Exploratory data analysis of the dependencies between skin permeability, molecular weight and log P.

Molecular weight and log P remain the most frequently used physicochemical properties in models that predict skin permeability. However, several reports over the past two decades have suggested that predictions made by these models may not be sufficiently accurate. In this study, exploratory data analysis of the probabilistic dependencies between molecular weight, log P and log Kp was performed on a dataset constructed from the combination of several popular datasets. The results suggest that, in general, molecular weight and log P are poorly correlated to log Kp. However, after employing several exploratory data analysis techniques, regions within the dataset of statistically significant dependence were identified. As an example of the applicability of the information extracted from the exploratory data analyses, a multiple linear regression model was constructed, bounded by the ranges of dependence. This model gave reasonable approximations to log Kp values obtained from skin permeability studies of selected non-steroidal ant-inflammatory drugs (NSAIDs) administered from a buffer solution and a lipid-based drug delivery system. A method of testing whether a given drug falls within the regions of statistical dependence was also presented. Knowing the ranges within which molecular weight and log P are statistically related to log Kp can supplement existing methods of screening, risk analysis or early drug development decision making to add confidence to predictions made regarding skin permeability.

Preparation and evaluation of metastable solid-state forms of lopinavir.

In this work, we present the preparation and evaluation of previously unreported metastable forms of the antiretroviral drug, lopinavir. By maintaining the chemical structure, physicochemical properties like the glass transition temperature (T(g)), dissolution and solubility can be readily attributed to the stability of the system. Commercially-available lopinavir was used to prepare partially amorphous crystals, semicrystalline needles, resins and glasses. The physicochemical properties of each were investigated using differential scanning calorimetry (DSC), Fourier transform infrared spectroscopy (FTIR) and powder X-ray diffraction (PXRD). Each sample's thermal and spectroscopic analyses, as well as dissolution and solubility studies were performed one month after sample preparation, for better comparability. Glass transition temperature, activation energy for global molecular mobility (deltaE(Tg)), and activation energy for local molecular mobility (deltaE(beta)) were assessed as primary indicators for structural stability of the systems. Relating these properties to aqueous solubility revealed that each metastable form possessed its own unique equilibrium solubility. Cumulative dissolved fractions (alpha) were fitted against deceleratory kinetics models, and from the data hereby obtained the dissolution process was determined to followed first-order kinetics (R2 = 0.998). From the rate constants, the activation energy for dissolution (deltaE(Diss)) of each sample was calculated. The results suggest that multiple metastable solid-state forms of lopinavir can exist under similar conditions, depending on the preparation conditions.

On the dynamics and function of ciliates in sequencing batch biofilm reactors.

Ciliates are known to directly influence the performance of wastewater treatment plants mainly by feeding on suspended particles. By monitoring two lab-scale sequencing batch biofilm reactors (SBBR), one filled with expanded shale (clay spheres), the other with "Kaldnes" particles (PE-carriers), the succession of biofilm communities with special emphasis on ciliates was monitored for one year. Ciliates were identified and quantified at the species level and compared to rotifer and nematode abundances. Members of the subclass Peritrichia clearly dominated the community of protozoa. Epistylis cf. coronata and Opercularia asymmetrica were the dominant species within this group. The tree-like structure of their colonies provided a distinctive augmentation of the area available for bacterial colonization. The flux of water, produced by E. cf. coronata due to cilia motility, has been visualized and measured by video processing. This flux of water was still measurable at distances > 500 microm and maximum water currents raised up to 180 microm s(-1). Therefore, the role of ciliates is not only restricted to the ingestion of bacteria and suspended particles. They also alter water flux and carry nutrients to the inner parts of the biofilm. Thus, monitoring biofilm formation in wastewater treatment plants should always consider the impact of protists such as ciliates.

Phylogenetic and physiological characterization of a "Nostocoida limicola"-like organism isolated from activated sludge.

Ten strains of "Nostocoida limicola"-like organisms were isolated from a municipal wastewater treatment plant with minor industrial effluent. The near complete 16S rDNA sequences were determined for four strains to analyze the position of the organisms within the phylogenetic tree. All four isolates showed the same 16S rRNA sequence and cluster within the green non sulfur bacteria. Hybridization with oligonucleotide probe AHW183 designed complementarily to diagnostic regions of the 16S rRNA sequences showed a positive reaction with all isolated strains. Hybridization of activated sludge samples with probe AHW183 indicates the filamentous "Nostocoida limicola"-like bacteria commonly to occur inside the sludge flocs and to hardly be detected within the flocs without a specific staining. On HA-medium all strains grow as long coiled, unbranched and unsheathed filaments which usually stain gram positive and show a variable Neisser reaction. The isolates grow well in liquid HA-medium at COD values between 1 to 4 g O2 h(-1) and a pH range from 6.0-7.8. No growth is detected in liquid R2A-medium and several synthetic mineral salts media containing various carbon and nitrogen sources.

Note on Sawilowsky's paper on the rank transform.

It is shown that the undesirable properties of the rank transform in two-way anaylsis of variance with an interaction are due to the test construction and not to the sampling distribution used. It is furthermore confirmed that a safe procedure is to test for main effects only if the interaction is shown not to be significant.

Population analysis in a denitrifying sand filter: conventional and in situ identification of Paracoccus spp. in methanol-fed biofilms.

The microbial community of a denitrifying sand filter in a municipal wastewater treatment plant was examined by conventional and molecular techniques to identify the bacteria actively involved in the removal of nitrate. In this system, denitrification is carried out as the last step of water treatment by biofilms growing on quartz grains with methanol as a supplemented carbon source. The biofilms are quite irregular, having a median thickness of 13 to 20 microns. Fatty acid analysis of 56 denitrifying isolates indicated the occurrence of Paracoccus spp. in the sand filter. 16S rRNA-targeted probes were designed for this genus and the species cluster Paracoccus denitrificans-Paracoccus versutus and tested for specificity by whole-cell hybridization. Stringency requirements for the probes were adjusted by use of a formamide concentration gradient to achieve complete discrimination of even highly similar target sequences. Whole-cell hybridization confirmed that members of the genus Paracoccus were abundant among the isolates. Twenty-seven of the 56 isolates hybridized with the genus-specific probes. In situ hybridization identified dense aggregates of paracocci in detached biofilms. Probes complementary to the type strains of P. denitrificans and P. versutus did not hybridize to cells in the biofilms, suggesting the presence of a new Paracoccus species in the sand filter. Analysis using confocal laser scanning microscopy detected spherical aggregates of morphologically identical cells exhibiting a uniform fluorescence. Cell quantification was performed after thorough disruption of the biofilms and filtration onto polycarbonate filters. An average of 3.5% of total cell counts corresponded to a Paracoccus sp., whereas in a parallel sand filter with no supplemented methanol, and no measurable denitrification, only very few paracocci (0.07% of cells stained with 4',6-diamidino-2-phenylindole) could be detected. Hyphomicrobium spp. constituted approximately 2% of all cells in the denitrifying unit and could not be detected in the regular sand filter. This clear link between in situ abundance and denitrification suggests an active participation of paracocci and hyphomicrobia in the process. Possible selective advantages favoring the paracocci in this habitat are discussed.

Prevalence of viral infections in Mozambican refugees in Swaziland.

The seroprevalence for antibodies to HIV-1, HTLV-1, hepatitis B virus (HBV), hepatitis C virus (HCV) and hepatitis E virus (HEV) were determined in a large group of Mozambican refugees living in Swaziland. Serum samples were collected from a total of 398 refugees located in the two camps (Ndzevane and Malindza). The prevalence for antibodies in the two camps were as follows: Ndzevane: 1.2% (HIV-1); 2.8% (HTLV-1); 0.3% (HCV); 4% (HEV) and 66% for any HBV marker. Malindza: 10.8% (HIV-1); 5.4% (HTLV-1); nil (HCV); 2% (HEV) and 65.7% for any HBV marker. The difference in the HIV-1 seroprevalence between the two camps was statistically highly significant. The phenomenon is possibly related to the location of the Malindza camp in the northern most populous area of Swaziland, resulting in more frequent contact between refugees and the local Swazi population.

Development of an rRNA-targeted oligonucleotide probe specific for the genus Acinetobacter and its application for in situ monitoring in activated sludge.

Enhanced biological phosphate removal in an anaerobic-aerobic activated sludge system has generally been ascribed to members of the genus Acinetobacter. A genus-specific 16S rRNA-targeted oligonucleotide probe was developed to investigate the role of Acinetobacter spp. in situ. Nonisotopic dot blot hybridization to 66 reference strains, including the seven described Acinetobacter spp., demonstrated the expected probe specificity. Fluorescent derivatives were used for in situ monitoring of Acinetobacter spp. in the anaerobic and aerobic compartments of a sewage treatment plant with enhanced biological phosphate removal. Microbial community structures were further analyzed with oligonucleotide probes specific for the alpha, beta, or gamma subclasses of the class Proteobacteria, for the Cytophaga-Flavobacterium cluster, for gram-positive bacteria with a high G + C DNA content, and for all bacteria. Total cell counts were determined by 4',6-diamidino-2-phenylindole staining. In both the anaerobic and the aerobic basins, the activated sludge samples were dominated by members of the class Proteobacteria belonging to the beta subclass and by gram-positive bacteria with a high G + C DNA content. Acinetobacter spp. constituted less than 10% of all bacteria. For both basins, the microbial community structures determined with molecular techniques were compared with the compositions of the heterotrophic saprophytic microbiota determined with agar plating techniques. Isolates on nutrient-rich medium were classified by whole-cell hybridization with rRNA-targeted probes and fatty acid analysis.(ABSTRACT TRUNCATED AT 250 WORDS)

Probing activated sludge with oligonucleotides specific for proteobacteria: inadequacy of culture-dependent methods for describing microbial community structure.

Bacterial community structures in activated sludge samples from aeration tanks of a two-stage system with a high-load first stage and a low-load second stage were analyzed with oligonucleotide probes. The probes were complementary to conserved regions of the rRNA of the alpha, beta, and gamma subclasses of proteobacteria and of all bacteria. Group-specific cell counts were determined by in situ hybridization with fluorescent probe derivatives. Contributions of the proteobacterial subclasses to total bacterial rRNA were quantified by dot blot hybridization with digoxigenin-labeled oligonucleotides. The activated sludge samples were dominated by proteobacteria from the alpha, beta, or gamma subclass. These proteobacteria account for about 80% of all active bacteria found in the activated sludge. For both samples the community structures determined with molecular techniques were compared with the composition of the heterotrophic saprophyte flora isolated on nutrient-rich medium. Probes were used to rapidly classify the isolates and to directly monitor population shifts in nutrient-amended, activated sludge samples. The rich medium favored growth of gamma-subclass proteobacteria (e.g., enterobacteria) and selected against beta-subclass proteobacteria. The culture-dependent community structure analysis of activated sludge produced partial and heavily biased results. A more realistic view will be obtained by using in situ techniques.