Ultrasonication processing for the production of plant-based nanoemulsions

Plant-derived proteins have been emerging and growing in interest over the past few years, due to their interesting properties and the trend to replace animal-derived proteins [1]. Ultrasonication processing can be used to develop nanoemulsions based on plant proteins that are kinetically stabilized by their small dimension, unlike classic emulsions [2]. In this work, oil-in-water nanoemulsions were produced through high-speed homogenization, followed by ultrasonic homogenization (US), using different plant-derived proteins, including potato (Solanum tuberosum), lupin (Lupinus angustifolius), pea (Pisum sativum), chickpea (Cicer arietinum) and faba bean (Vicia faba) protein as emulsifiers. A central composite rotatable experimental design was used to evaluate the influence of three independent variables: water/oil ratio (65-75% of water), protein content (1-6%) and US time (1-7 min) on the size average (by intensity) and polydispersity index (PDI) of the nanoemulsions. A total of 17 experiments were performed with 14 three-level experimental points, and 3 replicates at the central point. The effect of the US time (0, 3, 4.5 and 6 min) in the potato and lupin proteins primary and secondary structures were analysed through SDS-PAGE electrophoresis and circular dichroism, respectively. Results showed that the use of potato, lupin and pea proteins lead to the formation of stable nanoemulsions, while chickpea and faba bean proteins resulted in non-stable nanoemulsions, with phase separation. The smallest mean droplet size for potato protein was 439.9 nm and PDI value 0.464 [21:73 (w/w) oil/water ratio, 6% of protein and 6 min of US]. The smallest mean droplet size for lupin protein was 505.5 nm and PDI value 0.434, and for pea protein the droplet size was 551.3 nm and PDI value 0.249 [23.6:73 (w/w) oil/water ratio, 3.4% of protein and 6 min of US]. Electrophoresis results show that for native potato and lupin samples the ultrasonication did not induce significant changes in the protein pattern, indicating that the US treatment did not modify the primary structure. Regarding the second structure, US did not change the secondary structure of potato protein but induced a slight increase of -helix for all US treatments for lupin protein. Stable nanoemulsions can be developed using plant-derived proteins and ultrasonication, foreseeing different applications in the food industry.This study was supported by the project cLabel+ (POCI-01-0247-FEDER-046080) cofinanced by Compete 2020, Lisbon 2020, Portugal 2020 and the European Union, through the European Regional Development Fund (ERDF).info:eu-repo/semantics/publishedVersio

Carboxylic ester hydrolases from hyperthermophiles

Carboxylic ester hydrolyzing enzymes constitute a large group of enzymes that are able to catalyze the hydrolysis, synthesis or transesterification of an ester bond. They can be found in all three domains of life, including the group of hyperthermophilic bacteria and archaea. Esterases from the latter group often exhibit a high intrinsic stability, which makes them of interest them for various biotechnological applications. In this review, we aim to give an overview of all characterized carboxylic ester hydrolases from hyperthermophilic microorganisms and provide details on their substrate specificity, kinetics, optimal catalytic conditions, and stability. Approaches for the discovery of new carboxylic ester hydrolases are described. Special attention is given to the currently characterized hyperthermophilic enzymes with respect to their biochemical properties, 3D structure, and classification

Changes in End-of-Life Practices in European Intensive Care Units From 1999 to 2016

Importance: End-of-life decisions occur daily in intensive care units (ICUs) around the world, and these practices could change over time. Objective: To determine the changes in end-of-life practices in European ICUs after 16 years. Design, Setting, and Participants: Ethicus-2 was a prospective observational study of 22 European ICUs previously included in the Ethicus-1 study (1999-2000). During a self-selected continuous 6-month period at each ICU, consecutive patients who died or had any limitation of life-sustaining therapy from September 2015 until October 2016 were included. Patients were followed up until death or until 2 months after the first treatment limitation decision. Exposures: Comparison between the 1999-2000 cohort vs 2015-2016 cohort. Main Outcomes and Measures: End-of-life outcomes were classified into 5 mutually exclusive categories (withholding of life-prolonging therapy, withdrawing of life-prolonging therapy, active shortening of the dying process, failed cardiopulmonary resuscitation [CPR], brain death). The primary outcome was whether patients received any treatment limitations (withholding or withdrawing of life-prolonging therapy or shortening of the dying process). Outcomes were determined by senior intensivists. Results: Of 13 625 patients admitted to participating ICUs during the 2015-2016 study period, 1785 (13.1%) died or had limitations of life-prolonging therapies and were included in the study. Compared with the patients included in the 1999-2000 cohort (n = 2807), the patients in 2015-2016 cohort were significantly older (median age, 70 years [interquartile range {IQR}, 59-79] vs 67 years [IQR, 54-75]; P < .001) and the proportion of female patients was similar (39.6% vs 38.7%; P = .58). Significantly more treatment limitations occurred in the 2015-2016 cohort compared with the 1999-2000 cohort (1601 [89.7%] vs 1918 [68.3%]; difference, 21.4% [95% CI, 19.2% to 23.6%]; P < .001), with more withholding of life-prolonging therapy (892 [50.0%] vs 1143 [40.7%]; difference, 9.3% [95% CI, 6.4% to 12.3%]; P < .001), more withdrawing of life-prolonging therapy (692 [38.8%] vs 695 [24.8%]; difference, 14.0% [95% CI, 11.2% to 16.8%]; P < .001), less failed CPR (110 [6.2%] vs 628 [22.4%]; difference, -16.2% [95% CI, -18.1% to -14.3%]; P < .001), less brain death (74 [4.1%] vs 261 [9.3%]; difference, -5.2% [95% CI, -6.6% to -3.8%]; P < .001) and less active shortening of the dying process (17 [1.0%] vs 80 [2.9%]; difference, -1.9% [95% CI, -2.7% to -1.1%]; P < .001). Conclusions and Relevance: Among patients who had treatment limitations or died in 22 European ICUs in 2015-2016, compared with data reported from the same ICUs in 1999-2000, limitations in life-prolonging therapies occurred significantly more frequently and death without limitations in life-prolonging therapies occurred significantly less frequently. These findings suggest a shift in end-of-life practices in European ICUs, but the study is limited in that it excluded patients who survived ICU hospitalization without treatment limitations.status: publishe

Changes in End-of-Life Practices in European Intensive Care Units From 1999 to 2016

Key PointsQuestionHave end-of-life practices in European intensive care units (ICUs) changed from 1999-2000 to 2015-2016? FindingsIn this prospective observational study of 1785 patients who had limitations in life-prolonging therapies or died in 22 European ICUs in 2015-2016, compared with data previously reported from the same ICUs in 1999-2000 (2807 patients), treatment limitations (withholding or withdrawing life-sustaining treatment or active shortening of the dying process) occurred significantly more frequently (89.7\% vs 68.3\%), whereas death without any limitations in life-prolonging therapies occurred significantly less frequently (10.3\% vs 31.7\%). MeaningThese findings suggest that end-of-life care practices in European ICUs changed from 1999-2000 to 2015-2016 with more limitations in life-prolonging therapies and fewer deaths without treatment limitations. ImportanceEnd-of-life decisions occur daily in intensive care units (ICUs) around the world, and these practices could change over time. ObjectiveTo determine the changes in end-of-life practices in European ICUs after 16 years. Design, Setting, and ParticipantsEthicus-2 was a prospective observational study of 22 European ICUs previously included in the Ethicus-1 study (1999-2000). During a self-selected continuous 6-month period at each ICU, consecutive patients who died or had any limitation of life-sustaining therapy from September 2015 until October 2016 were included. Patients were followed up until death or until 2 months after the first treatment limitation decision. ExposuresComparison between the 1999-2000 cohort vs 2015-2016 cohort. Main Outcomes and MeasuresEnd-of-life outcomes were classified into 5 mutually exclusive categories (withholding of life-prolonging therapy, withdrawing of life-prolonging therapy, active shortening of the dying process, failed cardiopulmonary resuscitation {[}CPR], brain death). The primary outcome was whether patients received any treatment limitations (withholding or withdrawing of life-prolonging therapy or shortening of the dying process). Outcomes were determined by senior intensivists. ResultsOf 13625 patients admitted to participating ICUs during the 2015-2016 study period, 1785 (13.1\%) died or had limitations of life-prolonging therapies and were included in the study. Compared with the patients included in the 1999-2000 cohort (n=2807), the patients in 2015-2016 cohort were significantly older (median age, 70 years {[}interquartile range \{IQR\}, 59-79] vs 67 years {[}IQR, 54-75]; P&lt;.001) and the proportion of female patients was similar (39.6\% vs 38.7\%; P=.58). Significantly more treatment limitations occurred in the 2015-2016 cohort compared with the 1999-2000 cohort (1601 {[}89.7\%] vs 1918 {[}68.3\%]; difference, 21.4\% {[}95\% CI, 19.2\% to 23.6\%]; P&lt;.001), with more withholding of life-prolonging therapy (892 {[}50.0\%] vs 1143 {[}40.7\%]; difference, 9.3\% {[}95\% CI, 6.4\% to 12.3\%]; P&lt;.001), more withdrawing of life-prolonging therapy (692 {[}38.8\%] vs 695 {[}24.8\%]; difference, 14.0\% {[}95\% CI, 11.2\% to 16.8\%]; P&lt;.001), less failed CPR (110 {[}6.2\%] vs 628 {[}22.4\%]; difference, -16.2\% {[}95\% CI, -18.1\% to -14.3\%]; P&lt;.001), less brain death (74 {[}4.1\%] vs 261 {[}9.3\%]; difference, -5.2\% {[}95\% CI, -6.6\% to -3.8\%]; P&lt;.001) and less active shortening of the dying process (17 {[}1.0\%] vs 80 {[}2.9\%]; difference, -1.9\% {[}95\% CI, -2.7\% to -1.1\%]; P&lt;.001). Conclusions and RelevanceAmong patients who had treatment limitations or died in 22 European ICUs in 2015-2016, compared with data reported from the same ICUs in 1999-2000, limitations in life-prolonging therapies occurred significantly more frequently and death without limitations in life-prolonging therapies occurred significantly less frequently. These findings suggest a shift in end-of-life practices in European ICUs, but the study is limited in that it excluded patients who survived ICU hospitalization without treatment limitations. This study compares changes in end-of-life practices (withholding or withdrawing of life-prolonging therapy, active shortening of the dying process, failed CPR, documentation of brain death) in 22 European ICUs between 1999-2000 and 2015-2016

Temperature-induced molecular transport through polymer multilayers coated with PNIPAM microgels

Polyelectrolyte multilayers serve as effective reservoirs for bioactive molecules which are stored and released from the multilayers for cellular applications. However, control over the release without significantly affecting the multilayers and biomolecules is still a challenge. On the other hand, externally stimulated release would make the multilayers promising for the development of stimuli-sensitive planar carriers with release performance switched on demand. In this study soft composite films are designed by coating hyaluronic acid/poly-l-lysine (HA/PLL) multilayers with temperature responsive poly(N-isopropylacrylamide) (PNIPAM) microgels. Microgels are flattened and immersed into the multilayers to maximize the number of contacts with the surrounding polyelectrolytes (HA and PLL). The microgel coating serves as an efficient switchable barrier for the PLL transport into the multilayers. PLL diffusion into the film is significantly hindered at room temperature but is dramatically enhanced at 40 °C above the volume phase transition temperature (VPTT) of PNIPAM at 32 °C associated with microgel shrinkage. Scanning force microscopy micrographs show that the mechanism of volume phase transition on soft surfaces cannot be directly deduced from the processes taking place at solid substrates

Temperature-induced molecular transport through polymer multilayers coated with PNIPAM microgels

Polyelectrolyte multilayers serve as effective reservoirs for bioactive molecules which are stored and released from the multilayers for cellular applications. However, control over the release without significantly affecting the multilayers and biomolecules is still a challenge. On the other hand, externally stimulated release would make the multilayers promising for the development of stimuli-sensitive planar carriers with release performance switched on demand. In this study soft composite films are designed by coating hyaluronic acid/poly-l-lysine (HA/PLL) multilayers with temperature responsive poly(N-isopropylacrylamide) (PNIPAM) microgels. Microgels are flattened and immersed into the multilayers to maximize the number of contacts with the surrounding polyelectrolytes (HA and PLL). The microgel coating serves as an efficient switchable barrier for the PLL transport into the multilayers. PLL diffusion into the film is significantly hindered at room temperature but is dramatically enhanced at 40 °C above the volume phase transition temperature (VPTT) of PNIPAM at 32 °C associated with microgel shrinkage. Scanning force microscopy micrographs show that the mechanism of volume phase transition on soft surfaces cannot be directly deduced from the processes taking place at solid substrates