Phylogenetic analysis of Histoplasma capsulatum based on partial sequence of the D1/D2 region of the 28S rRNA gene.

In order to confirm the phylogenetic relationships of Histoplasma capsulatum, the partial sequences of large subunit (28S) ribosomal gene (D1/D2 region) of 49 isolates were studied. The similarity values of the 49 isolates were more than 99.0% across 617 base pairs, however, the 49 isolates were divided into 9 groups. These 9 groups were independent of 3 varieties, var. capsulatum, var. farciminosum and var. duboisii. These results showed that analysis of the nucleotide sequence of the 28S rRNA gene was very effective for identification of H. capsulatum and that three varieties of H. capsulatum should be reclassified according to the phylogenetic relationship established from analysis of the D1/D2 region sequences.

Dissolution mechanism of poorly water-soluble drug from extended release solid dispersion system with ethylcellulose and hydroxypropylmethylcellulose.

The purpose of this study is to investigate the release mechanism of poorly water-soluble drug from the extended release solid dispersion systems with water-insoluble ethylcellulose (EC) and water-soluble hydroxypropylmethylcellulose (HPMC) (1:1). Indomethacin (IND) was used as a model of poorly water-soluble drug. Two kinds of solid dispersions were prepared by the solvent evaporation methods, which consist of the same formulation but exhibit different physical performance. It appeared that the dissolution behavior of IND depended on the structures of EC-HPMC matrices, which were governed by the preparation method. In addition, the dissolution behavior showed pH dependency that the dissolution rate of IND was slower in acidic medium than that in neutral medium. The experimental results revealed that the hydrophobic interaction between IND and EC occurred under lower pH and strongly delayed the dissolution rate of IND. The relationship between this hydrophobic interaction and the dissolution rate of IND was also proposed.

Effect of structural relaxation on the physical and aerosol properties of amorphous form of FK888 (NK1 antagonist).

FK888 (NK1 antagonist) is a candidate drug for migraine and selected as a model of amorphous drug. FK888 was micronized to develop as dry powder inhalers (DPIs) taking into consideration of its water insoluble property. The glass transition temperature (Tg) and fragility (m) were 90 degrees C and 118, respectively, and it was categorized as a fragile glass based on Angell's concept. FK888 was structurally relaxed by aging below Tg, then the effect of aging on their physical and aerosol properties were investigated. The investigation on the moisture sorption-desorption isotherms of FK888 indicated that aged FK888 adsorbed less amount of water than that of unaged FK888. This unique moisture sorption-desorption behavior of the aged sample is explained by structural relaxation accompanying decrease of free volume and/or increase of density. As for the dissolution rate of unaged and aged FK888, they showed the similar value, suggesting that there would be no difference in bioavailability. In relation to the stability, FK888 DPIs prepared by unaged and aged FK888 were stored at 70 degrees C, and the respirable fraction of FK888 DPIs was evaluated by using multistage cascade impactor (USP apparatus 3). As a result, the respirable fraction of FK888 DPIs prepared by unaged sample was significantly decreased compared to the aged sample, suggesting that agglomeration may occur in the unaged sample during the storage. This phenomenon was supported by that the unaged sample showed a significant decrease in the surface area compared to that of the aged sample when stored at various conditions.

Quantitative determination method for trace amount of penicillin contaminants in commercially available drug product by HPLC coupled with tandem mass spectrometry.

A quantitative determination method for trace amount of penicillin contaminants in an active pharmaceutical ingredient (API) has been developed. Selective extraction of penicillin contaminants from the matrix containing API and specific separation among penicillin contaminants were achieved through an on-line column switching technique with gradient elution, followed by tandem mass spectrometric determination. Validation was conducted on the developed method in terms of specificity, linearity, accuracy, precision, and detection limit, and appeared reasonable. The detection limit was estimated as 0.03 ng/ml or lower of the concentration of penicillin contaminants in the preparation, corresponding to 4 parts par billion (ppb) against the API. This fulfilled the regulatory requirement by the authorities.

Effect of water content on the solid-state stability in two isomorphic clathrates of cephalosporin: cefazolin sodium pentahydrate (alpha form) and FK041 hydrate.

This study clearly demonstrates that clathrated water molecules can contribute to both chemical stabilization and destabilization of clathrates. The solid-state stabilities for two isomorphic clathrates of cephalosporin, cefazolin sodium and FK041, were investigated in terms of the effects of water content. The isomorphic ranges of water content were estimated to be 3.5-5 mol/mol for alpha-form cefazolin sodium and 2-4 mol/mol for FK041 hydrate. Upon the isomorphic dehydration, alpha-form cefazolin sodium was destabilized as the water content decreased below 4.25 mol/mol owing to the disruption of hydrogen bonding network in lattice channels. In this case, the hydration of clathrated water up to 4.25 mol/mol contributed to the physical and chemical stability of the crystals. On the contrary, the isomorphic hydration in FK041 hydrate contributed to the chemical destabilization owing to the high water activity. The difference in water activity between alpha-form cefazolin sodium and FK041 hydrate could be attributed to the size of water channels.

[Studies on residue on ignition test in JP].

The Residue on Ignition Test specified in General Tests, Japanese Pharmacopeia is a method to measure the weight of the residual substance not volatilized when the sample is ignited under a constant condition. Generally, this test is intended to determine the content of inorganic substances contained as impurities in an organic substance, and, occasionally, to determine the amount of inorganic substances contained as components in an organic substance. Our laboratories have adopted this test method for the above two purposes in the case of the quality control for drug substances, synthetic intermediates, starting materials in the course of drug development. Some validation studies have been conducted for this test in order to clarify the quantitation limit in the case of an application for the former purpose, and trueness and precision in the case of the latter purpose. Consequently, the quantitation limit was determined as around 0.50 mg. The amount of sample should be determined based on this quantitation limit (mg) and the level of quantitation limit (%) or that of specification limit (%) should be considered from the view point of each testing purpose. The test method was also confirmed to have a good trueness and precision as the determination method for an alkaline salt, such as sodium, potassium or calcium salt, of organic compounds, as long as the test is operated with great care not to scatter the content out of crucible during ignition to incineration.