Identifying polymorphs

Different crystal structures can be identified by their melting points, so in principle by scanning a sample in a DSC the different crystal forms present can be distinguished. Whereas in the above application the aim is to prevent recrystalHsation, when recrystalhsation does occur it may be of interest to characterise the forms produced, and so identify a number of different polymorphs. By scanning at a range of different rates, different forms may be produced depending upon the kinetics of the crystallisation reaction. Therefore, it can be informative to scan a sample from slow to high rates to see what transitions can be found. 

Where a material does not recrystaUise on cooling or where the cooling rate is sufficient to prevent any recrystalhsation, it may cool into a glass, and the Tg can be measured on heating. 

Subsequent crystallisation may occur with these materials and melting measurements can then be made. 

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Fig. 8.2 Single nucleotide polymorphisms of the human MDR1 gene (compiled from [10, 11, 97]). The locations of the identified polymorphisms (white arrow non-coding polymorphisms black arrow polymorphisms leading to amino acid exchanges) in relation to the exon-intron...

Laser excitation causes characteristic photon emissions from the solid phase, identifying polymorphic form and transformation rates. This technique is only suitable for non-fluorescing materials... 

F. Laplant and A. De Paepe, Raman spectroscopy for identifying polymorphs, in Pharmaceutical Applications of Raman Spectroscopy, S. Sasic (Ed), Technology for the Pharmaceutical Industry Series, John WUey Sons, Ltd, Hoboken, 2008. 

Association studies, which can be used to identify genes of small effect, have also been used in studies of adult-onset schizophrenics to identify polymorphisms in the SHTA receptor gene and the DRD3 receptor gene, which may be involved in schizophrenia (Owen, 2000). 

Gene expression, a determinant of a cell s characteristics, is another phenotype that can be studied using lymphoblastoid cell lines. Studies have shown that gene expression levels in humans differ not only among cell types within an individual, but also among individuals (16,31). As a result, there have been several recent studies that have identified polymorphic genetic variants that influence gene expression levels (15,25,32). 

The majority of all APIs manufactured demonstrate structural polymorphism. In order to formulate a drug product that is physically and chemically stable, the formulation team must identify the most thermodynamically stable polymorph of the API. By identifying the proper polymorph, the patient s need for a drug product with reproducible bioavailability during the course of typical and atypical shelf-life conditions will be met. This section will review the use of spectroscopic techniques for identifying polymorphism of APIs during API crystallization and formulation. For a more comprehensive discussion of polymorphism the reader is directed to a work by Singhal85 and references cited therein. 

Mui S, Briggs M, Chung H, et al. 1996. A newly identified polymorphism in the apolipoprotein E enhancer gene region is associated with Alzheimer s disease and strongly with the E 4 allele. Neurology 47 196-201. 

The effects of different rates of cooling down to isothermal crystallization temperature, different crystallization temperatures and different rates of heating during melting, on polymorph formation and transformation are easily studied by DSC. However, it is not possible unequivocally to identify polymorphs by DSC this must be done by X-ray diffractometry (Rossell, 2003 see below). 

However, it is often very difficult to grow single crystals of suitable size for a successful diffraction experiment and the researcher has to resort to powder diffraction experiments on a polycrystalline sample. Powder patterns are fingerprints of the solid materials and are therefore used to identify polymorphs. In the pharmaceutical industry, and in associated laboratories, it is becoming common practice to accompany the routine quality control analyses on the production fine with the measurements of powder patterns. 

X-ray diffractometry Identifying polymorphs quantification of degree of crystallinity crystal lattice geometry and solid-state transformations... 

In 2004, the FDA approved a microarray chip designed to routinely identify polymorphisms of drug-metabolizing enzymes related to cytochrome P450 drug metabolism. 

Hyperserotonemia has been a consistent finding in subjects with autism, which may be due to activity of serotonin-associated platelet proteins (Hranilovic et al., 2008, 2009). Interestingly, 99% of blood serotonin is contained in platelets (Anderson et al., 1987) and studies have shown that there is an approximate 50% increase in blood-levels of serotonin in subjects with autism vs. controls (McBride et al., 1998). Hypotheses for increased serotonin include increased synthesis of serotonin by tryptophan hydroxylase (TPHl), increased uptake of serotonin into platelets via serotonin transporters (5-HTT), diminished release of serotonin from platelets via serotonin 2A receptor, and decreased breakdown of serotonin by monoamine oxidase (MAOA) (Hranilovic et al., 2008). A study by Hranilovic et al. (2008) identified polymorphisms of tryptophan hydroxylase and MAOA with increased serum serotonin levels. Similarly, haplotype analysis has shown a significant association between polymorphisms of TPHl and increased serotonin in whole blood (Cross et al., 2008). 

The inconsistency of these results may be attributable to differences in populations, such as ethnicity and ascertainment of the families studied and/or differences in definition of phenotype. However the linkage between 5q31 and atopy and asthma phenotypes is the most reproduced of all the possible linkages reported. Based on the results of 5q31—33 linkage studies, studies have begun to identify polymorphisms in candidate genes at this locus and to examine possible association of these polymorphisms with disease traits. 

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