Combined effect of light and temperature on triacylglycerol accumulation in Dicranum elongatum

In the subarctic moss Dicranum elongatum Schleich & Schwaegr., the level of total lipids and triacylglycerols (TAG) was high in late winter and spring and low in autumn and winter. Four-week exposure of field material to continuous light (135μmol m⁻²s⁻¹) at 1°C resulted in a considerable increase in the amount of TAG in the autumn material acclimated to low temperatures and rhythmic light in the field. In contrast, the same treatment did not cause any increase in TAG in the spring material, acclimated to low temperatures and continuous light in the field. Results from experiments, in which moss cultivated for 4 months at 9°C on 12-h photoperiods (135μmol m⁻²s⁻¹) was kept for 3 weeks at low temperatures (9°C and −3°C) either in continuous light (135 or 70 μmol m⁻²s⁻¹) or with 12-h photoperiods (135 μmol m⁻²s⁻¹), indicated that the TAG level was higher at higher light intensity. At 9°C it was also higher in continuous light of both intensities than in rhythmic light. These results strongly suggest that decreasing irradiance and decreasing daylength limits the accumulation of TAG in D. elongatum during autumn in the subarctic.     

The development of non-separation time-resolved fluoroimmunoassays for the measurement of urinary metabolites

We describe the principles of a new generation of sequential or simultaneous time-resolved fluoroimmunoassays, namely, simple, rapid, liquid-phase non-separation procedures which may be applied to the measurement of urinary steroid and drug metabolites. As an example, a method for the measurement of estrone-3-glucuronide in undiluted urine is reported. This method has a similar sensitivity, specificity and accuracy to a conventional separation fluoroimmunoassay or radioimmunoassay but in terms of speed, convenience, precision, reliability and clinical utility the new method has many advantages. The labelled antigen is a novel fluorescent europium chelate covalently linked to estrone-3-glucuronide. The antibody-binding reaction involves the incubation of the labelled antigen (2ng) with a limited concentration of polyclonal or monoclonal antibodies to estrone-3-glucuronyl-6-BSA and an aliquot of standard or sample (undiluted urine; 10 μl) in microtitre wells. After a 10 min incubation, the fluorescence which emanates from the antibody-free label is measured in a time-resolved fluorometer and is proportional to the concentration of estrone-3-glucuronide in the standard or sample. The method may be applied for the monitoring of ovarian function in women.     

Introduction of aliphatic amino and hydroxy groups to keto steroids using O-substituted hydroxylamines.

(Aminooxy)butyl- and -hexylamines and alcohols were synthesized by the Ing-Manske modification of the Gabriel synthesis. The aminooxy group of these heterobifunctional spacer reagents is a far more powerful nucleophile than the amino or hydroxy group because of the oxygen atom adjacent to the amino group (alpha-effect). The aminooxy group reacts readily with keto groups while the amino or hydroxy (or other) group remains free for further reactions. These excellent heterobifunctional spacer reagents were used here to derivatize keto steroids in an alkaline alcoholic solution. Using the described, general, and easy one-step synthesis, aliphatic amino or hydroxy groups with a spacer arm have been introduced to testosterone, 6-ketoestradiol, and cortisol.     

Evaluation of a novel Wave Bioreactor® cellbag for aerobic yeast cultivation

The Wave Bioreactor is widely used in cell culture due to the benefits of disposable technology and ease of use. A novel cellbag was developed featuring a frit sparger to increase the system's oxygen transfer. The purpose of this work was to evaluate the sparged cellbag for yeast cultivation. Oxygen mass transfer studies were conducted in simulated culture medium and the sparged system's maximum oxygen mass transfer coefficient (kLa) was 38 h(-1). These measurements revealed that the sparger was ineffective in increasing the oxygen transfer capacity. Cultures of Saccharomyces cerevisiae were successfully grown in oxygen-blended sparged and oxygen-blended standard cellbags. Under steady state conditions for both cellbag designs, kLa values as high as 60 h(-1) were obtained with no difference in growth characteristics. This is the first report of a successful cultivation of a microbe in a Wave Bioreactor comparing conventional seed expansion in shake flasks and stirred tank bioreactors.     

Time-resolved fluoroimmunoassay of plasma daidzein and genistein

We present a method for the determination of the phytoestrogens daidzein and genistein in plasma (serum). These weakly estrogenic isoflavones occur in soybeans and in smaller amounts in some other beans and plants. It has been suggested that they may afford protection against prostate and breast cancer. The method is based on time-resolved fluoroimmunoassay (TR-FIA) using a europium chelate as a label. After synthesis of 4'-O-carboxymethyl-daidzein and 4'-O-carboxymethyl-genistein the compounds are coupled to bovine serum albumin (BSA), then used as antigens to immunize rabbits. The tracers with the europium chelate are synthesized using the same 4'-O-derivative of the isoflavones. After enzymatic hydrolysis and ether extraction the immunoassay is carried out using the VICTOR 1420 multilabel counter (Wallac Oy, Turku, Finland). The antisera cross-reacted to some extent with some isoflavonoids but not with flavonoids. The cross-reactivity seems not to influence the results, which were highly specific for both compounds. The correlation coefficients between the TR-FIA methods and the reference method based on isotope dilution gas chromatography-mass spectrometry were high; r-values were about 0.95-0.99 depending on concentration. The intra-assay coefficients of variation (CV%) for daidzein and genistein at three different concentrations vary 3.2-4.5 and 3.2-4.1, respectively. The inter-assay CVs vary 5.0-6.3 and 4.5-5.3, respectively. The working ranges of the daidzein and genistein assays are 1.0-216 and 1.7-370 nmol/l, respectively. The plasma values (n = 80) of daidzein and genistein are very low in Finnish subjects (mean for daidzein, 3.8+/-6.8 and for genistein, 3.2+/-7.6 nmol/l; median value for daidzein 1.5 and for genistein 1.4 nmol/l).     

Separation and partial characterization of two alkaline peptidases from cotyledons of resting kidney beans, Phaseolus vulagaris

Extracts prepared from cotyledons of resting kidney beans ( Phaseolus vulgaris L., cv. Processor) rapidly hydrolyzed two dipeptides, Leu-Tyr and Ala-Gly, with pH optima at 9.2 and 8.5, respectively. On ion exchange chromatography on DEAE-Sephacel the two activities eluted as separate peaks, showing that they were due to two different peptidases. The extracts also hydrolyzed Leu-β-naphthylamide optimally at pH 6.4; this activity eluted as a third peak berween the other peaks. The activity peak acting on Leu-Tyr and Ala-Gly rapidly hydrolized two tripeptides, showing that it was an aminopeptidase, whereas the Ala-Gly hydrolyzing peak acted only on dipeptides. The activities against Leu-Tyr and Ala-Gly were also separated by gel chromatography on Sephacryl S-300 with elution positions corresponding to M, values of about 360 000 and 105 000. The aminopeptidase was inhibited by bestatin, and the dipeptidase was inhibited by p-hydroxymercuribenzoate. Both enzymes were inhibited by o-phenanthroline. In most of their properties the two kidneys bean enzymes resembled the alkaline aminopeptidase and the dipeptidase earlier characterized from barley grains.     

Species-specific effects of live roots and shoot litter on soil decomposer abundances do not forecast plant litter-nitrogen uptake

Plant species produce litter of varying quality and differ in the quality and quantity of compounds they release from live roots, which both can induce different decomposer growth in the soil. To test whether differences in decomposer growth can forecast the amount of N species acquire from plant litter, as suggested by theory, we grew individuals of three grassland plants—Holcus lanatus, Plantago lanceolata and Lotus corniculatus—in soils into which ¹⁵N-labelled litter of either Holcus, Plantago or Lotus was added. We measured the effects of live roots and litter of each species on soil microbes and their protozoan and nematode feeders, and to link decomposer growth and plant nutrient uptake, we measured the amount of N taken up by plants from the added litter. We hypothesised that those species that induce the highest growth of microbes, and especially that of microbial feeders, will also take up the highest amount of N from the litter. We found, however, that although numbers of bacterial-feeding Protozoa and nematodes were on average lower after addition of Holcus than Plantago or Lotus litter, N uptake was higher from Holcus litter. Further, although the effects on Protozoa and bacterial- and fungal-feeding nematodes did not differ between the live plants, litter-N uptake differed, with Holcus being the most efficient compared to Plantago and Lotus. Hence, although microbes and their feeders unquestionably control N mineralization in the soil, and their growth differs among plant species, these differences cannot predict differences in litter-N uptake among plant species. A likely reason is that for nutrient uptake, other species-specific plant traits, such as litter chemistry, root proliferation ability and competitiveness for soil N, override in significance the species-specific ability of plants to induce decomposer growth.     

Trampling and Spatial Heterogeneity Explain Decomposer Abundances in a Sub-Arctic Grassland Subjected to Simulated Reindeer Grazing

Mammal grazing is composed of three mechanisms—removal of foliar tissue (defoliation), return of nutrients via dung and urine (fertilization), and trampling. To evaluate the relative role of these mechanisms in the effect of reindeer grazing on soil biota in northern grasslands, we subjected experimental plots in a sub-arctic alpine meadow to defoliation, fertilization (using NPK-solution), simulated trampling, and their factorial combinations once a year from 2002 to 2004 and measured the response of plants and decomposers (including microbes, nematodes, collembolans, and enchytraeids) in 2004. Trampling affected both plant and decomposer communities: the coverage of the moss Pleurozium schreberi and the sedge Carex vaginata, as well as the abundance of collembolans and enchytraeids were reduced in trampled plots. Trampling and fertilization also interacted significantly, with fertilization increasing the abundance of bacteria and bacterial-feeding and omnivorous nematodes in trampled plots only, and trampling decreasing fungal biomass in non-fertilized plots only. Defoliation had no overall effects on plants or decomposers. Nematode genera were not affected by the experimental treatments, but nematode and plant communities were significantly associated, and all decomposer biota, except collembolans, were strongly affected by the spatial heterogeneity of the study site. Our results indicate that trampling may have larger and defoliation and fertilization smaller roles than anticipated in explaining reindeer grazing effects in sub-arctic grasslands. However, even the effects of trampling seem to be outweighed by the spatial heterogeneity of decomposer abundances. This suggests that in sub-arctic grasslands spatial variation in abiotic factors can be a more important factor than grazing in controlling soil biota abundances. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users. Author contributions  LIS was involved in planning of the study, fieldwork, lab work, analysis, and wrote the article with contributions from all other authors; JM and MMK conceived the study and contributed to the data analyses; MMK further contributed to the field work and JM refined the final appearance of the text; JO started and was responsible for managing the field experiment, collecting the plant data and gave advice on statistical analysis.     

Occurrence of acid and neutral carboxypeptidases in germinating cereals

High neutral metallocarboxypeptidase activity (EC 3.4.17) has earlier been detected in young seedlings of rice ( Oryza sativa L.) using benzyloxycarbonyl-L-phenylalanyl-L-alanine (Z-Phe-Ala) as substrate at pH 7. This finding was confirmed, and it was observed that the activity could be assayed with higher specificity and sensitivity by using Z-Gly-Ala or Z-Gly-Phe as substrate at pH 6.5–7. No corresponding activity was detected in seedlings of barley ( Hordeum vulgare L. cv. Himalaya), oats ( Avena sativa L.) or maize ( Zea mays L.). The seedlings of the four cereals possessed similar activities of acid carboxypeptidases (EC 3.4.16; hydrolysis of Z-Phe-Ala and Z-Ala-Phe at pH 5.2 and of Z-Ala-Arg at pH 5.7). However, in endosperms of germinating rice and maize these activities were only about 1–5% of those in barley and oats. A corresponding, although less pronounced, difference was evident between the scutella of the two pairs of cereals. The possible relationship between neutral carboxypeptidase activity and ability to grow in anaerobic conditions is discussed.     

Temperature and soil fertility as regulators of tree line Scots pine growth and survival—implications for the acclimation capacity of northern populations

The acclimation capacity of leading edge tree populations is crucially important in a warming climate. Theoretical considerations suggest that adaptation through genetic change is needed, but this may be a slow process. Both positive and catastrophic outcomes have been predicted, while empirical studies have lagged behind theory development. Here we present results of a 30‐year study of 55,000 Scots pine (Pinus sylvestris) trees, planted in 15 common gardens in three consecutive years near and beyond the present Scots pine tree line. Our results show that, contrary to earlier predictions, even long‐distance transfers to the North can be successful when soil fertility is high. This suggests that present northern populations have a very high acclimation capacity. We also found that while temperature largely controls Scots pine growth, soil nutrient availability plays an important role—in concert with interpopulation genetic variation—in Scots pine survival and fitness in tree line conditions. These results suggest that rapid range expansions and substantial growth enhancements of Scots pine are possible in fertile sites as seed production and soil nutrient mineralization are both known to increase under a warming climate. Finally, as the ontogenetic pattern of tree mortality was highly site specific and unpredictable, our results emphasize the need for long‐term field trials when searching for the factors that control fitness of trees in the variable edaphic and climatic conditions of the far North.