Effect of carbohydrate type and concentration on polyhydroxy alcohol and trehalose content of conidia of three entomopathogenic fungi

The entomopathogenic fungi Beauveria bassiana, Metarhizium anisopliae and Paecilomyces farinosus were cultured on solid agar media containing different carbohydrate components (glycerol, glucose, trehalose or starch) at concentrations of ≤ 142.7 g added carbon 1-1 for 30 d at 25°C. The water activity (a(w)) of the media ranged from 0.925 to 0.998. Growth of M. anisopliae and P. farinosus was stimulated between 0.975 and 0.995 a(w) on glucose media and that of P. farinosus at 0. 975 a(w) on glycerol media. At < 0.970 a(w), growth of each fungal species was significantly reduced (P < 0.05). Polyhydroxy alcohols (polyols) and trehalose were extracted from conidia produced on different media and quantified using HPLC. Total polyol content of conidia produced on glucose media varied between 5.2 and 52.2 mg g-1 for B. bassiana, 77.3 and 90.3 mg g-1 for M. anisopliae, and 26.7 and 76.1 mg g-1 for P. farinosus. The amounts of specific polyols in conidia varied significantly from media of different glucose concentrations. Mannitol was the predominant polyol in conidia of all three species, with conidia of M. anisopliae, for example, containing as much as 75.2 mg mannitol g-1 when cultured on glucose media. The amount of the lower molecular mass polyols glycerol and erythritol was greater in conidia produced on glucose media with > 50.0 g added carbon 1-1 than that in conidia produced at lower glucose concentrations. Conidia contained between 10.8 and 20.8 mg glycerol plus erythritol g-1 on glucose media with 142.7 g added carbon 1-1, depending on species. Conversely, conidia of B. bassiana and P. farinosus contained maximum amounts of trehalose ( ≤ 23.5 mg g-1) when produced on glucose media with < 50.0 g added carbon l-1, and trehalose content was considerably less at higher glucose concentrations. There were accumulations of glycerol and erythritol in conidia of all three species when grown on glycerol media with > 25.0 g added carbon 1-1; conidia of B. bassiana contained up to 154.0 mg glycerol plus erythritol g-1. hen B. bassiana and P. farinosus were grown on trehalose media, conidia contained up to 222.1 mg trehalose g-1. By contrast, conidia of M. anisopliae contained < 17.0 mg trehalose g-1 under all conditions tested. The water availability of solutions of different polyols is discussed in relation to their potential to act in osmotic adjustment during germination. The ability to manipulate polyol and trehalose content of fungal propagules may be critical in enhancing the storage life and efficacy of biological control agents.

Distribution of assimilated carbon within the plant and rhizosphere of Lolium perenne:Influence of temperature

Perennial rye-grass plants were pulse labelled with [¹⁴C]-CO₂ over a range of temperatures (5-25°C). The fate of the label was determined within the plant and soil. The temperature at which plants were pulse labelled had a marked effect on the distribution of the label within the plant and soil system. Root-soil respiration increased from 5.7 to 24.15% when expressed as a percentage of net assimilated label. The percentage of label remaining in the plant root and in the soil was greater at 5 and 25°C, with a minimum for both these components at 15°C. At 15°C the percentage of net assimilated label that remained in the shoots was greater than at other temperatures, with this percentage decreasing at the lower and higher temperatures. © 1989.

A comparison of carbon flow from pre-labelled and pulse-labelled plants

Perennial rye-grass was subjected to two different¹⁴C labelling regimes to enable a partitioning of the carbon sources contributing to rhizosphere carbon-flow. Plant/soil microcosms were designed which enabled rye-grass plants to either receive a single pulse of¹⁴C-CO₂ or to be pre-labelled using a series of¹⁴C-CO₂ pulses, allowing the fate of newly photoassimilated carbon and carbon lost by root decomposition to be followed into the soil. For young rye-grass plants grown over a short period, rhizosphere carbon flow was found to be dominated by newly photoassimilated carbon. Evidence for this came from the observed percentage of the total¹⁴C budget (i.e. total¹⁴C-CO₂ fixed by the plants) lost from the root/soil system, which was 30 times greater for the pulse labelled compared to pre-labelled plants. Root decomposition was found to be less at 10°C compared to 20-25°C, though input of¹⁴C into the soil was the same at both temperatures. © 1988 Kluwer Academic Publishers.

Tributyltin (TBT) resistance in Aeromonas molluscorum Av27

O tributilestanho (TBT) é considerado um dos xenobióticos mais tóxicos, produzidos e deliberadamente introduzidos no meio ambiente pelo Homem. Tem sido usado numa variedade de processos industriais e subsequentemente descarregado no meio ambiente. O tempo de meia-vida do TBT em águas marinhas é de várias semanas, mas em condições de anóxia nos sedimentos, pode ser de vários anos, devido à sua degradação mais lenta. Embora o TBT tenha sido descrito como sendo tóxico para eucariotas e procariotas, muitas bactérias podem ser resistentes a este composto. O presente trabalho teve como objetivo principal elucidar o mecanismo de resistência ao TBT em bactérias. Para além disso, pretendeu-se desenvolver um biorepórter para detectar TBT no ambiente. Para atingir estes objetivos foram delineadas várias tarefas cujos principais resultados obtidos se apresentam a seguir. Várias bactérias resistentes ao TBT foram isoladas de sedimento e água do Porto de Pesca Longínqua (PPL) na Ria de Aveiro, Portugal. Entre estas, Aeromonas molluscorum Av27 foi selecionada devido à sua elevada resistência a este composto (concentrações até 3 mM), à sua capacidade de degradar o TBT em compostos menos tóxicos (dibutilestanho, DBT e monobutilestanho, MBT) e também por usar o TBT como fonte de carbono. A. molluscorum Av27 foi caracterizada genotipica e fenotipicamente. Os fatores de virulência estudados mostraram que esta estirpe i) possui atividade lipolítica; ii) não é citotóxica para células de mamíferos, nomeadamente para células Vero; iii) não possui integrões de classe I e II e iv) possui cinco plasmídeos com aproximadamente 4 kb, 7 kb, 10 kb, 100 kb e mais de 100 kb. Estes resultados mostraram que a estirpe Av27 não é tóxica, aumentando assim o interesse nesta bactéria para futuras aplicações, nomeadamente na bioremediação. Os testes de toxicidade ao TBT mostraram que este composto tem um impacto negativo no crescimento desta estirpe, bem como, na densidade, no tamanho e na atividade metabólica das células e é responsável pela formação de agregados celulares. Assim, o TBT mostrou ser bastante tóxico para as bactérias interferindo com a atividade celular geral. O gene Av27-sugE, que codifica a proteína SugE pertencente à família das “small multidrug resistance proteins” (SMR), foi identificado como estando envolvido na resistência ao TBT nesta estirpe. Este gene mostrou ser sobreexpresso quando as células crescem na presença de TBT. O promotor do gene Av27-sugE foi utilizado para construir um bioreporter para detetar TBT, contendo o gene da luciferase do pirilampo como gene repórter. O biorepórter obtido reúne as características mais importantes de um bom biorepórter: sensibilidade (intervalo de limite de detecção de 1-1000 nM), rapidez (3 h são suficientes para a deteção de sinal) e, possivelmente, não é invasivo (pois foi construído numa bactéria ambiental). Usando sedimento recolhido no Porto de Pesca Longínqua da Ria de Aveiro, foi preparada uma experiência de microcosmos com o intuito de avaliar a capacidade de Av27 para bioremediar o TBT, isoladamente ou em associação com a comunidade bacteriana indígena. A análise das amostras de microcosmos por PCR-DGGE e de bibliotecas de 16S rDNA revelaram que a comunidade bacteriana é relativamente estável ao longo do tempo, mesmo quando Av27 é inoculada no sedimento. Para além disso, o sedimento estuarino demonstrou ser dominado por bactérias pertencentes ao filo Proteobacteria (sendo mais abundante as Delta e Gammaproteobacteria) e Bacteroidetes. Ainda, cerca de 13% dos clones bacterianos não revelaram nenhuma semelhança com qualquer dos filos já definidos e quase 100% afiliou com bactérias não cultiváveis do sedimento. No momento da conclusão desta tese, os resultados da análise química de compostos organoestânicos não estavam disponíveis, e por essa razão não foi possível tirar quaisquer conclusões sobre a capacidade desta bactéria remediar o TBT em sedimentos. Esses resultados irão ajudar a esclarecer o papel de A. molluscorum Av27 na remediação de TBT. Recentemente, a capacidade da estirpe Av27 remediar solo contaminado com TBT foi confirmada em bioensaios realizados com plantas, Brassica rapa e Triticum aestivum (Silva 2011a), e também com invertebrados Porcellionides pruinosus (Silva 2011B). Assim, poder-se-á esperar que a bioremediação do sedimento na experiência de microcosmos também tenha ocorrido. No entanto, só a análise química dos compostos organostânicos deverá ser conclusiva. Devido à dificuldade em realizar a análise analítica de organoestânicos, um método de bioensaio fácil, rápido e barato foi adaptado para avaliar a toxicidade do TBT em laboratório, antes de se proceder à análise química das amostras. O método provou a sua utilidade, embora tenha mostrado pouca sensibilidade quando se usam concentrações de TBT baixas. Em geral, os resultados obtidos contribuíram para um melhor entendimento do mecanismo de resistência ao TBT em bactérias e mostraram o potencial biotecnológico de A. molluscorum Av27, nomeadamente, no que refere à sua possível aplicação na descontaminação de TBT no ambiente e também no desenvolvimento de biorepórteres.

Ecological interactions in populations facing environmental stress

O zooplâncton, particularmente os cladóceros, são organismos de água doce importantes na regulação da produção primária dos ecossistemas de água doce. No entanto, também podem adaptar-se a condições salobras. Tendo em conta as previsões no âmbito das alterações climáticas, a intrusão salina pode ocorrer a par com a subida de temperatura. As populações de água doce podem ficar vulneráveis aos efeitos interativos da salinidade e da temperatura, de acordo com os seus limites de tolerância e capacidade de adaptação ao stress ambiental. Assim, a presente tese analisou as interações resultantes das alterações destes agentes de stress em populações de cladóceros de água doce. Primeiro, comparou-se a halotolerância de diferentes genótipos de Simocephalus vetulus provenientes de populações de água doce e de água salobra de modo a avaliar a existência de uma componente genética de resistência à salinidade. A sensibilidade aguda dos genótipos variou na mesma gama de concentrações; todavia, todos os genótipos da população salobra, exceto um, foram mais tolerantes do que os de água doce, em termos de tempo à imobilização. Contudo, não foi possível estabelecer uma relação entre a performance reprodutiva em condições salobras e o contexto ambiental de origem destes genótipos. Mais, estes ensaios mostraram que as populações de água doce têm potencial para tolerar incrementos de salinidade. Como tal, pode-se concluir que a seleção a que os genótipos estão sujeitos no seu local de origem foi mais fraca do que o esperado. Segundo, investigou-se a capacidade de aclimatação de Daphnia galeata à salinidade e temperatura, de modo a avaliar a halotolerância de Daphnia a duas temperaturas num cenário de aclimatação multigeracional. O objetivo foi compreender se a pré-adaptação ao stress ambiental (20ºC e 25ºC versus 0 g/L e 1 g/L de NaCl) influenciou posteriormente as respostas a estes agentes de stress. Verificou-se uma tendência para um aumento de sensibilidade ao NaCl, a temperaturas mais elevadas. No entanto, este efeito foi anulado após nove gerações, mas apenas quando os organismos foram aclimatados aos dois agentes de stress em simultâneo (salinidade e temperatura elevada). Terceiro, demonstrou-se experimentalmente que a salinidade interferiu com a competição interespecífica, alterando a composição das comunidades zooplanctónicas. Este conjunto de evidências permitiu-nos refletir nos múltiplos impactos de agentes de stress, particularmente os relacionados com as previsões de alterações climáticas. Em paralelo aos estudos de natureza experimental, e numa perspetiva de Educação para o Desenvolvimento Sustentável (EDS), importa também promover o desenvolvimento de competências necessárias à compreensão de mudanças ambientais globais (e.g., o impacto da salinidade e da temperatura) para implementar estratégias de mitigação e adaptação. Neste contexto, foi realizada uma atividade com estudantes do ensino secundário, que se tornou uma boa oportunidade para a sua aprendizagem e aquisição de competências de interpretação de dados experimentais, assim como de sensibilização para as questões ambientais.

The effects of chemicals in isopods : a multi-organizational evaluation

The global aim of this thesis was to evaluate and assess the effects of a pesticide (dimethoate) and a metal (nickel), as model chemicals, within different organization levels, starting at the detoxification pathways (enzymatic biomarkers) and energy costs associated (energy content quantification, energy consumption and CEA) along with the physiological alterations at the individual and population level (mortality), leading to a metabolomic analysis (using liquid 1H-NMR) and finally a gene expression analysis (transcriptome and RT-qPCR analysis). To better understand potential variations in response to stressors, abiotic factors were also assessed in terrestrial isopods such as temperature, soil moisture and UV radiation. The evaluation performed using biochemical biomarkers and energy related parameters showed that increases in temperature might negatively affect the organisms by generating oxidative stress. It also showed that this species is acclimated to environments with low soil moisture, and that in high moisture scenarios there was a short gap between the optimal and adverse conditions that led to increased mortality. As for UV-R, doses nowadays present have shown to induce significant negative impact on these organisms. The long-term exposure to dimethoate showed that besides the neurotoxicity resulting from acetylcholinesterase inhibition, this stressor also caused oxidative stress. This effect was observed for both concentrations used (recommended field dose application and a below EC50 value) and that its combination with different temperatures (20ºC and 25ºC) showed different response patterns. It was also observed that dimethoate’s degradation rate in soils was higher in the presence of isopods. In a similar study performed with nickel, oxidative stress was also observed. But, in the case of this stressor exposure, organisms showed a strategy where the energetic costs necessary for detoxification (biomarkers) seemed to be compensated by positive alterations in the energy related parameters. In this work we presented for the first time a metabolomic profile of terrestrial isopods exposed to stressors (dimethoate and niquel), since until the moment only a previous study was performed on a metabolomic evaluation in nonexposed isopods. In the first part of the study we identify 24 new metabolites that had not been described previously. On the second part of the study a metabolomic profile variation of abstract non-exposed organism throughout the exposure was presented and finally the metabolomic profile of organisms exposed to dimethoate and nickel. The exposure to nickel suggested alteration in growth, moult, haemocyanin and glutathione synthesis, energy pathways and in osmoregulation. As for the exposure to dimethoate alterations in osmoregulation, energy pathways, moult and neurotransmission were also suggested. In this work it was also presented the first full body transcriptome of a terrestrial isopod from the species Porcellionides pruinosus, which will complement the scarce information available for this group of organisms. This transcriptome also served as base for a RNA-Seq and a RT-qPCR analysis. The results of the RNA-Seq analysis performed in organisms exposed to nickel showed that this stressor negatively impacted at the genetic and epigenetic levels, in the trafficking, storage and elimination of metals, generates oxidative stress, inducing neurotoxicity and also affecting reproduction. These results were confirmed through RT-qPCR. As for the impact of dimethoate on these organisms it was only accessed through RT-qPCR and showed oxidative stress, an impact in neurotransmission, in epigenetic markers, DNA repair and cell cycle impairment. This study allowed the design of an Adverse Outcome Pathway draft that can be used further on for legislative purposes.

Ecofisiologia de Gelidium sesquipedale: efeitos da temperatura e intensidade luminosa na fotossíntese

Dissertação de mestrado, Aquacultura, Unidade de Ciências e Tecnologias dos Recursos Aquáticos, Universidade do Algarve, 1998

Factors Involved in the Phenological Mechanism of Alnus Flowering in Central Europe

The objectives of this paper are to ascertain the main factors involved in the phenological mechanism of alder flowering in Central Europe by understanding the in - fluence of the main meteorological parameters, the North Atlantic Oscillation (NAO) effect and the study of the Chill and Heat requirements to overcome dormancy. Airborne pollen (1995–2007) was collected in Poznań (Poland) by means a volumetric spore trap. Temperatures for February, and January and February averages of the NAO are generally key factors affecting the timing of the alder pollen seasons. Chilling accumulation (which started in Poznań at the beginning of November, while the end took place during the month of January) of 985 CH with a threshold temperature of -0.25ºC, followed by 118 GDDºC with a threshold temperature of 0.5ºC, were necessary to overcome dormancy and produce the onset of flowering. The calculated dormancy requirements, mean tem - peratures of the four decades of the year, and January and February average NAO index recorded during the period before flowering, were used to construct linear and multiple regression models in order to forecast the start date of the alder pollen seasons Its ac - curacy was tested using data from 2007, and the difference between the predicted and observed dates ranged from 3–7 days

Effect of temperature and prey in the biology of Scymnus subvillosus (Goeze) (Coleoptera: Coccinellidae)

Dissertação de Mestrado, Biotecnologia em Controlo Biológico, 18 de Dezembro de 2013, Universidade dos Açores.

The influence of skeletal muscle cell volume on carbohydrate metabolism in contracting skeletal muscle

This study investigated the regulation of carbohydrate metabolism through changes in skeletal muscle cell volume immediately post contraction and during recovery. Using an established in vitro isolated muscle strip model, soleus (SOL) and extensor digitorum longus (EDL) were dissected from male rats and incubated in an organ bath (perfused with 95% O2; 5% CO2, pH 7.4, temperature 25°C) containing medium- 199 altered to a target osmotic condition (iso-, hypo- or hyper-osmotic; 290, 1 80, 400 mmol/kg). Muscles were stimulated for 10 minutes (40 Hz SOL; 30 Hz EDL) and then either immediately flash frozen or allowed to recover for 20 minutes before subsequent metabolite and enzyme analysis. Results demonstrated a relative water decrease in HYPER vs. HYPOosmotic condition (n=8/group; p<0.05) regardless of muscle type. Specifically, the SOL HYPER condition had elevated metabolite concentrations after 10 minutes of stimulation in comparison to both HYPO and ISO (p<0.05), while EDL muscle did not show any significant difTerences between the HYPER or HYPO conditions. After 20 minutes of recovery, metabolic changes occurred in both SOL and EDL with the SOL HYPER condition showing greater relative changes in metabolite concentrations versus HYPO. The results of the current study have demonstrated that osmotic imbalance induces metabolic change within the skeletal muscle cell and muscle type may influence the mechanisms utilized for cell volume regulation.

The effects of temperature on nickel tocicity in goldfish (carassius auratus L.)

Although it is widely assumed that temperature affects pollutant toxicity, few studies have actually investigated this relationship. Moreover, such research as has been done has involved constant temperatures; circumstances which are rarely, if ever, actually experienced by north temperate, littoral zone cyprinid species. To investigate the effects of temperature regime on nickel toxicity in goldfish (Carassius auratus L.), 96- and 240-h LCSO values for the heavy metal pollutant, nickel (NiCI2.6H20), were initially determined at 2DoC (22.8 mg/L and 14.7 mg/L in artificially softened water). Constant temperature bioassays at 10°C, 20°C and 30°C were conducted at each of 0, 240-h and 96-h LCSO nickel concentrations for 240 hours. In order to determine the effects of temperature variation during nickel exposure it was imperative that the effects of a single temperature change be investigated before addressing more complex regimes. Single temperature changes of + 10°C or -10°C were imposed at rates of 2°C/h following exposures of between 24 hand 216 h. The effects of a single temperature change on mortality, and duration of toxicant exposure at high and low temperatures were evaluated. The effects of fluctuating temperatures during exposure were investigated through two regimes. The first set of bioassays imposed a sinewave diurnal cycle temperature (20.±.1DOC) throughout the 10 day exposure to 240-h LeSO Ni. The second set of investigations approximated cyprinid movement through the littoral zone by imposing directionally random temperature changes (±2°C at 2-h intervals), between extremes of 10° and 30°C, at 240-h LC50 Ni. Body size (i.e., total length, fork length, and weight) and exposure time were recorded for all fish mortalities. Cumulative mortality curves under constant temperature regimes indicated significantly higher mortality as temperature and nickel concentration were increased. At 1DOC no significant differences in mortality curves were evident in relation to low and high nickel test concentrations (Le., 16 mg/L and 20 mg/L). However at 20°C and 30°C significantly higher mortality was experienced in animals exposed to 20 mg/L Ni. Mortality at constant 10°C was significantly lower than at 30°C with 16 mg/L and was significantly loWer than each of 2DoC and 39°C tanks at 20 mg/L Ni exposure. A single temperature shift from 20°C to 1DoC resulted in a significant decrease in mortality rate and conversely, a single temperature shift from 20°C to 30°C resulted in a significant increase in mortality rate. Rates of mortality recorded during these single temperature shift assays were significantly different from mortality rates obtained under constant temperature assay conditions. Increased Ni exposure duration at higher temperatures resulted in highest mortality. Diurnally cycling temperature bioassays produced cumulative mortality curves approximating constant 20°C curves, with increased mortality evident after peaks in the temperature cycle. Randomly fluctuating temperature regime mortality curves also resembled constant 20°C tanks with mortalities after high temperature exposures (25°C - 30°C). Some test animals survived in all assays with the exception of the 30°C assays, with highest survival associated with low temperature and low Ni concentration. Post-exposure mortality occurred most frequently in individuals which had experienced high Ni concentrations and high temperatures during assays. Additional temperature stress imposed 2 - 12 weeks post exposure resulted in a single death out of 116 individuals suggesting that survivors are capable of surviving subsequent temperature stresses. These investigations suggest that temperature significantly and markedly affects acute nickel toxicity under both constant and fluctuating temperature regimes and plays a role in post exposure mortality and subsequent stress response.

Palynology and foraminiferal geochemistry of the Lower Pleistocene Olduvai Subchron (ca. 1.8 Ma) in DSDP Hole 603C, western North Atlantic

Marine palynology and benthic and planktonic foraminiferal geochemistry are combined to reveal long- and short-term (Milankovitch-scale) paleoceanographic changes across the upper half of the Olduvai Subchron (ca. 1.86--1.77 Ma, lower Pleistocene) in DSDP Hole 603C from the lower New Jersey continental rise. Planktonic foraminiferal Mg/Ca ratios reveal annual sea-surface temperatures between 14.5° and 25°C, whereas modern values vary between 16° and 20°e. Despite evidence of downslope transport in much of the studied interval, dinoflagellate cyst and acritarch assemblages appear to reflect fluctuating temperate to subtropical water masses. These assemblages comprise both neritic and oceanic species, and are marked by a transition upsection from warm conditions, dominated by Lingulodinium machaerophorum, Polysphaeridium zoharyi and Cymatiosphaera? invaginata, to cooler conditions dominated by Filisphaera filifera. Combining dinoflagellate cyst proxies with planktonic foraminiferal geochemistry allows downslope transport events to be recognized during glacial episodes, and events dominated by intensified bottom-water circulation during interglacial episodes. Sixtytwo in-situ dinoflagellate cyst and acritarch taxa were recorded including several not previously described.

Phosphorylation of Skeletal Muscle Pyruvate Dehydrogenase Phosphatase in Response to Insulin Stimulation

Pyruvate dehydrogenase phosphatase (PDP) regulates carbohydrate oxidation through the pyruvate dehydrogenase (PDH) complex. PDP activates PDH, enabling increased carbohydrate flux towards oxidative energy production. In culture myoblasts, both PDP1 and PDP2 undergo covalent activation in response to insulin–stimulation by protein kinase C delta (PKCδ). Our objective was to examine the effect of insulin on PDP phosphorylation and PDH activation in skeletal muscle. Intact rat extensor digitorum longus muscles were incubated (oxygenated at 25°C, 1g of tension) for 30min in basal or insulin–stimulated (10 mU/mL) media. PDH activity increased 58% following stimulation, (p=0.057, n=11). Serine phosphorylation of PDP1 (p=0.047) and PDP2 (p=0.006) increased by 29% and 48%, respectively (n=8), and mitochondrial PKCδ protein content was enriched by 45% in response to stimulation (p=0.0009, n=8). These data suggest that the insulin–stimulated increase in PDH activity in whole tissue is mediated through mitochondrial migration of PKCδ and subsequent PDP phosphorylation.

Myosin Regulatory Light Chain Phosphorylation and Its Effect on the Contractile Economy of Mouse Fast Muscle

Activated by elevations in myoplasmic calcium concentration, myosin light chain kinase (skMLCK) phosphorylates the regulatory light chains (RLCs) of fast muscle myosin. This covalent modification potentiates force production, but requires an investment of ATP. Our objective was to investigate the effect of RLC phosphorylation on the contractile economy (mechanical output:metabolic input) of fast twitch skeletal muscle. Extensor digitorum longus muscles isolated from Wildtype and skMLCK-/- mice mounted in vitro (25°C) were subjected to repetitive low-frequency stimulation (10Hz,15s) known to cause activation of skMLCK, and staircase potentiation of force. With a 3-fold increase in RLC phosphate content, Wildtype generated 44% more force than skMLCK-/- muscles over the stimulation period (P = .002), without an accompanied increase in energy cost (P = .449). Overall, the contractile economy of Wildtype muscles, with an intact RLC phosphorylation mechanism, was 73% greater than skMLCK /- muscles (P = .043), demonstrating an important physiological function of skMLCK during repetitive contractile activity.

Stability studies of intravenous cyclosporine preparations stored in non-PVC containers

Dans cette étude, la stabilité de préparations intraveineuses de cyclosporine (0.2 et 2.5 mg/mL dans NaCl 0.9% ou dextrose 5%) entreposées dans des seringues de polypropylène, des sacs de polypropylène-polyoléfine et des sacs de vinyle acétate d’éthylène a été évaluée. Une méthode HPLC indicatrice de la stabilité à base de méthanol a été développée et validée suite a des études de dégradation forcée. Les solutions évaluées ont été préparées de façon aseptique, puis entreposées à 25°C. La stabilité chimique a été évaluée par HPLC et la stabilité physique a été évaluée par inspection visuelle et aussi par diffusion dynamique de la lumière (DLS). Tous les échantillons sont demeurés stables chimiquement et physiquement dans des sacs de polypropylène-polyoléfine (>98% de cyclosporine récupérée après 14 jours). Lorsqu’entreposés dans des seringues de polypropylène, des contaminants ont été extraits des composantes de la seringue. Toutefois, aucune contamination n’a été observée après 10 min de contact entre la préparation de cyclosporine non-diluée et ces mêmes seringues. Les préparations de 2.5 mg/mL entreposées dans des sacs de vinyle acétate d’éthylène sont demeurés stables chimiquement et physiquement (>98% de cyclosporine récupérée après 14 jours). Toutefois, une adsorption significative a été observée avec les échantillons 0.2 mg/mL entreposés dans des sacs de vinyle acétate d’éthylène (<90% de cyclosporine récupéré après 14 jours). Une étude cinétique a démontré une bonne corrélation linéaire entre la quantité adsorbée et la racine carrée du temps de contact (r2 > 0.97). Un nouveou modèle de diffusion a été établi. En conclusion, les sacs de polypropylène-polyoléfine sont le meilleur choix; les seringues de polypropylène présentent un risque de contamination, mais sont acceptables pour un transfert rapide. Les sacs de vinyle acétate d’éthylène ne peuvent être recommandés à cause d’un problème d’adsorption.