Late Holocene lake-level fluctuations in Walker Lake, Nevada, USA

Walker Lake, a hydrologically closed, saline, and alkaline lake, is situated along the western margin of the Great Basin in Nevada of the western United States. Analyses of the magnetic susceptibility (χ), total inorganic carbon (TIC), and oxygen isotopic composition (δ¹⁸O) of carbonate sediments including ostracode shells (Limnocythere ceriotuberosa) from Walker Lake allow us to extend the sediment record of lake-level fluctuations back to 2700 years B.P. There are approximately five major stages over the course of the late Holocene hydrologic evolution in Walker Lake: an early lowstand (> 2400 years B.P.), a lake-filling period ( 2400 to  1000 years B.P.), a lake-level lowering period during the Medieval Warm Period (MWP) ( 1000 to  600 years B.P.), a relatively wet period ( 600 to  100 years B.P.), and the anthropogenically induced lake-level lowering period (< 100 years B.P.). The most pronounced lowstand of Walker Lake occurred at  2400 years B.P., as indicated by the relatively high values of δ¹⁸O. This is generally in agreement with the previous lower resolution paleoclimate results from Walker Lake, but contrasts with the sediment records from adjacent Pyramid Lake and Siesta Lake. The pronounced lowstand suggests that the Walker River that fills Walker Lake may have partially diverted into the Carson Sink through the Adrian paleochannel between 2700 to 1400 years B.P.     

Structural characterization and antioxidant activity of a polysaccharide from the fruiting bodies of cultured Cordyceps militaris

The water-soluble crude polysaccharides were obtained from the fruiting bodies of cultured Cordyceps militaris by hot water extraction followed by ethanol precipitation. The polysaccharides were successively purified by chromatography on DEAE–cellulose-52 and Sephacryl S-100 HR columns, giving main three polysaccharide fractions termed P50-1, P70-1, and P70-2. Structural features of P70-1 were investigated by a combination of chemical and instrumental analysis, such as partial acid hydrolysis, methylation analysis, periodate oxidation – Smith degradation, GC–MS, ¹³C NMR, HPAEC-PAD, and FT-IR. The results indicated that P70-1 has a backbone of (1 → 6)-linked β-d-mannopyranosyl residues, which occasionally branches at O-3. The branches were mainly composed of (1 → 4)-linked -d-glucopyranosyl and (1 → 6)-linked β-d-galactopyranosyl residues, and terminated with β-d-galactopyranosyl residues and -d-glucopyranosyl residues. In the in vitro antioxidant assay, P70-1 was found to possess hydroxyl radical-scavenging activity with an IC₅₀ value of 0.548 mg/ml.     

The role of calcium channels in substance P-induced contractile response in the rat iris

This study was undertaken to assess the role of calcium channels in the contractile response induced by substance P in the isolated rat iris. Substance P produced graded and sustained contraction in the rat iris. Pre-incubation of preparations with thapsigargin (1 μM), verapamil (1 μM), isradipine (1 μM) or with ω-conotoxin MCIIA (0.1 μM) did not significantly inhibit substance P-mediated contraction in the isolated rat iris. However, pre-incubation of the preparations with nicardipine (1 μM) or ruthenium red (1 mM) caused parallel displacement to the right of the substance P concentration–response curve without affecting its maximal response. In contrast, amiloride (1 μM), markedly inhibited substance P-mediated contraction (73±5%), while econazole (1 mM) also significantly inhibited (44±11%) substance P-mediated contraction in the isolated rat iris. Collectively, these results suggest that substance P-mediated contractile response in the isolated rat iris depends largely on the influx of external Ca²⁺, by a mechanism which might involve the T-type calcium channels.     

The activities of several detoxication enzymes are differentially induced by juices of garden cress, water cress and mustard in human HepG2 cells

It has been previously demonstrated in a human-derived hepatoma cell line (HepG2) that juices from cruciferous vegetables protect against the genotoxicity caused by dietary carcinogens. HepG2 cells possess different enzymes involved in the biotransformation of xenobiotics. Therefore, we investigated the effect of cruciferous juices on the activities of CYP 1A and several phase II enzymes in this cell model. For each experiment, 1 × 10⁶ cells were seeded on Petri dishes. After 2 days, the juices (0.5–8 μl/ml of culture medium) were added for 48 h prior to cell harvesting. The addition of juice from water cress (Nasturtium officinalis R. Br) significantly increased the activities of ethoxyresorufin-O-deethylase at high doses only and NAD(P)H-quinone reductase in a dose-dependent manner (1.8- and 5-fold, respectively). The addition of juice from garden cress (Lepidum sativum L.) significantly increased the activities of NAD(P)H-quinone reductase and UDP-glucuronosyl-transferase with a maximal effect around the dose of 2 μl/ml juice (1.4- and 1.2-fold, respectively) while the other enzymes were not altered. Mustard (Sinapis alba L.) juice increased the activities of NAD(P)H-quinone reductase (2.6-fold at the dose of 8 μl/ml), and N-acetyl-transferase (1.4-fold at the dose of 8 μl/ml) in a dose-dependent manner while a maximal induction of UDP-glucuronosyl-transferase was obtained with a dose of 2 μl/ml (1.8-fold). These observations show that the three juices have different induction profiles: only water cress acted as a bifunctional inducer by enhancing both phase I and phase II enzymes. As a consequence, each juice may preferentially inhibit the genotoxicity of specific compounds.     

Dithioerythritol (DTE) prevents inhibitory effects of triphenyltin (TPT) on the key enzymes of the human sex steroid hormone metabolism

Organotins are known to induce imposex (pseudohermaphroditism) in marine neogastropods and are suggested to act as specific endocrine disruptors, inhibiting the enzyme-mediated conversion of steroid hormones. Therefore, we investigated the in vitro effects of triphenyltin (TPT) on human 5-reductase type 2 (5-Re 2), cytochrome P450 aromatase (P450arom), 17β-hydroxysteroid dehydrogenase type 3 (17β-HSD 3), 3β-HSD type 2 and 17β-HSD type 1 activity. First, the present study demonstrates that significant amounts of TPT occurred in the blood of eight human volunteers (0.17–0.67 μg organotin cation/l, i.e. 0.49–1.92 nmol cation/l). Second, TPT showed variable inhibitory effects on all the enzymes investigated. The mean IC₅₀ values were 0.95 μM for 5-Re 2 (mean of n=4 experiments), 1.5 μM for P450arom (n=5), 4.0 μM for 3β-HSD 2 (n=1), 4.2 μM for 17β-HSD 3 (n=3) and 10.5 μM for 17β-HSD 1 (n=3). To exclude the possibility that the impacts of TPT are mediated by oxidizing essential thiol residues of the enzymes, the putative compensatory effects of the reducing agent dithioerythritol (DTE) were investigated. Co-incubation with DTE (n=3) resulted in dose-response prevention of the inhibitory effects of 100 μM deleterious TPT concentrations on 17β-HSD 3 (EC₅₀ value of 12.9 mM; mean of n=3 experiments), 3β-HSD 2 (0.90 mM; n=3), P450arom (0.91 mM; n=3) and 17β-HSD 1 (0.21 mM; n=3) activity. With these enzymes, the use of 10 mM DTE resulted in an at least 80% antagonistic effect, whereas, the effect of TPT on 5-Re 2 was not compensated. In conclusion, the present study shows that TPT acts as an unspecific, but significant inhibitor of human sex steroid hormone metabolism and suggests that the inhibitory effects are mediated by the interaction of TPT with critical cysteine residues of the enzymes.     

Examination of cyclic changes in bovine luteal echotexture using computer-assisted statistical pattern recognition techniques

B-mode sonography is a well-established diagnostic tool for determination of cycle stage in gynaecology. The aim of this study was to determine whether computer-assisted texture analysis of B- mode sonographic images of bovine luteal glands provides further information about the animal's plasma progesterone concentration and cycle stage. Four Simmenthal cows were examined during two consecutive estrous cycles with an ultrasound device equipped with a 7.5 MHz microconvex probe. During each examination three B-mode images of the corpus luteum (CL) were digitized and analyzed off-line using a computer-assisted texture analysis program. Size, echogeneity, and echotexture of the CL were characterized by the following texture parameters: area of cross-sectional planes of the CL (A), mean gray level (MGL), correlation (CORR), run percentage (RPERC), and long-run emphasis (LREM). Plasma progesterone levels (P4) were also determined. All parameters showed characteristic changes during the estrous cycle (P < 0.05). Variance component estimates for the effect of Day of estrous cycle on A, MGL, CORR, RPERC, and LREM were 56.6%, 64.6%, 77.6%, 89.9%, and 86.0%, respectively, and 20.6%, 24.5%, 7.2%, 0.0%, and 14.0% for the influence of the individual cow. The factor estrous cycle within cows was responsible for 22.8%, 10.9%, 15.2%, 10.1%, and 0.0% of the variability of A, MGL, CORR, RPERC and LREM values, respectively. Cyclic changes were similar in A and P4. In contrast to P4, which decreased already between Days –5 and –3 (Day 0 = ovulation), A stayed at constant high values until Day –3. Mean MGL values were higher (P < 0.05) on Days 7, 9, and 13 compared to Days 3 and –3. Mean CORR values were constantly high (P > 0.05) during the first days after ovulation and decreased continuously (P < 0.05) between Days 5 and 13. Thereafter, mean CORR values remained low (P < 0.05) until the next ovulation, except on Day –3 (P < 0.05). Mean RPERC rose between Days 1 and 9 from low to high values (P < 0.0001) remained at these high values (P > 0.05) between Days 9 and 15, and decreased (P < 0.05) afterwards to baseline values on Day –1. Mean LREM inclined steeply (P < 0.0001) from minimum to maximum between Days 1 and 5. From Days 7 to –3, mean LREM remained (P > 0.05) at a constant level close below the maximum value, and decreased to baseline values on Day –1. The results of this study show that statistical pattern recognition techniques provide new information about the luteal glands, thus facilitating a more accurate differentiation between different cycle stages in cows.     

Dual-acting agents that possess reversing resistance and anticancer activities: Design, synthesis, MES-SA/Dx5 cell assay, and SAR of Benzyl 1,2,3,5,11,11a-hexahydro-3,3-dimethyl-1-oxo-6H-imidazo[3',4':1,2]pyridin[3,4-b]indol-2-substitutedacetates

Based on the structural analysis of fumitremorgin C (FTC), imidazoline and β-carboline amino acid benzylester, 14 novel 2-substitutedtetracyclic derivatives of tetrahydrocarboline 4a–n were prepared. We demonstrated that the exposure of MES-SA/Dx5 cells to some of 4a–n resulted in significant reduction of resistance of the cells against doxorubicin. This reduced resistance was accompanied by lowering of IC₅₀ value to doxorubicin from 1.55 ± 0.26 μmol/L to 0.33 ± 0.05 μmol/L for 2-(2-butyl)-derivative 4c, to 1.03 ± 0.22 μmol/L for 2-methyl-derivative 4d, to 0.46 ± 0.04 μmol/L for 2-benzyl-derivative 4f, to 0.98 ± 0.25 μmol/L for 2-indole-3-yl-methyl-derivative 4h, to 0.36 ± 0.03 μmol/L for 2-benzyloxycarbonylmethyl-derivative 4i, to 0.77 ± 0.08 μmol/L for 2-benzyloxycarbonylethyl-derivative 4j, and to 0.77 ± 0.08 μmol/L for 2-benzyloxycarbonylamino-n-butyl-derivative 4l. Proliferation assays of 4a–n indicated 4c,f,i,j were able to inhibit the proliferation of doxorubicin resistant MES-SA/Dx5 cells. The SAR analysis revealed that the benzylester form and the tetracyclic structure of 4a–n were critical for both sensitizing doxorubicin and the cellular anti-proliferative effect.     

Reducing plant abiotic and biotic stress: Drought and attacks of greenbugs, corn leaf aphids and virus disease in dryland sorghum

Multi-year spatial overlay patterns of plants, insects and soil water may yield insights for management for reducing biotic and abiotic stresses in dryland crops. A study of non-irrigated grain sorghum (Sorghum bicolor (L.) Moench) was conducted in a Pullman clay loam on the semi-arid High Plain of Texas during 2002–2005. The objectives of the 4-year study were to understand the mechanisms of plant spatial and temporal responses to stress from drought, infestations of greenbug, corn leaf aphid (CLA) and maize dwarf mosaic virus (MDMV) disease and soil water content (SWC) heterogeneity, and to reduce plant biotic and abiotic stress using their underlying relationships in space and time. Infrared IRt/c sensed-canopy temperature was measured at 18 or 54 sites along transects in a 6 m × 6 m grid across the years. Greenbugs, CLA, MDMV, SWC and hyperstectral reflectance were determined at each IRt/c site. Natural infestations of greenbugs and CLA on sorghum occurred in early July and insect populations peaked in late July or early August. Insect attacks resulted in plant water stress and sorghum yield loss except a late replanting in early July in 2004. Sorghum grain yield was negatively correlated with canopy temperature, greenbug and CLA (−0.38 < r < −0.75, P < 0.05), and positively correlated with SWC and plant near infrared reflectance (0.25 < r < 0.67, P < 0.05). The IRt/c temperature decreased with SWC but increased with greenbugs and CLA (0.26 < R² < 0.64). Crosscorrelation analysis showed that these insect, crop, and soil variables were correlated in space within 48–54 m. Late planting in July or spray control in late July or early August would be options to reduce dryland sorghum water stress and yield loss from drought and insect attacks.     

Effects of floor-feeding and the presence of a foraging substrate on the behaviour and stress physiological response of individually housed gilts

Both restricted feeding and barren housing have a negative influence on sow welfare. The aim of this study was to test whether sows that have to search for their feed in a substrate on the floor show less stereotyped (and other abnormal) behaviour and have a lower physiological stress response. In three batches, 96 gilts were housed individually in two rooms in 3.1 m² pens with 1.9 m² solid floor. In a 2×2 factorial design either wood shavings (S) or no substrate (NS) were provided on the floor, and 900 g of feed was provided twice daily (06:30 and 15:00 h) either in a trough (T) or on the floor (F). In weeks 8–12, behaviour was scan-sampled once in the periods 07:00–09:00 h (P1), 10:00–12:00 h (P2), and 13:00–15:00 h (P3). Data from the 5 weeks were pooled per animal. Video recordings (24 h) in week 12 or 13 were scan-sampled for ‘standing’. Saliva samples were taken in week 11 at 2 h intervals during 24 h and measured for cortisol. Spontaneously voided morning-urine was sampled in weeks 2, 7 and 12 or 13 for determination of ratios of adrenaline (A) and noradrenaline (NA) with creatinine (CR). Most effects that were found were due to substrate presence. Main findings were that compared with S-animals, NS-animals stood more during the dark period (4.8% versus 3.0%; P<0.05) and showed more (visible) oral behaviour in P1 (56.8% versus 47.6%; P<0.05), P2 (35.8% versus 30.8%; tendency) and P3 (44.1% versus 33.8%; P<0.05). This included more sham chewing in P1 (tendency), P2 and P3, more pen manipulation in P3, and more other oral behaviour (e.g. teeth grinding) in P1 (tendency) and P2. They had higher cortisol levels before feeding (peaks) and in the early evening (24 h average: 1.49 ng/ml versus 1.02 ng/ml; P<0.05) and higher NA/CR ratios in weeks 7 (tendency) and 12 (5.5 ng/mg versus 3.7 ng/mg; P<0.05). No treatment effects were found on A/CR ratios. Substantial interactive effects of feeding method and substrate were only found in floor manipulation. In all three periods NS-T-animals manipulated the floor less than other animals, probably because they had no attraction to the floor and were not rewarded for searching. Results imply that the presence of a substrate on the floor improves welfare, whereas provision of feed in the substrate in order to stimulate foraging behaviour is of less importance.     

Effect of dietary polyunsaturated/saturated fatty acid ratio on heat production and growth performance of chicks under different ambient temperature

Two trials were conducted to investigate the influence of dietary polyunsaturated/saturated fatty acid ratio (P/S) on growth performance and heat production (HP) of chicks under different ambient temperature (Ta). With the exception of Ta, all other treatment processes for two trials were the same. In each trial, 120 Arbor Acres 1-day-old male chicks were allotted to 2 (P/S, 0.6 or 2.4) × 2 (high or low Ta) factorial design with six replicates of five chicks each treatment. Chicks were reared in wire-floored metabolism cages in two temperature-controlled chambers, and were exposed to high Ta (37 and 38 °C at the first day for trial 1 and 2, respectively, and decreased 0.2 °C/day) or low Ta (33 and 28 °C at the first day for trial 1 and 2, respectively, and decreased 0.3 °C/day), for 3 weeks. Chicks were fed ad libitum. Body weight, feed intake and gain:feed were recorded. Excreta were collected for determining metabolizable energy. Energy retention was measured by the comparative slaughter technique; HP was calculated as the difference between metabolizable energy intake and body energy retention.

The results showed that high Ta decreased weight gain (P < 0.01) and feed intake (P < 0.01) in both trials, improved gain:feed in trial 2 (P < 0.01), and decreased HP in trial 2 (P < 0.01) of chicks during 0–3 weeks of age. Increasing dietary P/S did not affect the growth performance and HP of chicks during 0–3 weeks of age. No Ta × dietary P/S interaction among growth performance and HP in chicks was observed.

In summary, increasing dietary P/S did not affect HP, therefore, it is neither detrimental to the growth of chicks reared under high Ta nor is beneficial to the growth of chicks reared under low Ta.     

The presence of the potentially toxic genera Ostreopsis and Coolia (Dinophyceae) in the North Aegean Sea, Greece

The examination of macrophyte, water and sediment samples, collected at depths less than 1.5 m from 50 different sites along the North Aegean coasts, has revealed, for the first time in Greek coastal waters, the presence of two Ostreopsis species (O. ovata and O. cf. siamensis) and Coolia monotis in the majority of the sampling sites (94% and 100%, respectively). Other epiphytic dinoflagellates of the genera Prorocentrum and Amphidinium and diatoms were accompanying species in this epiphytic community. Morphometric features, plate formula and thecal ornamentation were used for species identification. O. ovata cells were smaller in dorsoventral (DV) diameter and width (W) (26.18–61.88 μm and 13.09–47.60 μm, respectively) in comparison with O. cf. siamensis (35.70–65.45 μm and 23.80–49.98 μm, respectively). In contrast, the anterioposterior (AP) diameter of O. cf. siamensis was smaller (14.28–26.18 μm) resulting in DV/AP ≈ 3, whereas the above ratio for O. ovata was less than 2 (AP ranging between 14.28–35.70 μm). Moreover, the theca of O. ovata cells was ornamented with scattered pores, which fluctuated in a wider range (0.07–0.32 μm) than those of O. cf. siamensis (0.23–0.29 μm). Coolia monotis cells were almost round with average DV diameter 26.88 μm, AP 25.66 μm and width 26.76 μm. Small and large cells were recorded in both field and culture populations of Ostreopsis spp. and C. monotis, while hyaline cysts were observed for O. ovata. The presence of O. ovata and O. cf. siamensis exhibited a clear seasonal pattern dominating (maximum abundance up to 4.05 × 10⁵ cells gr⁻¹ fwm) the period from midsummer to late autumn in years 2003 and 2004, while C. monotis was found also in winter and spring months.     

Different effects of ATP on the contractile activity of mice diaphragmatic and skeletal muscles

Apart from acetyl-choline (Ach), adenosine-5′-trisphosphate (ATP) is thought to play a role in neuromuscular function, however little information is available on its cellular physiology. As such, effects of ATP and adenosine on contractility of mice diaphragmatic and skeletal muscles (m. extensor digitorum longa—MEDL) have been investigated in in vitro experiments. Application of carbacholine (CCh) in vitro in different concentrations led to pronounced muscle contractions, varying from 9.15 ± 4.76 to 513.13 ± 15.4 mg and from 44.65 ± 5.01 to 101.46 ± 9.11 mg for diaphragm and MEDL, respectively. Two hundred micromolars of CCh in both muscles caused the contraction with the 65% (diaphragm) to 75% (MEDL) of maximal contraction force—this concentration was thus used in further experiments. It was found that application of ATP (100 μM) increased the force of diaphragmatic contraction caused by CCh (200 μM) from 335.2 ± 51.4 mg (n = 21) in controls to 426.5 ± 47.8 mg (n = 10; P < 0.05), but decreased the contractions of MEDL of CCh from 76.6 ± 6.5 mg (n = 26) in control to 40.2 ± 9.0 mg (n = 8; P < 0.05). Application of adenosine (100 μM) had no effect on CCh-induced contractions of these muscles.

Resting membrane potential (MP) measurements using sharp electrodes were done at 10, 20 and 30 min after the application of ATP and adenosine. Diaphragm showed depolarization from 75 ± 0.6 down to 63.2 ± 1.05, 57.2 ± 0.96 and 53.6 ± 1.1 mV after 10, 20 and 30 min of exposition, respectively (20 fibers from 4 muscles each, P < 0.05 in all three cases). Adenosine showed no effect on diaphragmatic MP. Both agents were ineffective in case of MEDL.

The effects of ATP in both tissues were abolished by suramin (100 μM), a P2-receptor antagonist, and chelerythrin (50 μM), a specific protein-kinase C (PKC) inhibitor, but were not affected by 1H-[1,2,4]-oxadiazolo-[4,3-]-quinoxalin-1-one (ODQ, 1 μM), a guanylyl-cyclase inhibitor, or by adenosine-3,5-monophosphothioate (Rp-cAMP, 1 μM), a protein-kinase A (PKA) inhibitor.

Besides the action on contractile activity, ATP (100 μM) led to a significant (P < 0.001) depolarization of diaphragm muscle fibers from 74.5 ± 2.3 down to 64 ± 2.1, 58.2 ± 2.2 and 54.3 ± 2.4 mV after 10, 20 and 30 min of incubation, respectively. Incubation of MEDL with the same ATP concentration showed no significant change of MP.

Denervation of the muscles for 28 days led to a decrease of CCh-induced contractions of diaphragm down to 171.1 ± 34.5 mg (n = 11, P < 0.05), but increased the contractile force of MEDL up to 723.9 ± 82.3 mg (n = 9, P < 0.01). Application of ATP elevated the contractility of denervated diaphragm caused by CCh up to normal values (311.1 ± 79.7 mg, n = 6, P > 0.05 versus control), but did not significantly affect of contractility of MEDL, which became 848.1 ± 62.7 mg (n = 6).

These results show that the effects of ATP on both diaphragmatic and skeletal muscles are mediated through P2Y receptors coupled to chelerytrin-sensitive protein-kinase C.     

Aspiration of oocytes from transitional, cycling, and pregnant mares

The aim of this study was to compare the efficacy of three approaches for recovering equine oocytes via transvaginal ultrasound-guided follicular aspiration. Fourteen mares were used as oocyte donors during the spring transition period and physiologic breeding season, and 11 mares were bred for use as oocyte donors during early gestation. In all mares, large (>20 mm) and small (10–20 mm) follicles were aspirated in eight rounds every 10–11 days. In each of the four rounds during the transition period, half the mares received 12.5 mg eFSH once daily for 4 days prior to aspiration. For each of the four rounds during the cycling season, half the mares received 12.5 mg eFSH twice daily for 3 days prior to aspiration. Pregnant mares were aspirated on days 25, 40 and 55 of gestation and received no eFSH. There were more large (>20 mm) follicles in cycling controls (2.25 ± 0.27) and cycling FSH-treated (2.64 ± 0.27) mares than in transitional FSH-treated mares (1.18 ± 0.27). The number of oocytes recovered from small (10–20 mm) follicles varied by mare (P < 0.05) and averaged 1.08 ± 0.22 per aspiration for transitional mares and 1.23 ± 0.22 per aspiration for cycling mares (P > 0.1). The number of oocytes per aspiration from large follicles was greater in cycling FSH-treated mares (0.46 ± 0.09) than in transitional control mares (0.11 ± 0.09). In pregnant mares, more large follicles were present at day 25 than at any other time, and the number of oocytes per aspiration from large follicles was greater at day 25 (0.73 ± 0.16) than at day 55 (0.04 ± 0.18). When compared across all seasons and treatments, the day 25 pregnant mares yielded the greatest number of oocytes per aspiration (2.91 ± 0.66 per mare).     

Twice single doses of 100,000 IU of vitamin D in winter is adequate and safe for prevention of vitamin D deficiency in healthy children from Ushuaia, Tierra Del Fuego, Argentina

In order to improve vitamin D status of children from Ushuaia (55°S), at the South of Argentina, double supplementation with 100.000 IU of vitamin D was administered at the beginning of winter (March 2004), and 3 months later during winter (June 2004). In 2004, serum 25-hydroxyvitamin D (25OHD) was measured before the first supplementation, a month after, and 3 months after receiving the second supplementation (March, April and September). We studied 18 healthy children from Ushuaia, age (mean ± S.D.) 7.3 ± 4.4 years old (range 1.2–14.6), seven girls and 11 boys. Before treatment, serum 25OHD was 29.3 ± 5.9 ng/ml. It increased significantly 1 month after the first supplementation (April): 35.3 ± 4.4 ng/ml (p < 0.001), and decreased significantly 3 months after the second supplementation: 22.4 ± 4.6 ng/ml (September (p < 0.001). No child was neither deficient (<10 ng/ml) nor insufficient (10–15 ng/ml) of vitamin D. On April, a month after the first supplementation, no children had vitamin D intoxication levels (>50 ng/ml). These results disclosed that to prevent vitamin D deficiency for children at zones of risk at the south of our country, double supplementation of 100,000 IU of vitamin D during autumn and winter, would be adequate and safe.     

Stability of cumin oleoresin microencapsulated in different combination of gum arabic, maltodextrin and modified starch

Microencapsulations of cumin oleoresin by spray drying using gum arabic, maltodextrin, and modified starch (HiCap^® 100) and their ternary blends as wall materials were studied for its encapsulation efficiency and stability under storage. The microcapsules were evaluated for the content and stability of volatiles, and total cuminaldehyde, γ-terpinene and p-cymene content for six weeks. Gum arabic offered greater protection than maltodextrin and modified starch, in general, although the order of protection offered was volatiles > cuminaldehyde > p-cymene > γ-terpinene. A 4/6:1/6:1/6 blend of gum arabic/maltodextrin/modified starch offered a protection, better than gum arabic as seen from the t_1/2, i.e. time required for a constituent to reduce to 50% of its initial value. However protective effect of ternary blend was not similar for the all the constituents, and followed an order of volatiles > p-cymene > cuminaldehyde > γ-terpinene.     

Synthesis and preliminary evaluation of [3H]PSB-0413, a selective antagonist radioligand for platelet P2Y12 receptors

The selective antagonist radioligand [³H]2-propylthioadenosine-5′-adenylic acid (1,1-dichloro-1-phosphonomethyl-1-phosphonyl) anhydride ([³H]PSB-0413) was prepared by catalytic hydrogenation of its propargyl precursor with a high specific radioactivity of 74 Ci/mmol. In preliminary saturation binding studies, [³H]PSB-0413 showed high affinity for platelet P2Y₁₂ receptors with a K_D value of 4.57 nM. Human platelets had a high density of P2Y₁₂ receptors exhibiting a B_max value of 7.66 pmol/mg of protein.     

Biological phosphorus removal in sequencing batch reactor with single-stage oxic process

The performance of biological phosphorus removal (BPR) in a sequencing batch reactor (SBR) with single-stage oxic process was investigated using simulated municipal wastewater. The experimental results showed that BPR could be achieved in a SBR without anaerobic phase, which was conventionally considered as a key phase for BPR. Phosphorus (P) concentration 0.22–1.79 mg L⁻¹ in effluent can be obtained after 4 h aeration when P concentration in influent was about 15–20 mg L⁻¹, the dissolved oxygen (DO) was controlled at 3 ± 0.2 mg L⁻¹ during aerobic phase and pH was maintained 7 ± 0.1, which indicated the efficiencies of P removal were achieved 90% above. Experimental results also showed that P was mainly stored in the form of intracellular storage of polyphosphate (poly-P), and about 207.235 mg phosphates have been removed by the discharge of rich-phosphorus sludge for each SBR cycle. However, the energy storage poly-β-hydroxyalkanoates (PHA) was almost kept constant at a low level (5–6 mg L⁻¹) during the process. Those results showed that phosphate could be transformed to poly-P with single-stage oxic process without PHA accumulation, and BPR could be realized in net phosphate removal.     

Modulation of ERK 1/2 and p38 MAPK signaling pathways by ATP in osteoblasts: involvement of mechanical stress-activated calcium influx, PKC and Src activation

There is evidence that extracellular nucleotides, acting through multiple P2 receptors, may play an important role in the regulation of bone metabolism by activating intracellular signaling cascades. We have studied the modulation of mitogen-activated protein kinase (MAPK) signaling pathways and its relationship to changes in intracellular calcium concentration ([Ca²⁺]_i) induced by ATP in ROS-A 17/2.8 osteoblastic cells. ATP and UTP (10 μM) increased [Ca²⁺]_i by cation release from intracellular stores. We have found that when the cells are subsequently subjected to mechanical stress (medium perturbation), a transient calcium influx occurs. This mechanical stress-activated calcium influx (MSACI) was not observed after ADP stimulation, indicating that P2Y₂ receptor activation is required for MSACI. In addition, ERK 1/2 and p38 MAPK were activated by ATP in a dose- and time-dependent manner. This activation was almost completely blocked using neomycin (2.5 mM), an inhibitor of phosphoinositide-phospholipase C (PI-PLC), Ro 318220 (1 μM), a protein kinase C (PKC) inhibitor, and PP1 (50 μM), a potent and selective inhibitor of the Src-family tyrosine kinases. Ca²⁺-free extracellular medium (containing 0.5 mM EGTA) and the use of gadolinium (5 μM), which suppressed MSACI, prevented ERK 1/2 and p38 phosphorylation by ATP. Altogether, these results represent the first evidence to date suggesting that P2Y₂ receptor stimulation by ATP in osteoblasts sensitizes mechanical stress activated calcium channels leading to calcium influx and a fast activation of the ERK 1/2 and p38 MAPK pathways. This effect also involves upstream mediators such as PI-PLC, PKC and Src family kinases.     

Galactans from cystocarpic plants of the red seaweed Callophyllis variegata (Kallymeniaceae, Gigartinales)

The polysaccharide extracted from cystocarpic Callophyllis variegata was fractionated with potassium chloride yielding three small fractions that precipitated in the ranges of 0–0.05 M KCl, 1.20–1.25 M KCl, and 1.80–2.00 M KCl, and a main product soluble in 2.00 M KCl. These fractions were analyzed, and as the first one contained very high amounts of protein, it was not studied further. Structural analysis of the rest of the fractions (F1–F3) was carried out by methylation, desulfation–methylation, IR, and ¹³C NMR spectroscopy. The results are consistent for F1 with a carrageenan-type backbone mainly constituted by β-d-galactose 2-sulfate linked to -d-galactose 2,3,6-trisulfate and β-d-galactose 2,4-disulfate linked to 3,6-anhydro-d-galactose 2-sulfate as dominant diads. In F2 these diads are present together with low amounts of β-d-galactose 2-sulfate linked to 3,6-anhydro-d-galactose 2-sulfate, whose contribution becomes higher in F3. In addition, minor but significant amounts of l-galactose were detected. F1–F3 showed potent antiviral activity against herpes simplex types 1 and 2 and dengue type 2.