
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