Search

• Understanding of the enzyme immobilization is required to optimize the processes. • Immobilization parameters and courses become critical to understand the process. • Changes in enzyme properties during immobilization can complicate the calculations. • Likely artifacts and problems to make these calculations are discussed. • Some advices to improve the understanding of immobilization processes are given. Biocatalytic processes continue to find increasing application in industry. Therefore enzyme immobilization has also become of increasing importance as a means of allowing enzyme containment within reactors operating in continuous mode or else separation of enzyme after use in (fed-)batch reactors, as well as potential recycle. Whilst much has been reported in the scientific literature about enzyme immobilization methods, in many cases the protocol leads to losses in enzyme activity. In this review we outline the reasons for loss of activity during immobilization and highlight suitable diagnostic tests to elucidate the precise cause and thereby methods to restore activity. The need for standardized reporting of immobilization methods is also emphasized as a means of benchmarking alternative approaches. [ABSTRACT FROM AUTHOR]

Abstract: Two hundred years ago, Henri Braconnot described a polysaccharide containing a substantial percent of nitrogen, later to be called chitin: that discovery stemmed from investigations on the composition of edible mushrooms and their nutritional value. The present interdisciplinary article reviews the major research topics explored by Braconnot, and assesses their importance in the light of our most advanced knowledge. Thus, the value of fungi, seafoods and insects is described in connection with the significance of the presence of chitin itself in foods, and chitinases in the human digestive system. The capacity of chitin/chitosan to depress the development of microbial pathogens, is discussed in terms of crop protection and food preservation. Other topics cherished by Braconnot, such as the isolation of pectin from a large number of plants, and inulin from the Helianthus tubers, are presented in up-to-date terms. Acids isolated from plants at that early time, led to enormous scientific advancements, in particular the glyoxylic acid and levulinic acid used for the preparation of soluble chitosan derivatives that paved the way to a number of applications. An opportunity to trace the origins of the carbohydrate polymers science, and to appreciate the European scientific heritage. [Copyright &y& Elsevier]

Abstract: A modelling approach is proposed to represent the decreasing evolutions observed on cell concentration, biosynthetic activity and substrate consumption at late fermentation times during processing when using a cell recycling (membrane) apparatus and continuous feeding. Indeed, during such processes, which are over rather long time periods, the cell populations may have to withstand various stresses (physical, environmental and chemical) involving a decrease of the global activity of the cell populations. This study was carried out with l-lactic acid fermentation using Lactobacillus casei as a model system and the modelling approach was based on the definitions of a critical cell concentration and a stress zone. The results obtained corresponded to what was expected. [Copyright &y& Elsevier]

Alkaline phosphatases (APs), E.C. 3.1.3.1, are non-specific phosphomonoesterases optimally active under alkaline conditions. They are classically known to be homodimeric metalloenzymes. This quaternary structure has been considered necessary for activity, although the relationship between quaternary structure and activity is not well understood. Recombinant Pyrococcus abyssi AP was previously isolated and characterized, appearing to have two active quaternary structures on native polyacrylamide gel electrophoresis: a monomer and a homodimer. The purpose of the present work was to determine the actual quaternary structure of P. abyssi AP in solution, by isolating each of the two quaternary forms and establishing the parameters governing the assembly and dissociation of the dimer. pH appeared to be an important parameter: in acidic media, the monomer/dimer ratio shifted towards monomer. Buffer composition also affected the quaternary structure: at the same pH, in potassium phosphate buffer, the two quaternary structures were observed, whereas in tris(hydroxymethyl)aminomethane hydrochloride buffer, only the dimer was observed. Metals bound to the enzyme were found to be involved in the stability of the quaternary structure. Indeed, the P. abyssi AP obtained upon removal of the metals was monomeric. Reactivation of the latter was achieved with variable efficiency. From these experiments, no active monomer could be isolated, leading the conclusion that the active form of P. abyssi AP is the homodimer. [Copyright &y& Elsevier]

In the present work, L-glutamic acid production by Corynebacterium glutamicum fermentation on date juice extracts applying two fed-batch feeding modes, pulsed and continuous, were investigated. According to the obtained results, the continuous feeding fed-batch mode was found to be the most efficient process. Moreover the continuous feeding rate mode with a feeding medium containing date juice sugars enriched with ammonium sulfate was found even more favorable as it enhances the L-glutamic acid production by approximately 2.35 fold more than the batch culture and by about 1.17 fold more than the pulsed feeding. In this respect, comparing the traditional batch culture to the continuously fed culture with a medium containing date juice sugars with ammonium sulfate showed increases of 135.47% in L-glutamic acid production, 104% in productivity, 39.09% in biomass, and 47.69% in the yield respectively allowing us to reach a final L-glutamic acid concentration of about 138 g/L, the highest ever published. [ABSTRACT FROM AUTHOR]

Abstract: Dehydration of microorganisms, rendering them anhydrobiotic, is often an efficient method for the short and long term conservation of different strain-producers. However, some biotechnologically important recombinant bacterial strains are extremely sensitive to conventional treatment. We describe appropriate conditions during dehydration of the recombinant Escherichia coli strain HB 101 (GAPDH) that can result dry cells having a ∼88% viability on rehydration. The methods entails air-drying after addition of 100mM trehalose to the cultivation medium or distilled water (for short term incubation). [Copyright &y& Elsevier]

Purpose: In diseases where there is no real consensus regarding treatment modalities, promoting shared decision-making can contribute to improving safety and quality of care. This is the case in low- or intermediate-risk localized prostate cancer (PC) treatment. The aim of this study was to investigate the preferences guiding men's decisions regarding the characteristics of the treatment strategies for PC to help physicians adopt a more patient-centered approach. Methods: This prospective multicenter study used a discrete choice experiment (DCE). The attributes and the modalities were identified from a qualitative study and a literature review. Relative preferences were estimated using a logistic regression model. Interaction terms (demographic, clinical and socio-economic characteristics) were added to the model to assess heterogeneity in preferences. Results: 652 men were enrolled in the study and completed a questionnaire with 12 pairs of hypothetical therapeutic alternatives between which they had to choose. Men's choices were significantly negatively influenced by the risk of impotence and urinary incontinence, death, and the length and frequency of care. They preferred treatments with a rescue possibility in case of deterioration or recurrence and the use of innovative technology. Surprisingly, the possibility of undergoing prostate ablation negatively influenced their choice. The results also highlighted differences in trade-offs according to socio-economic level. Conclusion: This study confirmed the importance of considering patients' preferences in the decision-making process. It appears essential to better understand these preferences to allow physicians to improve communication and promote case-by-case decision-making. [ABSTRACT FROM AUTHOR]

Alkaline phosphatases (APs) are non-specific phosphomonoesterases which are of interests for basic research as well as for practical applications. So far, Pyrococcus abyssi AP is the only AP which was isolated from a hyperthermophilic archaeon. In the present work, expression of this enzyme using mesophilic hosts was investigated. When P. abyssi AP was produced by Escherichia coli using the regulation of the T7 promoter, only 0.6 mg of recombinant enzyme per liter of culture were obtained. Using the same system with the co-expression of tRNAs related to the rare codons AGA, AGG and ATA, the expression level was 14-fold higher. The methylotrophic yeast Pichia pastoris was also used to produce P. abyssi AP. A recombinant pPICZαA was constructed in order to have the AP secreted in the culture medium. A Pc. pastoris cell line producing P. abyssi AP was studied in 1.2 l working volume fermentor. Secretion was achieved with low expression yield but the recombinant enzyme appeared more active than when produced in E. coli. [Copyright &y& Elsevier]

The feasibility of baker's yeast production using fruits and peels of Opuntia ficus indica (OFI) as carbohydrate feedstock was investigated. Two response surface methodologies involving central composite face centered design (CCFD) were successfully applied. The effects of four independent variables on baker's yeast production from OFI fruit juice was evaluated using the first CCFD. The best results were obtained with 24 H of inoculum age, 30 °C temperature, 200 rpm of agitation, and 10% inoculum size. At the maximum point, the biomass concentration reached 9.29 g/L. A second CCFD was performed to optimize the sugar extraction from OFI fruit peels. The potential of these latter as a fermentation substrate was determined. From the experimental results, the OFI fruit peel is an appropriate carbon source for the production of baker's yeast. The maximum biomass concentration was 12.51 g/L. Different nitrogen supplements were added to promote the yields of baker's yeast. Corn steep liquor was found to be the best alternative nutrient source of casein hydrolysate and yeast extract for baker's yeast production. [ABSTRACT FROM AUTHOR]

Purpose: Family caregivers play an important role in caring for patients with advanced cancer. To become competent, individuals must draw on and mobilise an adequate combination of resources. Our goal was to identify the skills developed by caregivers of patients with advanced cancer and the associated resources mobilised. We chose to do it with partners of patients with colon cancer.Methods: The study used a cross-sectional qualitative design based on 20 individual interviews and a focus group. Partners were recruited from patients treated in three hospitals of France. Semi-structured interviews were conducted until data saturation was achieved. Each interview was transcribed verbatim, and thematic analyses were performed to extract significant themes and subthemes.Results: Results from the individual and focus group interviews showed that the skills implemented by the partners (in domains of social relationships and health, domestic, organisational, emotional and well-being dimensions) were singular constructs, dependant on if resources (personal, external and schemes) may have been missing and insufficient. In addition, partners may have had these resources but not mobilised them.Conclusion: The identification of the skills and associated resources could allow healthcare professionals better identifying and understanding of the difficulties met by partners in taking care of patients. This could enable them to offer appropriate support to help the caregivers in their accompaniment. [ABSTRACT FROM AUTHOR]

The article presents a study that investigates the fermentation potential of the lactate production of Corynebacterium glutamicum 2262. It describes the method of the study that observes coupling aerobic cell growth and fermentation phase. It notes the result of the study which reveals the capability of Corynebacterium glutamicum 2262 to produce various fermentation products from pyruvate.

Highlights: [•] A new application of cloud point phenomenon in phenanthrene biodegradation. [•] Stable increase of the solubility of phenanthrene after a cloud point treatment. [•] Obtained higher concentration of soluble phenanthrene improved its biodegradation. [•] Perspective application of cloud point in the enhancement of PAHs biodegradation. [Copyright &y& Elsevier]

This study describes the biodegradation of phenanthrene in aqueous media in the presence and in the absence of a surfactant, Brij 30. Biodegradations were performed using either Pseudomonas putida DSMZ 8368 or a bacterial consortium Pyr01 isolated from one PAHs-polluted site. P. putida degraded phenanthrene to form 1-hydroxy-2-naphthoic acid (1H2Na) as the major metabolite. LC-MS analysis revealed the production of complementary intermediates in the presence of Brij 30, showing intense ions at mass-to-charge ratios ( m/z) 97 and 195. Higher phenanthrene biodegradation rate was obtained in the presence of Brij 30. Conversely, in the case of Pyr01consortium, the addition of Brij 30 (0.5 g L) had a negative effect on biodegradation: no phenanthrene biodegradation products were detected in the medium, whereas a production of several intermediates ( m/z 97, 195 and 293) was obtained without surfactant. New results on phenanthrene metabolism by P. putida DSMZ 8368 and Pyr01 consortium in the presence and in the absence of Brij 30 we obtained. They confirm that the knowledge of the effect of a surfactant on bacterial cultures is crucial for the optimization of surfactant-enhanced PAHs biodegradation. [ABSTRACT FROM AUTHOR]

To improve glutamic acid production from Corynebacterium glutamicum 2262 using date juice, a culture medium was screened and optimized using the statistical experimental designs of Plackett-Burman and response-surface methodology. In the first step, a two-level Plackett-Burman design was adopted to select the most important nutrients influencing the glutamic acid production, which showed that the date juice sugars, urea, peptone, and glycine betaine were the most significant ingredients ( P < 0.05). Finally, response surface Box-Behnken design was employed to develop a mathematical model to identify the optimum concentrations of key components for higher glutamic acid production, which revealed the following: date juice (45 g/L), urea (16.9 g/L), peptone (15 g/L), and glycine betaine (12 g/L). The high correlation between the predicted and observed values indicated the validity of the model. Glutamic acid concentration increased significantly with optimized medium (33.2 g/L) when compared with non-optimized medium (12 g/L). [ABSTRACT FROM AUTHOR]

Abstract: In the present study surfactant addition with the help of either a mechanical dispersion or a thermal treatment was applied in order to increase the solubility and the bioavailability of phenanthrene in aqueous media, and therefore to promote its biodegradation. Among four tested surfactants (Tween 80, Brij 30, sodium dodecyl sulphate and rhamnolipids), Brij 30 (0.5gL−1) showed the best results allowing us to attain about 20mgL−1 of soluble phenanthrene. An additional thermal treatment at 60°C for 24h, 200rpm permitted to increase the solubility of phenanthrene in the presence of Brij 30 (0.5gL−1) to about 30mgL−1. Higher dispersions of phenanthrene particles as well as the reduction of their size were obtained using Ultra-Turrax and French press. The biodegradation of phenanthrene by Pseudomonas putida was then investigated. The reduction of size of phenanthrene particles by mechanical dispersion did not influence its biodegradation, suggesting that P. putida consumed only soluble phenanthrene. The addition of Brij 30 (0.5gL−1) permitted to obtain more phenanthrene metabolized. The use of Brij 30 coupled with a transitory heating of phenanthrene-containing medium at 60°C led to an even more complete biodegradation. This might be a promising way to enhance biodegradation of PAHs. [Copyright &y& Elsevier]

In Corynebacterium glutamicum, the activity of the 2-oxoglutarate dehydrogenase complex was shown to be controlled by the phosphorylation of a 15-kDa protein OdhI by different serine/threonine protein kinases. In this paper, the phosphorylation status and kinetics of OdhI dephosphorylation were assessed during glutamate producing processes triggered by either a biotin limitation or a temperature upshock from 33°C to 39°C. A dephosphorylation of OdhI in C. glutamicum 2262 was observed during the biotin-limited as well as the temperature-induced glutamate-producing process. Deletion of pknG in C. glutamicum 2262 did not affect the phosphorylation status of OdhI during growth and glutamate production phases triggered by a temperature upshock, though a 40% increase in the specific glutamate production rate was measured. These results suggest that, under the conditions analyzed, PknG is not the kinase responsible for the phosphorylation of OdhI in C. glutamicum 2262. The phosphorylation status of OdhI alone is, as expected, not the only parameter that determines the performance of a specific strain, as no clear relation between the specific glutamate production rate and OdhI phosphorylation level was demonstrated. [ABSTRACT FROM AUTHOR]

[Display omitted] • Mineral concentration is important in cladode powder with a granule size of 350 µm. • A low amount of HMF (inhibitor) was detected in the prickly pear medium. • No furfural (inhibitor) was detected in the prickly pear medium. • Prickly pear cladodes are a potential raw material for the production of lactic acid. • Prickly pear fruit peels can be used as a cheap medium for lactic acid production. Opuntia ficus indica (OFI) waste was evaluated as a fermentation feedstock for lactic acid production using Lactobacillus plantarum. Dilute acid pretreatment of the OFI cladodes (OFIC) was performed for extracting maximum fermentable sugars by optimizing process parameters using statistical optimization method. The best results were obtained with HCl 1% (v/v), temperature 120 °C, residence time 40 min, granulation 350 µm and substrate loading 5% (w/v), the sugar concentration reached 24 g/L with low concentration of hydroxymethylfurfural. The feasibility of producing lactic acid from OFI fruit peel (OFIFP) as a source of carbon was also investigated. Lactobacillus plantarum was shown to have a capacity for lactic acid production from OFIC 350 (granulation 350 µm) hydrolysate and OFIFP extract without detoxification. The highest lactic acid yields of 0.46 and 0.78 g/g were obtained from enzymatic hydrolysate of pretreated OFIC 350 and OFIFP extract, respectively. [ABSTRACT FROM AUTHOR]

Abstract: The galactose-β1,3-glucuronosyltransferase I (GlcAT-I) catalyzes the transfer of glucuronic acid from UDP-α-d-glucuronic acid onto the terminal galactose of the trisaccharide glycosaminoglycan–protein linker region of proteoglycans. This enzyme plays a key role in the process of proteoglycan assembly since the completion of the linkage region is essential for the conversion of a core protein into a functional proteoglycan. To investigate the enzymatic properties of human GlcAT-I, we established an expression system for producing a soluble form of enzyme in the methylotrophic yeast Pichia pastoris and developed a three-step purification procedure using a combination of anion exchange, cation exchange and heparin chromatographies. This procedure yielded 1.6mg homogeneous enzyme from 200ml yeast cell culture, with a specific activity value of 1.5μmol/min/mg protein. Analysis of the specificity of GlcAT-I towards Galβ1–3Gal and Galβ1–4GlcNAc derivatives known as substrates of the β1,3-glucuronosyltransferases, showed that the enzyme exhibited a strict selectivity towards Galβ1–3Gal structures. Thus, the large source of purified active enzyme allowed the determination of the kinetic parameters of GlcAT-I towards the donor substrate UDP-GlcA and the acceptor substrate digalactoside Galβ1–3Gal. [Copyright &y& Elsevier]

The use of date juice as a substrate for lactic acid production was investigated. Various nitrogen sources were compared with yeast extract for efficient lactic acid production by Lactobacillus casei subsp. rhamnosus. Among different nitrogen sources added to date juice (yeast extract, ammonium sulfate, tryptic soy, urea, peptone, and casein hydrolysate), yeast extract was the most efficient. The effect of yeast extract could have been due to its B vitamin content. The addition of five B vitamins at less than 25 mg/l to date juice with any nitrogen source enhanced lactic acid production to some extent, except for date juice with yeast extract or urea or peptone. The most significant increase was obtained with ammonium sulfate. Half of the yeast extract content (10 g/l) in a supplemented date juice could be replaced by a mixture of B vitamins at less than 25 mg/l, and ammonium sulfate at 2.6 g/l with no significant decrease in lactic acid production. [Copyright &y& Elsevier]

Describes the characterization of a highly thermostable alkaline phosphatase from the thermophilic archaeon Pyrococcus abyssi. Classical and unique features of the Pyrococcus abyssi AP gene; Dependence of Pyrococcus abyssi AP on divalent cations; Effectivity of Pyrococcus abyssi recombinant AP for DNA dephosphorylation.