Rapid one-step separation and purification of recombinant phenylalanine dehydrogenase in aqueous two-phase systems

Background: Phenylalanine dehydrogenase (PheDH; EC 1.4.1.20) is a NAD+-dependent enzyme that performs the reversible oxidative deamination of L-phenylalanine to phenylpyruvate. It plays an important role in detection and screening of phenylketonuria (PKU) diseases and production of chiral intermediates as well. The main goal of this study was to find a simple and rapid alternative method for purifying PheDH. Methods: The purification of recombinant Bacillus sphaericus PheDH was investigated in polyethylene glycol (PEG) and ammonium sulfate aqueous two-phase systems (ATPS). The influences of system parameters including PEG molecular weight and concentration, pH and (NH4)2SO4 concentration on enzyme partitioning were also studied. The purity of enzyme was analyzed by sodium dodecyl sulfate polyacrylamide gel electrophoresis. Results: A single extraction process was developed for separation and purification of recombinant PheDH from E. coli BL21 (DE3). The optimized conditions for partitioning and purification of PheDH were 9% (w/w) PEG-6,000 and 16% (w/w) (NH4)2SO4 at pH 8.0. The partition coefficient, recovery, yield, purification factor and specific activity values were achieved 58.7, 135%, 94.42%, 491.93 and 9828.88 U/mg, respectively. Also, the Km values for L-phenylalanine and NAD+ in oxidative deamination were 0.21 and 0.13 mM, respectively. Conclusion: The data presented in this paper demonstrated the potential of ATPS as a versatile and scaleable process for downstream processing of recombinant PheDH.     

Microalgae as Contributors to Produce Biopolymers

Biopolymers are very favorable materials produced by living organisms, with interesting properties such as biodegradability, renewability, and biocompatibility. Biopolymers have been recently considered to compete with fossil-based polymeric materials, which rase several environmental concerns. Biobased plastics are receiving growing interest for many applications including electronics, medical devices, food packaging, and energy. Biopolymers can be produced from biological sources such as plants, animals, agricultural wastes, and microbes. Studies suggest that microalgae and cyanobacteria are two of the promising sources of polyhydroxyalkanoates (PHAs), cellulose, carbohydrates (particularly starch), and proteins, as the major components of microalgae (and of certain cyanobacteria) for producing bioplastics. This review aims to summarize the potential of microalgal PHAs, polysaccharides, and proteins for bioplastic production. The findings of this review give insight into current knowledge and future direction in microalgal-based bioplastic production considering a circular economy approach. The current review is divided into three main topics, namely (i) the analysis of the main types and properties of bioplastic monomers, blends, and composites; (ii) the cultivation process to optimize the microalgae growth and accumulation of important biobased compounds to produce bioplastics; and (iii) a critical analysis of the future perspectives on the field.     

Anti-inflammatory activities of cucurbitacin E isolated from Citrullus lanatus var. citroides: role of reactive nitrogen species and cyclooxygenase enzyme inhibition

The in vivo and in vitro mechanistic anti-inflammatory actions of cucurbitacin E (CE) (Citrullus lanatus var. citroides) were examined. The results showed that LPS/INF-γ increased NO production in RAW264.7 macrophages, whereas L-NAME and CE curtailed it. CE did not reveal any cytotoxicity on RAW264.7 and WRL-68 cells. CE inhibited both COX enzymes with more selectivity toward COX-2. Intraperitoneal injection of CE significantly suppressed carrageenan-induced rat's paw edema. ORAC and FRAP assays showed that CE is not a potent ROS scavenger. It could be concluded that CE is potentially useful in treating inflammation through the inhibition of COX and RNS but not ROS.     

Modulating gut microbiota and digestive enzyme activities of Artemia urmiana by administration of different levels of Bacillus subtilis and Bacillus licheniformis

The aim of this study was to evaluate the effects of Bacillus subtilis and Bacillus licheniformis on growth, gut microbiota, and digestive enzyme activities of Artemia urmiana. Three diets containing 10² (T₁), 10⁴ (T₂), 10⁶ (T₃) CFU of probiotics/g feed, and a control diet (C) without probiotic were used through a completely randomized design (treatments with triplicates). Twelve plastic tanks with the capacity of 60- l and density of 20 nauplii/ml were used and the trial continued for 15?days. Results showed that probiotics significantly increased the total length of A. urmiana (P?<?0.05). Although the total aerobic gastrointestinal bacteria count showed no significant differences among the treatments, the total Bacillus count significantly increased in experiments (P?<?0.05). The ratio of TCBS to total aerobic bacteria count was significantly lower in T₁ (0.31?±?0.05), T2 (0.27?±?0.15), and T₃ (0.25?±?0.05) compared to the control (0.76?±?0.34) (P?<?0.05). The probiotics were able to increase the protease and amylase activities (P?<?0.05). No significant effect on lipase activity. The study determined T₂ and T₃ as the most effective treatments for improving growth, bacterial flora, and digestive enzyme activities. As less probiotic needed in T₂, using 10⁴ bacteria per g diet is recommended for rearing Artemia up to the maturity stage.     

Extrapituitary gonadotropin-releasing hormone (GnRH) binding sites in goldfish

In teleosts, as in other vertebrates, the secretion of pituitary gonadotropin (GTH) is mediated by the hypothalamic decapeptide, gonadotropin-releasing hormone (GnRH). Recent findings in teleosts indicate that GnRH receptors are not restricted to the pituitary gonadotropes and are also associated with somatotropes as well as being present in a number of other tissues. In the present study, we provide novel information on GnRH binding in a number of extrapituitary tissues in goldfish. However, we do not intend to provide full characterization of GnRH binding sites in various extrapituitary tissues in goldfish as this would clearly be outside the scope of this paper. In this study we examined GnRH binding in a number of extrapituitary tissues in goldfish and observed specific binding in ovary, testis, brain, liver and kidney. No specific GnRH binding was observed in muscle, skin, gut, gill and heart. In general, the present findings together with the results of other studies carried out in our laboratory demonstrate that mature goldfish ovary and testis contain two classes of GnRH binding sites, high affinity/low capacity and low affinity/high capacity sites with binding characteristics similar to those of the pituitary GnRH receptors. The brain of goldfish was also found to contain two classes of GnRH binding sites, a super-high affinity/low capacity and a low affinity/high capacity sites. Furthermore, study of goldfish liver and kidney demonstrated the presence of a single class of GnRH binding sites with characteristics different from those of pituitary, ovary, testis and brain. Overall, it is evident that goldfish contains a family of GnRH binding sites which can be classified into four groups based on binding affinities: 1) A class of high affinity binding sites present in the pituitary, ovary and testis, 2) a class of super high affinity sites so far only detected in the brain, 3) a class of intermediate-affinity GnRH binding sites in the liver and kidney, and 4) a class of low affinity binding sites present in all the tissues containing specific GnRH binding sites except for liver and kidney.     

Which source and level of dietary sodium is appropriate for broiler chickens reared in a high‐altitude area?

This study investigated effect of increasing level of dietary sodium using sodium bicarbonate or sodium chloride on growth performance, mortality, characteristics of carcass, organs and tibia, calcium and phosphorus of serum in broilers reared in a high‐altitude area (1,700 m above sea level). A total of 588 Ross 308 male broiler chicks were used in seven treatments, six replicates per treatment of 14 birds per each from 1 to 38 d of age. Seven dietary treatments consisted of a basal diet (with 0.16% sodium and 0.23% chloride), top‐dressed for six diets to give three supplementary levels of sodium (0.07%, 0.14% and 0.21%) from sodium bicarbonate (respectively by 0.26%, 0.52% and 0.78%) or sodium chloride (respectively by 0.18%, 0.36% and 0.54%), resulting in seven diets with total sodium and chloride levels of 0.16% and 0.23%, 0.23% and 0.23%, 0.30% and 0.23%, 0.37% and 0.23%, 0.23% and 0.33%, 0.30% and 0.44%, 0.37% and 0.55% respectively. Increasing sodium level improved feed conversion ratio (FCR) linearly and quadratically. However, when FCR was calculated without adjusting for feed intake of mortalities, the enhanced sodium level did not improve this parameter. Increasing sodium level via sodium chloride enhanced ascites mortality, total mortality, relative weight of heart and right ventricle linearly. Increasing sodium level reduced serum calcium and enhanced serum phosphorus linearly; however, there was a linear tendency to increase tibia ash when sodium level was enhanced by sodium bicarbonate (p = 0.08) or sodium chloride (p = 0.07). Increasing sodium level via sodium bicarbonate tended (p = 0.08) to reduce tibia strength linearly. In conclusion, a diet with 0.16% sodium and 0.23% chloride is enough for broiler chicken reared in a high‐altitude area, and increasing dietary sodium level via sodium chloride has detrimental effect on survivability of broiler in such condition.     

Western blot analysis of Leishmania infantum antigens using sera from pentamidine-treated dogs

Pentamidine treatment has been used successfully to establish immune cellular responses in recovered dogs. In this paper, we examined the appearance and disappearance of antibodies over time against crude Leishmania antigens and purified gp63 or gp70 proteins in sera from cured dogs using a Western blotting technique.Following the treatment, a pattern of antibody specificities to parasite antigens was observed in the sera of cured dogs. Antibodies to gp63 and gp70 were maintained after cure. In addition, the reaction with a 26 kD band observed during the clinical phase was no longer recognized by sera taken from recovery dogs. Interestingly, two proteins, 85 and 110 kD, not observed during the patent phase were detected by sera taken from treated dogs. Such patterns of antibody specificities to various parasite antigens might represent a useful parameter to determine the actual phase of the disease process.     

近60年来克里雅绿洲耕地动态变化驱动力及生态环境效应

利用1950-2008年统计资料和土地详查数据,采用多元统计分析与文献调查,分析了塔克拉玛干沙漠南缘克里雅绿洲近60年来耕地动态变化情况及其驱动机理,并讨论了耕地动态变化对绿洲生态环境的影响。结果表明:(1) 1950-2008年克里雅绿洲耕地面积经历了急剧增加(1950-1961年)→缓慢减少(1962-1990年)→快速增加(1991-2008年)的变化过程,耕地数量总体呈增加趋势,人均耕地面积呈减少趋势;(2)影响克里雅绿洲耕地动态变化的驱动因子可归纳为政策变动、经济发展、人口增长、农业科技进步等四大因素;(3)耕地动态变化某种程度上改善了克里雅绿洲生态环境的同时,导致了绿洲外围沙漠化,绿洲内部土壤盐渍化,湿地减少,河道缩短等一系列生态环境问题,威胁了绿洲稳定性。     

Comparison of effectiveness of introduced barn owls,Tyto javanica javanica,and rodenticide treatments on rat control in oil palm plantations

Journal of Pest Science - In Peninsular Malaysia, barn owls (Tyto javanica javanica) have been utilized as biological control of rats since the 1960s. In this study, the impact of introduced barn...