Quantitative analysis of natural cyclodextrins by high-performance liquid chromatography with pulsed amperometric detection: application to cell permeation study

Simple HPLC-PAD methods were developed for quantitation of cyclodextrins (CDs) in aqueous matrices from in vitro cell permeation studies. C-18 solid-phase extraction was used for sample pretreatment. Samples were analysed using acetonitrile-water mobile phase with post-column alkalization by 0.5M NaOH. Zorbax SB-Aq (for alpha-CD) and Zorbax SB-Phenyl (for beta-CD and gamma-CD) columns gave excellent peak shape and sufficient resolution of CD to glucose (2.7-3.2). The methods showed good concentration-response relationship (r > or = 0.999), precision (RSD% 0.7-5.1), repeatability (RSD% 3.4-13.7) and accuracy (87-107%). The limits of quantitation were 0.78, 0.46 and 0.52 microg/ml for alpha-CD, beta-CD and gamma-CD (RSD% of 10.6, 8.1 and 16.3, respectively).     

Toxicity of Selected Cationic Drugs in Retinoblastomal Cultures and in Cocultures of Retinoblastomal and Retinal Pigment Epithelial Cell Lines

Tamoxifen and toremifene are antiestrogenic drugs successfully used in the therapy of breast cancer. Rheumatoid arthritis and malaria have been treated with chloroquine for decades. Unfortunately, tamoxifen and chloroquine are reported to induce retinal changes as a side effect. We now studied the effects of tamoxifen, toremifene, and chloroquine on the viability of the human retinoblastomal cell line Y79, using the WST-1 test or measurement of the cellular ATP content. The studies were made on Y79 cell cultures and on cocultures of Y79 cells and retinal pigment epithelial cell line ARPE-19. The cocultures were used to clarify the effect of retinal pigment epithelium on toxicity to Y79 cells. In the coculture, the drugs were applied to ARPE-19 cells growing in the culture inserts on top of Y79 cells and the viability of ARPE-19 and Y79 cells was assessed separately. Tamoxifen, toremifene, and chloroquine reduced dose-dependently the viability of Y79 cells after 24-h exposure. The ARPE-19 cells proved to be protective after chloroquine exposure in the coculture. The results shed light on the toxicity of tamoxifen and chloroquine in Y79 cells in vitro. With the coculture we were able to simulate the in vivo route of chloroquine to the retina via the retinal pigment epithelium.     

Thermal responses during and after whole-body cryotherapy (−110°C)

The effect of whole-body cryotherapy (WBC) on rectal and skin temperatures was measured in healthy subjects before, during and after WBC exposure. WBC did not cause any significant change in rectal temperature. The lowest local skin temperatures were recorded in the forearm, 5.2 (2.8)°C, and in the calf, 5.3 (3.0)°C. WBC involves no risk for frostbites. After WBC, all skin temperatures recovered rapidly, indicating that the analgetic effects of WBC only occur during a limited period after the exposure.     

Frost hardiness of mycorrhizal (Hebeloma sp.) and non-mycorrhizal Scots pine roots

The frost hardiness (FH) of mycorrhizal [ectomycorrhizal (ECM)] and non-mycorrhizal (NM) Scots pine (Pinus sylvestris) seedlings was studied to assess whether mycorrhizal symbiosis affected the roots’ tolerance of below-zero temperatures. ECM (Hebeloma sp.) and NM seedlings were cultivated in a growth chamber for 18 weeks. After 13 weeks’ growth in long-day and high-temperature (LDHT) conditions, a half of the ECM and NM seedlings were moved into a chamber with short-day and low-temperature (SDLT) conditions to cold acclimate. After exposures to a range of below-zero temperatures, the FH of the roots was assessed by means of the relative electrolyte leakage test. The FH was determined as the inflection point of the temperature-response curve. No significant difference was found between the FH of mycorrhizal and non-mycorrhizal roots in LDHT (?8.9 and ?9.8 °C) or SDLT (?7.5 and ?6.8 °C). The mycorrhizal treatment had no significant effect on the total dry mass, the allocation of dry mass among the roots and needles or nutrient accumulation. The mycorrhizal treatment with Hebeloma sp. did not affect the FH of Scots pine in this experimental setup. More information is needed on the extent to which mycorrhizas tolerate low temperatures, especially with different nutrient contents and different mycorrhiza fungi.     

Aging three-spined sticklebacks Gasterosteus aculeatus: comparison of estimates from three structures

In order to assess the accuracy and reliability of age estimates from calcified structures in the three-spined stickleback Gasterosteus aculeatus, we evaluated intra and inter-reader repeatability from three structures: otoliths, gill covers and pelvic spines). Average age estimates were also compared between the structures. The overall intra-reader repeatability of age estimates were highest for otoliths (69%), lowest for gill covers (53%) and intermediate for spine cross-sections (63%). Although four of the seven readers had the highest intra-reader repeatability score for spine cross-sections, the inter-reader variance in this structure was much higher than in others. Otoliths were the easiest in terms of their pre-analysis treatment and exchange of materials (as digital images) between readers. In addition, otoliths are more well-studied compared with the other structures with respect to their development through ontogenesis; hence, age estimates based on otoliths should be the most reliable. Therefore, our recommendation is that whenever possible, analysis of otoliths should be the preferred approach for aging G. aculeatus.     

Characterization of a thermophilic sulfur oxidizing enrichment culture dominated by a Sulfolobus sp. obtained from an underground hot spring for use in extreme bioleaching conditions

A thermoacidophilic elemental sulfur and chalcopyrite oxidizing enrichment culture VS2 was obtained from hot spring run-off sediments of an underground mine. It contained only archaeal species, namely a Sulfolobus metallicus-related organism (96% similarity in partial 16S rRNA gene) and Thermoplasma acidophilum (98% similarity in partial 16S rRNA gene). The VS2 culture grew in a temperature range of 35–76°C. Sulfur oxidation by VS2 was optimal at 70°C, with the highest oxidation rate being 99 mg S⁰ l⁻¹ day⁻¹. At 50°C, the highest sulfur oxidation rate was 89 mg l⁻¹ day⁻¹ (in the presence of 5 g Cl⁻ l⁻¹). Sulfur oxidation was not significantly affected by 0.02–0.1 g l⁻¹ yeast extract or saline water (total salinity of 0.6 M) that simulated mine water at field application sites with availability of only saline water. Chloride ions at a concentration above 10 g l⁻¹ inhibited sulfur oxidation. Both granular and powdered forms of sulfur were bioavailable, but the oxidation rate of granular sulfur was less than 50% of the powdered form. Chalcopyrite concentrate oxidation (1% w/v) by the VS2 resulted in a 90% Cu yield in 30 days.     

Dynamics of voles and small mustelids in the taiga landscape of northern Fennoscandia in relation to habitat quality

In the forests of northern Fennoscandia during the I980's, the dynamics of microtine rodents changed from multiannual high amplitude fluctuations (cycles) to, depending on species, fluctuations with a strong seasonal component or fluctuations with smaller amplitude and lower frequency. Microtine and predator data from the Pallasjarvi area, Finnish Lapland, suggest that this transition took place at different rates in different parts of the taiga landscape. Generally, densities in forest habitats have been primarily seasonal since 198S-86. In mesic spruce taiga and in drier forest habitats microtines had a prolonged peak in 1981-83 and a crash in 1984-83. At the timberline, however, microtine populations dropped from peak to low densities already in 1982-83 but the final crash did not occur until spring 1985. The synchronous decrease in microtines densities in all habitat types in 1984-85 coincided with increase in weasel activity. Activity of other carnivores was consistently high in mesic lowland habitats. The data support following three conjectures. 1) Periodic abundance of least weasels is crucial for sustained vole cycles. 2) Predominance of stoats and other generalist predators lead to less regular fluctuations with a strong seasonal component where density declines occur in autumn and early winter. 3) In barren tundra areas, the vegetation cannot sustain high densities of microtines and. consequently, predation is not a necessary condition for population crashes.     

Molecular Evolution of Puumala Hantavirus

Puumala virus (PUUV) is a negative-stranded RNA virus in the genus Hantavirus, family Bunyaviridae. In this study, detailed phylogenetic analysis was performed on 42 complete S segment sequences of PUUV originated from several European countries, Russia, and Japan, the largest set available thus far for hantaviruses. The results show that PUUV sequences form seven distinct and well-supported genetic lineages; within these lineages, geographical clustering of genetic variants is observed. The overall phylogeny of PUUV is star-like, suggesting an early split of genetic lineages. The individual PUUV lineages appear to be independent, with the only exception to this being the Finnish and the Russian lineages that are closely connected to each other. Two strains of PUUV-like virus from Japan form the most ancestral lineage diverging from PUUV. Recombination points within the S segment were searched for and evidence for intralineage recombination events was seen in the Finnish, Russian, Danish, and Belgian lineages of PUUV. Molecular clock analysis showed that PUUV is a stable virus, evolving slowly at a rate of 0.7 x 10(-7) to 2.2 x 10(-6) nt substitutions per site per year.