An exploratory study of fibre microplastics pollution in different process stages of salt production by solar evaporation in Spain.

Microplastics are a pollutant of growing concern. Several studies have found microplastics in table salt worldwide in the last decade, although most have focused on already prepackaged salt. To the best of our knowledge, there is no previous research analysing the entire salt production process. In this study focused on solar evaporation salinas, brine and salt samples were obtained from each stage of production, starting with the entrance of seawater/brine until the final stage of ready-to-sell salt, in six sites in Spain. We extracted microplastics from each sample after 30 % H2O2 digestion and filtration through cellulose nitrate 5 µm pore filters. Microplastic fibres were optically analysed with an Olympus DSX1000. Results indicate that microplastics are present both in seawater and air, with atmospheric fallout identified as the primary source. Microplastic concentrations from the entrance to the salina till the inlet to the crystallizers ranges from 256 to 1500 items per liter and from 79 to 193 microplastics per kg for packaged salt were estimated. Artisanal salina F shows the highest content in microplastics. This study hopes to give insight into the origin and causes of microplastic pollution in solar evaporation salinas and contribute to preventing this form of pollution in food-grade salt.

Expression of enzymes involved in the urea cycle and muscle and mammary gland development of Holstein × Gyr heifers in a rotational grazing system supplemented with increasing protein levels.

Studies evaluating the crude protein (CP) supplementation strategies across the year for grazing cattle and its association with the enzymes involved in the urea cycle and muscle and mammary gland developments are scarce. Thus, we aimed to evaluate the effect of supplementation with different levels of CP on the expression of genes involved in the urea cycle and muscle and mammary gland development of Holstein × Gyr crossbreed heifers grazing intensively managed Brachiaria decumbens throughout the year. Thirty-eight heifers with average initial BW of 172.5 ± 11.15 kg (mean ± SE) and 8.2 ± 0.54 mo of age were randomly assigned to 1 of 4 treatments: 3 protein supplements (SUP) fed at 5g/kg of body weight, plus a control group (CON, non-supplemented animals). The supplement CP levels evaluated were: 12, 24, and 36%. The study was divided into 4 seasons: rainy, dry, rainy-dry transition (RDT), and dry-rainy transition (DRT). On the penultimate day of each season, ultrasound images of the carcass and mammary gland were taken. Five animals from each treatment were randomly chosen on the last day of each season, and liver and muscle tissue biopsies were performed. The target genes were the mammalian target of rapamycin (mTOR) and adenosine monophosphate-activated protein kinase (AMPK) in the muscle samples. Carbamoyl phosphate synthetase (CPS), ornithine transcarbamylase (OTC), argininosuccinate synthetase (ASS), arginosuccinate lyase (ASL), and arginase (ARG) were evaluated in the liver samples. Data were analyzed using PROC GLIMMIX of the SAS with repeated measures. We observed a greater rib eye area (cm2) and fat thickness (mm) in SUP animals than in non-supplemented animals. However, we did not observe differences among SUP levels for both variables. No effects of supplementation were detected on mammary gland development. Nevertheless, seasonal effects were observed, where the RDT and dry season had the most and least accumulated fat in the mammary gland. In muscle, we observed greater expression of AMPK in non-supplemented animals than SUP animals. On the other hand, no differences were observed in gene expression between SUP and non-supplemented animals and among SUP animals for mTOR. Season affected both AMPK and mTOR; heifers had a greater AMPK gene expression on rainy than RDT. For mTOR, we observed greater gene expression in RDT and DRT than in rainy. No differences were observed among RDT, dry, and DRT, and between dry and rainy seasons for mTOR. We observed greater CPS, ASL, and ARG gene expression in SUP animals than in non-supplemented animals. Among SUP animals, supplement CP linearly affected CPS. In conclusion, the supplementation strategy did not affect mammary gland development and mTOR expression in muscle tissue. However, we observed a seasonal effect on mammary gland development and AMPK and mTOR expression. The CP supplementation increased the rib eye area and fat thickness, directly affecting AMPK expression in the muscle. Moreover, the CP supplementation increased urea cycle enzyme expression, indicating greater urea production in the liver.

Association of housing and management practices with milk yield, milk composition, and fatty acid profile, predicted using Fourier transform mid-infrared spectroscopy, in farms with automated milking systems.

Milk fatty acid (FA) profile can be divided into (1) de novo (C4-C14) that are synthesized in the mammary gland; (2) preformed (≥C18) that are absorbed from blood and originate from mobilized adipose tissues or dietary fat; and (3) mixed (C16), which have both origins. Our objectives were to describe the FA profile, as predicted using Fourier transform mid-infrared spectroscopy, of bulk tank milk from automated milking system (AMS) farms and to assess the association of management and housing factors with the bulk tank milk composition and FA profile of those AMS farms. The data used were collected from 124 commercial Canadian Holstein dairy farms with AMS, located in the provinces of Ontario (n = 68) and Quebec (n = 56). The farms were visited once from April to September 2019, and information were collected on barn design and herd management practices. Information regarding individual cow milk yield (kg/d), days in milk, parity, and the number of milking cows were automatically collected by the AMS units on each farm. These data were extracted for the entire period that the bulk tank milk samples were monitored, from April 2019 to April 2020 in Quebec and from August 2019 to May 2020 in Ontario. Across herds, milk yield averaged (mean ± standard error) 35.9 ± 0.4 kg/d, with 3.97 ± 0.01% fat and 3.09 ± 0.01% protein, whereas FA profile averaged 26.2 ± 0.1, 33.1 ± 0.1, and 40.7 ± 0.2 g/100 g of FA for de novo, mixed, and preformed, respectively. The FA yield averaged 0.34 ± 0.01, 0.44 ± 0.01, and 0.54 ± 0.01 kg/d for de novo, mixed, and preformed, respectively. Multivariable regression models were used to associate herd-level housing factors and management practices with milk production, composition, and FA profile. Milk yield was positively associated with using a robot feed pusher (+2.1 kg/d) and the use of deep bedding (+2.6 kg/d). The use of a robot feed pusher, deep bedding, and greater stall raking frequency were positively associated with greater yield (kg/d) of de novo, mixed, preformed, and de novo + mixed FA. Use of deep bedding was negatively associated with concentration of fat, de novo FA, mixed FA, and de novo + mixed FA, expressed in grams per 100 g (%) of milk. A wider lying alley width (≥305 cm) was associated with a greater concentration (g/100 g of milk) of de novo and de novo + mixed FA. Greater frequency of partial mixed ration delivery (>2×/d vs. 1 and 2×/d) was positively associated with a greater proportion (g/100 g of FA) of de novo, mixed, and de novo + mixed FA and negatively associated with the proportion of preformed FA. Overall, these associations indicated that bulk tank FA profile can be used as a tool to monitor and adjust management and housing in AMS farms.

Desenvolvimento ponderal e parâmetros hematológicos e bioquímicos de cordeiros Pantaneiros submetidos a diferentes manejos de amamentação / [Ponderal development and hematological and biochemical parameters of Pantaneiro lambs subjected to different suckling regimens]

Este estudo teve por objetivo avaliar o desenvolvimento ponderal e a dinâmica dos parâmetros hematológicos e bioquímicos de cordeiros Pantaneiros submetidos a diferentes manejos de amamentação dos 15 aos 43 dias. Foram separados 30 cordeiros em três diferentes grupos (n=10). Os grupos foram caracterizados conforme o tempo de permanência das ovelhas com suas crias em diferentes sistemas de amamentação: MAM24 - ovelhas e cordeiros 24 horas em conjunto; MAM12 - ovelhas e cordeiros 12 horas em conjunto durante a noite; MAM2x30 - ovelhas e cordeiros 30 minutos de manhã e 30 minutos à tarde em conjunto. As coletas de amostras sanguíneas e as pesagens ocorriam a cada sete dias. Houve aumento significativo no peso com o avanço da idade dos cordeiros nos três manejos, mas os tratamentos não diferiram entre si para o ganho de peso. Houve diferenças entre os três tratamentos para a variável hematológica CHGM e para as variáveis bioquímicas AST, glicose, ureia e proteína total. As variáveis bioquímicas foram influenciadas pela faixa etária dos animais. Os diferentes manejos de amamentação não influenciaram o desenvolvimento ponderal dos cordeiros. Não ocorreram alterações patológicas. Alguns parâmetros hematológicos e bioquímicos podem ser influenciados pelo desenvolvimento etário dos cordeiros.(AU)

This study aimed to evaluate the weight development and the dynamics of hematological and biochemical parameters of Pantaneiro lambs submitted to different breastfeeding management from 15 to 43 days. First, 30 lambs were separated into three different groups (n = 10). The groups were characterized according to the length of stay of ewes with their young in different breastfeeding systems: MAM24 - ewes and lambs 24 hours together. MAM12- Sheep and lambs 12 hours together at night. MAM2x30 - sheep and lambs 30 minutes in the morning and 30 minutes in the afternoon together. Blood samples were collected and weighed every seven days. Significant increase in weight was observed with the age of the lambs in the three managements, but the treatments did not differ for weight gain. Differences were identified between the three treatments for the hematological variable CHGM and for the biochemical variables AST, glucose, urea and total protein. Correlating the age of the lambs with the biochemical variables, there was variation influenced by the age of the animals. The different management of breastfeeding did not influence the weight development of the lambs. Hematological and biochemical variations did not represent pathological changes. Some hematological and biochemical parameters may be influenced by the age development of lambs.(AU)

Energy and protein requirements of Holstein × Gyr crossbred heifers.

Nutrient requirements in cattle are dependent on physiological stage, breed and environmental conditions. In Holstein × Gyr crossbred dairy heifers, the lack of data remains a limiting factor for estimating energy and protein requirements. Thus, we aimed to estimate the energy and protein requirements of Holstein × Gyr crossbred heifers raised under tropical conditions. Twenty-two crossbred (½ Holstein × ½ Gyr) heifers with an average initial BW of 102.2 ± 3.4 kg and 3 to 4 months of age were used. To estimate requirements, the comparative slaughter technique was used: four animals were assigned to the reference group, slaughtered at the beginning of the experiment to estimate the initial empty BW (EBW) and composition of the animals that remained in the experiment. The remaining animals were randomized into three treatments based on targeted rates of BW gain: high (1.0 kg/day), low (0.5 kg/day) and close to maintenance (0.1 kg/day). At the end of the experiment, all animals were slaughtered to determine EBW, empty body gain (EBG) and body energy and protein contents. The linear regression parameters were estimated using PROC MIXED of SAS (version 9.4). Estimates of the parameters of non-linear regressions were adjusted through PROC NLIN of SAS using the Gauss-Newton method for parameter fit. The net requirements of energy for maintenance (NEm) and metabolizable energy for maintenance (MEm) were 0.303 and 0.469 MJ/EBW0.75 per day, respectively. The efficiency of use of MEm was 64.5%. The estimated equation to predict the net energy requirement for gain (NEg) was: NEg (MJ/day) = 0.299 × EBW0.75 × EBG0.601. The efficiency of use of ME for gain (kg) was 30.7%. The requirement of metabolizable protein for maintenance was 3.52 g/EBW0.75 per day. The equation to predict net protein requirement for gain (NPg) was: NPg (g/day) = 243.65 × EBW-0.091 × EBG. The efficiency of use of metabolizable protein for gain (k) was 50.8%. We observed noteworthy differences when comparing to ME and protein requirements of Holstein × Gyr crossbred heifers with other systems. In addition, we also observed differences in estimates for NEm, NEg, NPg, kg and k. Therefore, we propose that the equations generated in the present study should be used to estimate energy and protein requirements for Holstein × Gyr crossbred dairy heifers raised in tropical conditions in the post-weaning phase up to 185 kg of BW.

Determination of macromineral requirements for preweaned dairy calves in tropical conditions.

International committees that have published nutrient requirements for dairy cattle have used data from mineral studies conducted in the 1920s to 1970s, and no study has reported data from animals less than 100 kg; therefore, there is a need to update mineral requirements for preweaned dairy calves. Thus, a meta-analysis was performed to estimate the mineral requirements of Ca, P, K, Mg, and Na for Holstein and Holstein × Gyr crossbred preweaned dairy calves using data from 5 studies developed at the Universidade Federal de Viçosa (Viçosa, MG, Brazil). A total of 210 calves were separated into 2 breeds: purebred Holstein calves (animals with a Holstein pedigree higher than 87.5%) and Holstein × Gyr crossbred calves (animals with a Holstein pedigree lower than 87.5%). The comparative slaughter technique was used to estimate animal body composition and empty body weight (EBW). Mineral requirements for maintenance were estimated by the regression between retained mineral and mineral intake, whereas mineral requirements for gain were obtained from the first derivative of the mineral content in the animal's body. In addition, breed effect was tested on the intercept and slope of the models. The effect of breed was not observed for all analyzed variables. Thus, net requirements for maintenance were 12.73, 11.81, 20.28, 3.50, and 6.37 mg/kg of EBW per day for Ca, P, K, Mg, and Na, respectively. Retention coefficients were 73.18, 65.20, 13.16, 29.55, and 24.28% for Ca, P, K, Mg, and Na, respectively. The following equations were determined to estimate net requirements for gain (NRG, g/d): NRG for Ca = 14.402 × EBW-0.139 × empty body gain (EBG); NRG for P = 5.849 × EBW-0.027 × EBG; NRG for K = 1.140 × EBW-0.048 × EBG; NRG for Mg = 0.603 × EBW-0.036 × EBG; and NRG for Na = 1.508 × EBW-0.045 × EBG. Due to the high variation between the data found in this study and in the available literature, we suggest that further studies should be conducted to evaluate the estimates of this study.

An overview of patterns and dynamics of suspended sediment transport in an agroforest headwater system in humid climate: Results from a long-term monitoring.

Small headwater catchments deliver large quantities of suspended sediment (SS) to the ocean. However, there are relatively few studies focused on the study of patterns and dynamics of suspended sediment in headwater catchments over the long-term (10 year or more). In this study, the dynamics of suspended sediment transport were examined at different time scales in a small headwater catchment in NW Spain, based on a 12-year dataset from high-resolution monitoring. The results revealed that, similar to other humid catchments, the hydrological response was highly dependent on initial conditions, especially in autumn and summer. However, in winter and spring the hydrology was more influenced by rainfall amount. The annual suspended sediment was 117 Mg, which equates to a suspended sediment yield of 10 Mg km-2 y-1. The SS yield in the Corbeira catchment is related to runoff generation and flooding, which play a key role in sediment yield from the catchment. About 80% of the annual SS was transported over 12% of the study period. Rainfall and discharge at the beginning of the events were the most important factors in explaining the hydrological response at event scale. Suspended sediment transport in this catchment is determined by event magnitude, while the SS is mainly influenced by variables related to runoff erosivity.

Aluminum fractionation in acidic soils and river sediments in the Upper Mero basin (Galicia, NW Spain).

This study aims to determine aluminum fractions in the fine earth of acidic soils under different land uses (forest, pasture and cultivation) and in the river bed sediments of the headwater of the Mero River in order to identify and quantify Al-bearing phases to assess Al mobility and potential bioavailability (environmental availability) in the monitoring area. Sequential extraction is used to evaluate the Al partitioning into six fractions operationally defined: soluble/exchangeable/specifically adsorbed, bound to manganese oxides, associated with amorphous compounds, aluminum bound to oxidizable organic matter, associated with crystalline iron oxides, and residual fraction (aluminum within the crystal lattices of minerals). The mean concentration of total aluminum (24.01 g kg-1) was similar for the three considered uses. The mean percentage of the aluminum fractions, both in soils and sediments, showed the following order: residual fraction â‰« amorphous compounds ≈ crystalline iron oxides > water-soluble/exchangeable/specifically adsorbed > bound to oxidizable organic matter ≈ Mn oxides. However, in the soils, the amorphous compounds and water-soluble/exchangeable/specifically adsorbed fraction showed considerable differences between some types of uses, the percentage of aluminum linked to amorphous compounds being higher in forest soils (16% of total Al) compared to other uses (mean about 8% of total Al). The highest values of water-soluble/exchangeable/specifically adsorbed Al were also found in forest soils (mean 8.6% of the total Al versus about 4% of pasture and cultivation), which is consistent with the lower pH and higher organic matter content in forest soils. Nevertheless, the potentially bioavailable fraction (sum of the first three fractions) is low, suggesting very low geoavailability of this element in both soils and sediments; hence, the possibility to affect the crops and water quality is minimal.

An optimized protocol for DNA extraction in plants with a high content of secondary metabolites, based on leaves of Mimosa tenuiflora (Willd.) Poir. (Leguminosae).

Some species are characterized by a high content of tannins, alkaloids, and phenols in their leaves. These secondary metabolites are released during DNA extraction and might hinder molecular studies based on PCR (polymerase chain reaction). To provide an efficient method to extract DNA, Mimosa tenuiflora, an important leguminous plant from Brazilian semiarid region used in popular medicine and as a source of fuelwood or forage, was used. Eight procedures previously reported for plants were tested and adapted from leaf tissues of M. tenuiflora stored at -20°C. The optimized procedure in this study encompassed the utilization of phenol during deproteinization, increased concentrations of cetyltrimethylammonium bromide and sodium chloride, and a shorter period and lower temperature of incubation concerning other methods. The extracted DNA did not present degradation, and amplification via PCR was successful using ISSR, trnL, ITS, and ETS primers. Besides M. tenuiflora, this procedure was also tested and proved to be efficient in genetic studies of other plant species.

Energy and protein requirements of crossbred (Holstein × Gyr) growing bulls.

The objective of this study was to estimate the energy and protein requirements of crossbred (Holstein × Gyr) growing bulls. Twenty-four 10-mo-old bulls [initial body weight (BW) = 184 ± 23.4 kg] were used in a comparative slaughter trial. Six bulls were slaughtered at the beginning of the experiment as the reference group, to estimate initial empty body weight (EBW) and energy and protein contents of the remaining animals. The remaining bulls were assigned to a completely randomized design with 3 levels of dry matter intake and 6 replicates. The levels of dry matter intake were 1.2% of BW, 1.8% of BW, and ad libitum to target orts equal to 5% of the total amount that was fed. The remaining bulls were slaughtered at the end of the experiment. The bulls were fed a diet consisting of 59.6% corn silage and 40.4% concentrate on a dry matter basis. The equation that determined the relationship between EBW and BW was EBW = (0.861 ± 0.0031) × BW. The relationship between empty body gain (EBG) and average daily gain (ADG) was demonstrated by the following equation: EBG = (0.934 ± 0.0111) × ADG. Net energy for maintenance (NEM) was 74.8 ± 2.89 kcal/kg of EBW0.75 per day, and metabolizable energy for maintenance (MEM) was 120.8 kcal/kg of EBW0.75 per day. The detected efficiency of use of metabolizable energy for maintenance (km) was 61.9%. The equation used to estimate net energy for gain (NEG) was as follows: NEG = (0.049 ± 0.0011) × EBW0.75 × EBG0.729 ± 0.0532. The efficiency of use of metabolizable energy for gain (kg) was 35.7%. The metabolizable protein for maintenance (MPM) was 3.05 g/kg of BW0.75. The equation used to estimate net protein requirements for gain (NPG) = (87.138 ± 65.1378 × EBG) + [(40.436 ± 21.3640) × NEG]. The efficiency of use of metabolizable protein for gain (k) was 35.7%. We concluded that the estimates of energy and protein requirements presented herein are more appropriate than the National Research Council dairy cattle model and the Brazilian BR-CORTE system to balance the diets of crossbred (Holstein × Gyr) growing bulls.

Determination of energy and protein requirements for crossbred Holstein × Gyr preweaned dairy calves.

The objective was to quantify the energy and protein nutritional requirements of Holstein × Gyr crossbred preweaned dairy calves until 64 d of age. Thirty-nine Holstein × Gyr crossbred male calves with an average initial live weight (mean ± SEM; for all next values) of 36 ± 1.0 kg were used. Five calves were slaughtered at 4 d of life to estimate the animals' initial body composition (reference group). The remaining 34 calves were distributed in a completely randomized design in a 3 × 2 factorial arrangement consisting of 3 levels of milk (2, 4, or 8 L/d) and 2 levels of starter feed (presence or absence in diet). At 15 and 45 d of life, 4 animals from each treatment were subjected to digestibility trials with total collection of feces (for 72 h) and urine (for 24 h). At 64 d of age, all animals were slaughtered, their gastro-intestinal tract was washed to determine the empty body weight (EBW; kg), and their body tissues were sampled for subsequent analyses. The net energy requirement for maintenance was estimated using an exponential regression between metabolizable energy intake and heat production (both in Mcal/EBW0.75 per d) and was 74.3 ± 5.7 kcal/EBW0.75 per d, and was not affected by inclusion of starter feed in the diet. The metabolizable energy requirement for maintenance was determined at the point of zero energy retention in the body and was 105.2 ± 5.8 kcal/EBW0.75 per d. The net energy for gain was estimated using the EBW and the empty body gain (EBG; kg/d) as 0.0882 ± 0.0028 × EBW0.75 × EBG0.9050±0.0706. The metabolizable energy efficiency for gain (kg) of the milk was 57.4 ± 3.45%, and the kg of the starter feed was 39.3 ± 2.09%. The metabolizable protein requirement for maintenance was 3.52 ± 0.34 g/BW0.75 per d. The net protein required for each kilogram gained was estimated as 119.1 ± 32.9 × EBW0.0663±0.059. The metabolizable protein efficiency for gain was 77 ± 8.5% and was not affected by inclusion of starter feed in the diet. In conclusion, the energy efficiency for gain of milk is higher than that of starter and the net protein required per unit protein gain increases with empty body weight.

Performance and health of Holstein calves fed different levels of milk fortified with symbiotic complex containing pre- and probiotics.

The objective of this study was to evaluate the performance and health of Holstein calves fed low or high milk supply (MSP) with or without symbiotic complex (SYM) supplementation, consisting of prebiotics, probiotics, and fibrolytic enzymes. Thirty-two Holstein calves with body weight (BW) of 34 ± 7 kg were distributed in a randomized block design in a 2 × 2 factorial arrangement. Treatments consisted of low and high MSP: 10 % of BW from 1st to 8th weeks after birth (low) and 20 % BW from 1st and 2nd weeks after birth, 15 % BW for the 3rd and 4th weeks after birth, and 10 % BW from 5th and 8th weeks after birth (high). Solid ration was supplied in addition to milk. Intake, ADG, diet digestibility, and fecal consistency index were evaluated. Low and high MSP groups tended (P < 0.10) to differ in calf growth, final BW (69 vs. 73 kg), post-weaning average weight gain (548 vs. 788 g/day), and final average weight gain (549 vs. 646 g/day) in low and high MSP calves, respectively. There was an interaction between MSP level and SYM on the digestibilities of dry matter (DM) and neutral detergent fiber (NDF) (P < 0.10). In the low MSP group, inclusion of SYM increased digestibility of DM (0.720 to 0.736 g/kg) and NDF (0.758 to 0.783 g/kg). The inclusion of SYM improved calf health (P < 0.10) with a fecal score of 0.31 compared to 0.42 without SYM. Milk-feeding level was an important factor in calf performance, while SYM supplementation improved diet digestibility and animal health.

Ants in the Hospital Environment: Ecological Parameters as Support for Future Management Strategies.

Urban ants cause many losses to human society, and they represent a potential threat to public health in hospital environments due to their ability to transport pathogenic organisms. We evaluated several ecological parameters (richness, abundance, constancy, and evenness), their fluctuation during the seasons, and identified species that occur outside the natural range of the ant fauna of a hospital environment, as support for future management strategies. Ant sampling was held every 2 months by using attractive bait traps in the morning and evening, leading to the sampling of 10,342 individuals belonging to six subfamilies and 26 species. Myrmicinae showed higher richness (n = 12) and abundance (n = 7336), with Pheidole susannae Forel being the most abundant species. The most constant species (100%) were P. susannae and Tetramorium simillimum (Smith). Among the most abundant species, Monomorium floricola (Jerdon) and Tapinoma melanocephalum (Fabricius) are considered as species that occur outside the natural range. No difference was observed between species richness and abundance. The Shannon (2.247), dominance (0.1395) and evenness indices (0.6897) indicated a stability of the community throughout the year with high diversity and low dominance of species. The sampled data constitute a new series of information on a long-term ecological approach to support future management strategies in hospital environments and allow for more efficient pest control.

Quantification of miltefosine in peripheral blood mononuclear cells by high-performance liquid chromatography-tandem mass spectrometry.

Phagocytes, the physiological compartment in which Leishmania parasites reside, are the main site of action of the drug miltefosine, but the intracellular pharmacokinetics of miltefosine remain unexplored. We developed a bioanalytical method to quantify miltefosine in human peripheral blood mononuclear cells (PBMCs), expanding from an existing high performance liquid chromatography-tandem mass spectrometry method for the quantification of miltefosine in plasma. The method introduced deuterated miltefosine as an internal standard. Miltefosine was extracted from PBMC pellets by addition of 62.5% methanol. Supernatant was collected, evaporated and reconstituted in plasma. Chromatographic separation was performed on a reversed phase C18 column and detection with a triple-quadrupole mass spectrometer. Miltefosine was quantified using plasma calibration standards ranging from 4 to 1000ng/mL. This method was validated with respect to its PBMC matrix effect, selectivity, recovery and stability. No matrix effect could be observed from the PBMC content (ranging from 0.17 to 26.3×10(6)PBMCs) reconstituted in plasma, as quality control samples were within 3.0% of the nominal concentration (precision less than 7.7%). At the lower limit of quantitation of 4 ng/mL plasma, corresponding to 0.12ng/10(6) PBMCs in a typical clinical sample, measured concentrations were within 8.6% of the nominal value. Recovery showed to be reproducible as adding additional pre-treatment steps did not increase the recovery with more than 9%. This method was successfully applied to measure intracellular miltefosine concentrations in PBMC samples from six cutaneous leishmaniasis patients up to one month post-treatment.

Therapeutic properties of VO(dmpp)2 as assessed by in vitro and in vivo studies in type 2 diabetic GK rats.

The bis(1,2-dimethyl-3-hydroxy-4-pyridinonato)oxidovanadium(IV), VO(dmpp)2, has shown anti-diabetic effects by in vitro studies in Wistar (W) rat adipocytes and in vivo in obese Zucker rats. The aim of this work is to confirm the therapeutic properties of VO(dmpp)2 in non-obese type 2 diabetic Goto-Kakizaki (GK) rats. An in vivo study was carried out, treating W and GK rats during 21 days with a daily dose of VO(dmpp)2 (44 µmol/kg). It was shown that VO(dmpp)2 doesn't affect the normal increase of body weight of both W and GK rats, after 8 days of treatment ameliorates glycemia in GK rats (8.4 ± 0.3 vs 10.1 ± 0.2 mM in GK control, P<0.001) but doesn't interfere with glucose levels in W rats and, after 21 days of treatment, improves the glucose intolerant profile of GK rats (13.1 ± 0.5 vs 20.6 ± 0.7 mM/min in GK control, P<0.001), despite no increase of plasma insulin levels during glucose tolerance test. Additionally, it was demonstrated that VO(dmpp)2 significantly enhances [3-(3)H]-glucose uptake by W and GK rat adipocytes (non-toxic concentration of 100 µM: respectively 193 ± 20 and 254 ± 21%, P<0.001, relative to the basal value) showing an efficacy similar to insulin 1.72 nM and better than the same concentration of BMOV (P<0.01). Western blotting revealed that in W and GK rats VO(dmpp)2 significantly promotes IRS2 (P<0.05) and p-AKT expression (P<0.001 and P<0.05, respectively, relative to the respective controls) and in GK animals reduces the increase of PTP1ß expression (P<0.001, relative to GK control).

Phosphorus loss from a mixed land use catchment in northwest Spain.

This work examined P loss from a mixed land use catchment in northwest Spain and how it was related to hydrological factors. A stream draining a mixed land use catchment of 16 km in northwest Spain was monitored for 5 yr. Water samples were taken more frequently with increased stream flows. A total of 132 runoff events of different magnitudes were analyzed in this study. Mean annual P loss from the catchment was 11.5 kg km, about 68% being transported during runoff events, which only contributed to 29% of the flow. The contribution of runoff events to particulate P and dissolved P losses was 76 and 46% of total losses during the study period, respectively. A high interannual variability in P loss was observed, the differences being related to rainfall amount and distribution. At a seasonal scale, the highest P loads and concentrations were observed during rainy seasons; the lowest P loads occurred in summer because they occurred with stream flow. Particulate P accounted for the greatest proportion (74%) of transported P. There was a high variability in P transported between runoff events, with maximum discharge and discharge increase being the most important variables explaining total P concentration in these events through their effect on suspended sediment concentration.

Inhibitory effects of caspase inhibitors on the activity of matrix metalloproteinase-2.

Matrix metalloproteinase (MMP)-2, a zinc-dependent endopeptidase, plays a detrimental role in several diseases including ischemia and reperfusion (I/R) injury of the heart. Caspases are a group of cysteine-dependent, aspartate-directed proteases which regulate cellular apoptosis. Interestingly, protective effects of caspase inhibitors independent of apoptosis have been shown in I/R injury of the heart. The cardioprotective actions of both these classes of protease inhibitors led us to hypothesize that caspase inhibitors may also reduce MMP-2 activity. Five known caspase inhibitors (Z-IE(OMe)TD(OMe)-fmk, Ac-DEVD-CHO, Ac-LEHD-cmk, Z-VAD-fmk and Ac-YVAD-cmk) were tested for their possible inhibitory effects on MMP-2 activity in comparison to the MMP inhibitors ONO-4817 and ARP-100 (which themselves were unable to inhibit caspase-3 activity). MMP-2 activity was assessed by an in vitro troponin I (TnI) proteolysis assay and a quantitative kinetic fluorescence assay using a fluorogenic peptide substrate (OmniMMP). TnI proteolysis was also measured by western blot in neonatal cardiomyocytes subjected to hypoxia-reoxygenation injury. Using human recombinant MMP-2 and TnI as its substrate, the caspase inhibitors, in comparison with ONO-4817, significantly inhibited MMP-2-mediated TnI degradation in a concentration-dependent manner. The kinetic assay using OmniMMP revealed that these caspase inhibitors blocked MMP-2 activity in a concentration-dependent manner with similar IC50 values. TnI degradation in neonatal cardiomyocytes was enhanced following hypoxia-reoxygenation and this was blocked by ARP-100 and Ac-LEHD-cmk. Inhibition of MMP-2 activity is an additional pharmacological action which contributes to the protective effects of some caspase inhibitors.

Prevalence of human papillomavirus (HPV) type 16 variants and rare HPV types in the central Amazon region.

Infection by human papillomavirus (HPV) is one of the primary causes of mortality by cancer in northern Brazil. Sexually active women from Manaus, Amazonas, without cytological alterations and women with pre-malignant and malignant cytological alterations were examined for HPV virus, identified via PCR and sequencing. The target region for this study was part of the L1 capsid gene of HPV. Twenty-three samples that were PCR-positive were sequenced. Analysis of 336 bp demonstrated a high incidence of high-risk HPV types in the population of Manaus, identified as HPVs 16, 33, 58, 66, 68. HPV type 16 was the most prevalent, presenting two variants similar to the Asian-American (AA) and East-Asian type (As) variants. A rare HPV type 13 related to "Heck's disease" was also detected. This preliminary provides important information about the HPV circulating in Amazonas State.

[Systematic review of the implementation and evaluation of Pharmaceutical Care in hospitalised patients(Pharmaceutical Care implementation in hospitalised patients. Systematic review)]. / Revision sistemática sobre la implantación y la evaluación del seguimiento farmacoterapéutico en pacientes hospitalizados.

INTRODUCTION: The persistent morbidity and mortality related to pharmaceutical treatment for hospitalised patients mean that it is necessary to identify scientific criteria for implementing and evaluating Pharmaceutical Care (phC) on the hospital setting. OBJECTIVE: The purpose of the study is to perform a systematic literature review in order to locate, select and analyse studies on implementing and evaluating phC in hospitalised patients. MATERIAL AND METHODS: We searched for articles having to do with clinical pharmacy (CP) and phC published between 1990 and 2006, using a restricted search technique combining all descriptors. The databases we searched were Medline, Embase-Drug & Pharmacology and Cochrane Library. We selected original articles and reviews in English or Spanish describing a phC and clinical pharmacy programme having a participating pharmacist and used in hospitalised patients. RESULTS: We located 66 publications, of which 49 (74.2%) were included and 17 (25.8%) were excluded. We selected 15 (22.7%) on integrating CP and phC in the hospital environment, 18 (27.3%) on implementing phC and 16 (24.2%) relating to evaluating phC programmes. CONCLUSIONS: In the listed studies, pharmacists have managed to incorporate phC programmes in pharmacy divisions' treatment activities. Joining efforts in order to unify CP and phC criteria should be a plan for a common future in this profession. Patients under care should obtain concrete health benefits from phC use, and hospitals should recognise that they create beneficial effects at a reasonable cost.