Agitation Conditions Research Articles

Production of Recombinant Human Asparaginase from Escherichia coli under Optimized Fermentation Conditions: Effect of Physicochemical Properties on Enzyme Activity

The recombinant human asparaginase (rhASP) plays an important role in the treatment of acute lymphoblastic leukemia. In the present work, volumetric mass transfer coefficient (kLa) values are derived from the E. coli cultivation under different agitation and aeration conditions, in order to improve the rhASP productivity. The aeration and agitation conditions were systematically optimized by the kLa. The maximum biomass (2.4 g/L) and rhASP (1.68 g/L) are achieved with the kLa of 0.024 s-1 at 1.5 lpm and 700 rpm process conditions. The kinetic properties of purified rhASP are also extensively studied and optimized for the maximal enzyme activity. The optimal pH, temperature and incubation duration conditions for accomplishing maximum enzyme activity are found to be 9.0, 40°C, and 30 min, respectively. The optimum substrate concentration and substrate specificity for the highest enzyme activity are of 0.07 M and L-asparagine, respectively. The enzyme activity (204 IU/mL) is significantly improved in the presence of sodium metal (Na+) ions and the inhibitors 2- mercaptoethanol, bromoacetic acid and urea have presented the highest inhibition rate on rhASP activity. The enzyme kinetic parameters Km and Vmax of the purified rhASP are recorded as 2.25 mM and 250 IU/mL, respectively. This work provides the extensive characteristic properties of rhASP enzyme, which enables us to place in a competition for the development of oncology drugs.

Physical Simulation Behavior of the Slag in Gas Injection with Non-consumable Lance and Porous Plug for Secondary Refining Operations

In secondary refining operations, injection of inert gases into the metal bath is required to generate stirring in order to produce cleaner steels with low sulfur, ultra-low contents of nitrogen and hydrogen gases as well as improving both temperature and composition homogenization of the steel and control the inclusions and floating of them. To demonstrate the viability and competitiveness of the non-consumable lance versus porous plugs, a series of experiments in a model of secondary refining ladle were carried out in previous research (Liu et al. in ISIJ Int 57:1971, 2017). Simulation of gas injection was carried out to test the two methods: porous plug and non-consumable lance. It was found that the porous plug injection involves shorter mixing times (10–12 s) when it is positioned in an intermediate point between the center and edge of the ladle and with the increase in the airflow injected, as compared to the lance. However, one problem with this technique in the actual plant is the metal oxidation because the liquid metal is exposed to air as a result of excessive agitation. In this work, gas injection through porous plug and non-consumable lance into secondary refining ladle was simulated by the addition of oils of different densities, to simulate the behavior of the slag of different viscosities. The results show that the non-consumable lance exhibits less water surface area exposed compared with the porous plug. This means less metal is exposed to the environment and less oxidation of it, under the same agitation conditions.

Hydrogels for diabetic eyes: Naltrexone loading, release profiles and cornea penetration.

Naltrexone (NTX) is a potent opioid growth factor receptor (OGFR) antagonist proved to be useful for treatment of ocular surface complications. The aim of this work was to explore the feasibility of designing NTX-imprinted 2-hydroxyethyl methacrylate-based hydrogels for sustained drug release on the ocular surface. Acrylic acid (AAc) and benzyl methacrylate (BzMA) were chosen as functional monomers able to form binding cavities mimicking OGFR binding sites for NTX. Imprinted hydrogels containing functional monomers loaded higher amounts of NTX compared to non-imprinted ones by simple soaking in drug aqueous solution. In addition, possibility of carrying out the loading and sterilization processes in one step was investigated. NTX release was evaluated both under agitated sink conditions and in a microfluidic flow chamber mimicking the hydrodynamic conditions of the eye, namely the small volume of lachrymal fluid and its renovation rate. Sustained release profiles together with adequate swelling degree (46 to 57% w/w), light transparency (over 85%) and oxygen permeability may make these hydrogels suitable candidates to NTX-eluting contact lenses. NTX-loaded and non-loaded discs successfully passed the chorioallantoic membrane test for potential ocular irritation and were cytocompatible with human mesenchymal stem cells. Finally, NTX-imprinted hydrogels tested in the bovine corneal permeability assay provided therapeutically relevant amounts of NTX inside the cornea, reaching drug levels similar to those attained with a concentrated aqueous solution in spite the discs showed sustained release.

Adhesion of filamentous fungi isolated from drinking water under different process conditions

Filamentous fungi (ff) are consistently recognized as drinking water (DW) inhabitants, typically harboured in biofilms. Their sessile behaviour is still poorly understood. This study aimed the evaluation of the influence of several abiotic factors (substratum, hydrodynamic conditions and nutrient availability) on biofilm formation by Penicillium brevicompactum and P. expansum isolated from DW. Fungal adhesion was quantified on high density polyethylene (HDPE) and polyvinyl chloride (PVC) surfaces using synthetic tap water (STW) and R2A broth, under stagnant or agitated (25 and 150 rpm) conditions. Fungal spore numbers were assessed after staining with Calcofluor White MR2 and epifluorescence microscopy. The surface charge and physicochemical properties of spores were characterized for a mechanistic understanding on the adhesion process. The adhesion kinetics of spores was represented accurately by the Logistic model, in which adhesion increased with time until a maximum level attained before spore germination (8 h after incubation). In general, P. brevicompactum demonstrated to adhere in a higher extent than P. expansum (12 × 104 spores/cm2vs 1.7 × 104 spores/cm2). Moreover, fungal adhesion was potentiated under stagnation and using R2A broth. HDPE and PVC allowed spore adhesion at similar extents. Adhesion predictions based on the extended Derjaguin, Landau, Verwey and Overbeek (XDLVO) theory corroborated the experimental results, highlighting the role of physicochemical surface properties on the adhesion of spores. These results allowed to refine a model for ff biofilm formation. The overall results help to understand key aspects determining the presence of P. brevicompactum and P. expansum biofilms in DW, where stagnant conditions and the presence of nutrients should be avoided to prevent ff biofilm formation.

An efficient method of Yarrowia lipolytica immobilization using oil- and emulsion-modified bacterial cellulose carriers

Abstract Background Yarrowia lipolytica is a nonconventional, dimorphic yeast with multiple biotechnological applications. Considering the size of Y. lipolytica cells and a plethora of its morphological forms (spherical cells or hyphae and pseudohyphae), it is highly difficult to select a suitable carrier for this useful microorganism. Bacterial cellulose (BC) is currently considered one of the most promising immobilization carriers. In the current study, the usefulness of oil- and emulsion-modified BCs as a carrier for Y. lipolytica immobilization was investigated. Static and agitated cultures were conducted in media supplemented with oil or emulsion to improve carrier porosity. Results It was found that the application of oil- and emulsion-modified BCs correlated with significantly higher efficiency of Y. lipolytica immobilization and hence higher yield than the yield achieved with an unmodified carrier. Increased efficiency of immobilization correlated with BC porosity-related parameters, which, in turn, depended on the size of oil droplets introduced into the culture medium. Moreover, changes in porosity-related parameters caused by the addition of oil or emulsion to the medium were observed when the cultures were conducted only under static conditions and not under agitated conditions. Conclusion The application of oil- and emulsion-modified BCs as carriers significantly increased the efficiency of Y. lipolytica immobilization as compared to unmodified BC. The addition of oil or emulsion to the culture medium can be a simple but effective method to modify the porosity of BC-based carriers. How to cite: Żywicka A, Wenelska K, Junka A, et al. An efficient method of Yarrowia lipolytica immobilization using oil- and emulsion-modified bacterial cellulose carrier. Electron J Biotechnol 2019;41. https://doi.org/10.1016/j.ejbt.2019.06.004 .

Storm-induced concentration of sulfurized, marine-origin, organic matter as a possible mechanism in the formation of petroleum source-rock

Abstract Black shales, though laminated, are not systematically synonymous with quiet conditions of deposition. A number of papers report about black shales yielding sedimentary structures echoing relatively high hydrodynamic conditions. Here we examine two sedimentary sequences pertaining to the Late Jurassic Argiles de Châtillon Formation of the Boulonnais area (Northernmost France), each of them including a meter-thick black-shale horizon. The two laminated black shales contain sulfurized (i.e., organic S-rich) organic matter, reaching a maximum of 9%. However, some differences set the two black shales apart. The lower one was deposited under calm, suboxic to anoxic, bottom-water conditions (the « classic » way); the upper black shale was deposited under oxic, agitated bottom-water conditions, not compatible with stable sulfidic conditions required for organic-matter sulfurization. The upper black shale experienced hydrodynamically-induced concentration of sulfurized, recalcitrant organic matter of marine origin. The sulfurized organic matter could be preserved and quantitatively accumulated owing to its non-putrescible nature, leading to the formation of a potential hydrocarbon source rock. The association of pyrite inclusions with sulfurized organic matter probably modified the hydrodynamic behavior of organic particles. The storm-induced remobilization and concentration of sulfurized organic matter implies that the “sulfurization factory” operated in proximal, shallow environments such as estuaries, mud flats or mangrove environments. Such a model is in agreement with findings from modern coastal/shelf environments (notably mangroves) and fully transposable to many other sedimentary situations of any geological time period.

TR OPTIMAL CONDITIONS FOR β-MANNANASE PRODUCTION BY BACILLUS CIRCULANS NT 6.7

Optimum medium for mannanase production is very important and producing medium has been shown effective for mannanase production from Bacillus circulans NT 6.7 but expensive. This work aim to find low cost medium and optimum condition for mannanase production from Bacillus circulans NT 6.7. 1.5% concentration of defatted copra meal as carbon source in shake flask resulted in highest growth and enzyme activities of 10.51±0.53 Log CFU/ml and 12.84±4.38 U/ml at 18 hours accordingly. Also, 2% defatted copra meal in fermenter produced highest growth of 7.94±0.41 Log CFU/ml and enzyme activities of 32.07±2.94 U/ml at 15 hours. Using taguchi design, optimal conditions for inoculum, agitation and aeration were 1%, 600 rpm and 0.75 vvm, respectively. Under this optimized condition, the mannanase experimental yield (29.57±4.81U/ml) closely matched the yield predicted by the statistical model (24.15U/ml) with R2 = 0.75. Using mineral salts medium, slightly increased in mannanase product was observed compare to previously used producing medium. Also, the cost of using mineral salts medium (cost per liter) is 27 time lower than the cost of producing medium. Suggesting that, mineral salt medium provides an economically feasible medium for producing mannanase from Bacillus circulans NT 6.7.

Determination of the Growing Season for Dry Land Agriculture Based on Thornthwaite Method in Bulukumba Regency, South Sulawesi Province

Drought events in a region are strongly related to the surface water availability and low rainfall. Furthermore, drought conditions impact the threat of food security. Bulukumba Regency in South Sulawesi Province is one of the areas that are prone to drought and the risk of crop failure. Therefore it needs anticipatory action to understand the drought index in this region. This study aims to identify the conditions of drought and agitation in the Bulukumba District of South Sulawesi Province. The results of the identification then became the basis for determining the growing season in various types of dryland agriculture cultivation. This study used the Thornthwaite method with a water balance and drought index approach. The sampling method in this study was purposive sampling. Data collection techniques carried out are observation techniques, laboratory analysis techniques, and documentation techniques. Data processing used quantitative techniques and descriptive approaches. The results of the study show that the growing season in dryland agriculture in November, December, and January based on the availability of water in this region. Conversely, the availability of water is very low in May to September and not appropriate to handling the cultivation.

Cell Suspension Culture of Plumbago europaea L. Towards Production of Plumbagin.

BackgroundPlumbagin is as an important bioactive secondary metabolite found in the roots of Plumbago spp. The only one species, Plumbago europaea L., grows wild in Iran. The therapeutic use of plumbagin is limited due to its insufficient supply from the natural sources as the plants grow slowly and take several years to produce quality roots.ObjectivesTo develop an efficient protocol for the establishment of callus and cell suspension cultures of P. europaea and to evaluate production of plumbagin in callus and cell suspension cultures of P. europaea for the first time.Material and MethodsStems and leaves explants were cultured on agar solidified (7% w/v) MS media, supplemented with different combination of 2, 4-D and Kin or 6-Benzylaminopurin (BA) for callus induction. The rapid growing calli were cultured in liquid Murashige and Skoog (MS) media in agitated condition for establishing cell suspension cultures of P. europaea. Moreover, the effects of light and dark conditions on the cell growth, cell viability and plumbagin production in cell suspension cultures of P. europaea were assessed.ResultsFriable calli were successfully induced using stem segments of P. europaea in semisolid MS medium supplemented with 1 mg.L-1 2, 4-Dichlorophenoxy acetic acid (2, 4-D) and 0.5 mg.L-1of kinetin (Kin). Optimal cell growth was obtained when the cells were grown in MS liquid media supplemented with 1 mg.L-1 2, 4-D and 0.5 mg.L-1 kinetin with an initial cell density of ~3×105 cells per ml incubated in the dark at 25 ± 1 °C. Growth curve revealed that the maximum cell growth rate (14.83×105 cells per ml) achieved on the day 18 and the highest plumbagin content (0.9 mg.g-1 Dry Cell Weight (DCW)) in the cells was obtained at the late exponential phase under dark condition which determined by High Performance Liquid Chromatography (HPLC) technique. Based on the obtained results, cell viability remained around 82.73% during the 18 days of cell culture in darkness. These suspension cultures showed continuous and stable production of plumbagin.ConclusionsOur study suggests that cell suspension cultures of P. europaea represent an effective system for biosynthesis and production of plumbagin as a valuable bioactive compound.

Boosting productivity of heterotrophic microalgae by efficient control of the oxygen transfer coefficient using a microbubble sparger

We isolated and characterized an oleaginous Thraustochytrid microalga. Based on 18 s rRNA sequencing, we classified this strain as Schizochytrium sp. SH103. Growth of these cells in a 5 L fermentor with a microbubble-type sparger (MTS) efficiently increased the oxygen transfer coefficient (OTC), while inducing only low external stress. For fermentation under conditions of low agitation (100 rpm) and aeration (3 L/min), the OTC was 8.6-fold higher using a MTS than a general type sparger (GTS). Under these same conditions, dry cell weight (DCW) productivity (0.54 vs. 1.26 g/L·h) and total lipid productivity (0.29 vs. 0.61 g/L·h) were each about 2.2-fold higher using a MTS than a GTS. Under conditions that optimized the OTC in a MTS fermentor (876 h−1), DCW productivity, total lipid productivity and DHA productivity were each 1.3-fold greater than non-optimized condition. Fed-batch fermentation experiments indicated the maximal DCW was 100.6 g/L at 48 h, maximal DCW productivity was 2.1 g/L·h, maximal lipid content was 48.6 g/L at 48 h, and lipid productivity was 1.0 g/L·h. Additionally, the results of experiments using a 50 L MTS fermentor (96.0 g/L DCW and 44.3 g/L total lipids at 48 h) were similar to those using the 5 L fermentor. Our results suggest that use of a MTS may help to achieve the efficient fermentation of heterotrophic microalgae for high cell densities.

Phycoremediation of lithium ions from aqueous solutions using free and immobilized freshwater green alga Oocystis solitaria: mathematical modeling for bioprocess optimization.

Lithium is registered as a serious pollutant that causes environmental damage to an irrigation water supply. Freshwater green alga (Oocystis solitaria) was studied for its potential to remove lithium ions from aqueous solutions. The Plackett-Burman design was applied for initial screening of six factors for their significances for the removal of lithium from aqueous solutions using Oocystis solitaria cells. Among the variables screened, pH, lithium concentration, and temperature were the most significant factors affecting lithium removal. Hence, the levels of these significant variables were further investigated for their interaction effects on lithium removal using the Box-Behnken statistical design. The optimum conditions for maximum lithium removal from aqueous solutions by Oocystis solitaria were the initial lithium concentration of 200 mg/L, contact time of 60 min, temperature of 30 °C, pH 5, and biomass of Oocystis solitaria cells of 1 g/L with agitation condition. Under the optimized conditions, the percentage of maximum lithium removal was 99.95% which is larger than the percentage of lithium removal recorded before applying the Plackett-Burman design (40.07%) by 2.49 times. The different properties of Oocystis solitaria, as an adsorbent, were explored with SEM and via FTIR analysis. The spectrum of FTIR analysis for samples of Oocystis solitaria cells before lithium biosorption showed different absorption peaks at 3394 cm-1, 2068 cm-1, 1638 cm-1, 1398 cm-1, 1071 cm-1, and 649 cm-1 which has been shifted to 3446 cm-1, 2924 cm-1, 1638 cm-1, 1384 cm-1, 1032 cm-1, and 613 cm-1, respectively, after lithium biosorption by the alga. The treatment of aqueous solution containing lithium with Oocystis solitaria cells immobilized in alginate beads removed 98.71% of lithium at an initial concentration of 200 mg/L after 5 h. Therefore, Oocystis solitaria may be considered as an alternative for sorption and removal of lithium ions from wastewaters.

A novel manufacturing process for car-T cell therapies in an automated stirred-tank bioreactor

Background & Aim Engineered Chimeric Antigen Receptor (CAR) T-cells therapies have recently gained FDA and EMA approval. Although the outstanding clinical and commercial achievement these products demonstrated, the high cost (exceeding $300,000) and the 9% failure rate of commercial doses need to be promptly addressed. Methods, Results & Conclusion Previous work undertaken by the group has proven the ability of primary T-cells to be successfully grown in a small-scale single use stirred-tank ambr® 250 bioreactor under agitated conditions. Building on these results, the same platform was used for the growth of CAR-T cells. A range of speeds was tested in order to find the optimal expansion conditions under which T-cells can be grown without losing their functionality. The outcomes from different experiments were compared with respect to cell yield, CAR-T cell fold expansion, viability, metabolite flux and T-cell subpopulations. pH and dissolved oxygen (dO2) concentration were monitored throughout the 7 days expansion allowing a better understanding of the culture requirements. Stirred-tank bioreactors are not commonly used for the manufacture of CAR-T cell products, therefore a further comparison in terms of growth and quality of CAR-T cells achieved in routinely used static and dynamic systems, as for example T-Flasks, G-Rex platform and Rocking Motion bioreactors, was needed. Seeding density, feeding regime and media composition were kept constant across the different platforms in order to be able to compare the outcome. Each condition was tested with different donors in order to assess the inter-donor variability in terms of growth kinetic and starting material. The stirred-tank bioreactor was proven to perform better in terms of fold expansion. The outcome from this extensive comparability study will be used to improve the current manufacturing process and reduce the manufacturing cost of the commercial product. Furthermore, using a stirred tank bioreactor will allow for an easier scaling-up of the process to 1-5 litres bioreactors. These can be used for the initial development of allogeneic CAR-T therapies.

The effect of midazolam administration for the prevention of emergence agitation in pediatric patients with extreme fear and non-cooperation undergoing dental treatment under sevoflurane anesthesia, a double-blind, randomized study.

Background: Sevoflurane is generally the preferred anesthetic agent for general anesthesia in pediatric patients, due to its rapid induction and recovery characteristics. However, it has been recognized that a major complication is emergence agitation when awakening from general anesthesia. The aim of this study was to evaluate the occurrence rate of emergence agitation in the operating room and postoperative recovery area following intraoperative administration of midazolam to pediatric patients under general anesthesia.Patients and methods: One hundred and twenty pediatric patients undergoing dental treatment under sevoflurane anesthesia were enrolled in this study. The patients were divided into three groups (n=40 each in the 0.1 mg/kg midazolam, 0.05 mg/kg midazolam, and control with saline groups). Midazolam or saline was injected intravenously approximately 30 minutes before the end of the dental treatment. We used the Richmond Agitation and Sedation Scale (RASS) to assess the level of sedation and drowsiness at emergence phase in the operating room. We also used the Pediatric Anesthesia Emergence Delirium Scale (PAED) to assess the level of agitation and delirium at the full recovery phase from anesthesia in the recovery area.Results: At the emergence phase, the incidence of emergence agitation in the 0.1 mg/kg midazolam group was significantly lower than in the other groups (p=0.0010). At the recovery phase, there was no significant difference among the three groups. The odds ratio between PAED score and RASS score was 4.0 using logistic regression analysis. The odds ratio between PAED score and Disability was 2.5.Conclusion: Administration of a single dose of 0.1 mg/kg midazolam dose significantly decreases the incidence of severe emergence agitation at the emergence after sevoflurane anesthesia, but not at the recovery phase. Furthermore, the evaluation of sedative and agitation condition using RASS score at emergence from anesthesia is useful to predict occurrence of agitation in the recovery phase.

Role of impeller design for the cultivation of stem cells in stirred-tank bioreactors

Background & Aim The routine use of stem cells and stem cell-derived cells in drug research and regenerative medicine requires the constant supply of high cell numbers in consistent quality. Stirred-tank bioreactors have emerged as promising cultivation systems, which facilitate close control of critical process parameters and have proven their value for efficient process scaling. Cell cultivation on microcarriers or as cell aggregates are two widely used methods for expanding anchorage-dependent cells in stirred-tank bioreactors. Suitable agitation conditions have to be established to keep cell aggregates or microcarriers in suspension while avoiding damaging shear forces. Methods, Results & Conclusion We have developed a novel 8-blade impeller to improve the cultivation of stem cells as aggregates and on microcarriers in the DASbox Mini Bioreactor System. Use of this impeller supported the formation and growth of cell aggregates in the bioreactor. Cell aggregate formation was more efficient than when using a conventional pitched-blade or Rushton-type impeller. Furthermore, microcarriers could be efficiently brought to and kept in suspension. Our results suggest that the impeller shape is an important parameter to consider when optimizing the agitation conditions in stirred-tank bioreactors.

Analysis of liquid-to-gas mass transfer, mixing and hydrogen production in dark fermentation process

The aim of this work was to investigate mixing and liquid-to-gas mass transfer of hydrogen in relation to hydrogen production in the dark fermentation process as a function of agitation conditions and digestate viscosity. Experiments were carried out in a baffled mechanically-stirred reactor equipped with a dual-stage impeller using five levels of viscosity. Biohydrogen production was studied using glucose as substrate under controlled pH. Three experimental techniques, namely local conductimetry, chemical decolorization and Planar Laser Induced Fluorescence were used to measure mixing time tm and describe the flow pattern. The effects of inter-impeller clearance and tracer injection position were also studied. Then, kLaH2 was deduced from dynamic deaeration/aeration experiments. Experimental results showed that biohydrogen production presented a maximum in the transitional flow regime (Re about 200), and fell under turbulent flow (Re> 1000). Similarly, the evolution of kLaH2 was better described by Re than by the volumetric power input, contrary to literature data. Finally, the Damköhler number showed that hydrogen production was limited by liquid-to-gas mass transfer in the laminar regime and that maximum reaction rate could be reached only due to dissolved H2 supersaturation in the liquid phase. Conversely, the steep decrease of H2 production under turbulence conditions could be attributed neither to mass transfer, nor to mixing conditions, highlighting a probable negative interaction between turbulent eddies and biomass aggregates. Regarding kLa∙tm, the transitional flow region also approached ideal mixing, which strengthened the conclusion that H2 production was optimized in the transition region in the dark fermentation process.

Diffusion coefficients and volume changes of beef meat during osmotic dehydration in binary and ternary solutions

Mass transfer phenomena taking into consideration volume changes during osmotic dehydration of beef meat under static and agitation conditions were studied. Experiments were conducted at low temperatures, 5–25 °C, in binary (20 or 25% NaCl) and ternary (40, 50 or 60% maltodextrin plus 5% NaCl) osmotic solutions. Meat volume changes during osmotic processing were strongly correlated with water loss of samples. A predictive model was developed to describe the volume reduction as a function of water loss and solute concentration. Moisture, solids and salt content were measured, and their corresponding mass transfer coefficients were calculated applying Fick’s second law of diffusion. The calculated moisture diffusion coefficients considering the volume changes (1.51 × 10−9–2.43 × 10−9 m2/s) were lower than those without considering this phenomenon (1.52 × 10−9–2.55 × 10−9 m2/s). Finally, a mathematical model was developed to predict moisture diffusion coefficients as a function of maltodextrin concentration and temperature, during osmotic dehydration of beef meat.

Commercial and non-commercial pectinase and cellulase on the enzymatic hydrolysis efficacy of rice husk and Tifton 85 hay

The aim of this study was to evaluate the action of commercial and non-commercial cellulase and pectinase on rice husk and Tifton 85 hay hydrolyses. The hydrolysis kinetics of the substrates with commercial cellulase and pectinase were evaluated and the hydrolysis at different temperature and agitation conditions was maximized using experimental design. The combined use of commercial and non-commercial enzymes under optimized conditions was evaluated. The pre-treatment of the residues was also investigated by milling and different concentrations of NaOH. Finally, the effect of the hydrolysis on the bromatological composition of the residues was evaluated. The best hydrolysis times of rice husk and Tifton 85 hay were 10 and 12h for commercial cellulase, 12 and 14h for non-commercial cellulase, 10 and 14h for commercial pectinase and 16 and 20h for non-commercial pectinase, respectively. The highest hydrolysis values were obtained using commercial cellulase with 1:50 (w:v enzyme:water) dilution rate, at 45ºC and 300 rpm agitation for both substrates, reaching 20.6% maximum percentage for Tifton 85 hay and 11.6% for rice husk. The combined use of commercial enzymes did not increase hydrolysis percentage. The pre-treatment using 7.5% NaOH and 0.5 mm grain size significantly increased the rice husk and Tifton 85 hay hydrolyses (60-80%), either using commercial cellulase or pectinase enzymes. The use of non-commercial enzymes provided 18-30% hydrolysis obtained from commercial ones. Bromatological analyzes indicated a reduction in neutral detergent fiber and acid detergent fiber content for rice husk and Tifton 85 hay when using pectinases and commercial cellulases.

Differences in Engineered Nanoparticle Surface Physicochemistry Revealed by Investigation of Changes in Copper Bioavailability During Sorption to Nanoparticles in the Aqueous Phase.

Sorption of chemical substances to nanoparticles (NPs) in the aqueous phase strongly influences NP physicochemisty, and investigations of these complex interactions can provide important insights into the environmental fate of NPs. The objective of the present study was to use differences in copper (Cu) bioavailability to investigate aqueous-phase sorption with NPs that had different physicochemical characteristics (silicon [Si], perovskite, and titanium dioxide NPs [TiO2 NPs]). Sorption of Cu with NPs was assessed by the presence of adsorbent in water and onto the NP surface after ultracentrifugation, and by changes in Cu bioavailability under static conditions during exposure of larval zebrafish, as well as under conditions of continuous agitation during exposure of the alga Chlorella vulgaris. The presence of TiO2 NPs reduced total Cu in the water column and Cu bioavailability (measured by growth inhibition, mortality, and metallothionein 2 gene expression), confirming Cu sorption to TiO2 NPs. Nanoparticle surface area was the most important factor that affected Cu sorption, as indicated by less bioavailable Cu in the presence of smaller TiO2 NPs. The surface area effect was consistent regardless of exposure conditions (alga, continuous agitation; zebrafish, static water) and was further supported by the fact that the lowest total Cu concentration in the water column was found in the presence of the smallest NP. The results differed with other NP types, for example, silicon NPs, in which Cu sorption was indicated by analytical chemistry, but sorption was not sufficient to significantly alter Cu bioavailability. The bioavailability tests did not indicate Cu sorption with perovskite NPs. The results demonstrate that surface area critically influences sorption, that Cu sorption as measured by bioavailability is not affected by agitation or static conditions, and that Cu sorption differs among types of NPs, indicating differences in their surface physicochemistry. Environ Toxicol Chem 2019;9999:1-11. © 2019 SETAC.

Effect of aeration and agitation on yeast inulinase production: a biocalorimetric investigation.

Air flow rate and agitation speed for inulinase production by Kluyveromyces marxianus were optimized based on metabolic heat release profiles. Shear stress and oxygen transfer (kLa) values were compared to assess the effects of aeration and agitation. At agitation rates of ≤ 100rpm, the oxygen mass transfer rates were small and eventually led to less inulinase production, but at agitation rates > 150rpm, loss of biomass resulted in less inulinase activity. Bio-reaction calorimeter (BioRc1e) experiment with aeration rates ≤ 0.5 lpm showed low kLa while at 1.5 lpm frothing of reactor contents caused loss of biomass and inulinase activity. The optimum conditions for aeration and agitation rate for K. marxianus in BioRc1e were 1 lpm and 150rpm. Heat yield values obtained for the substrate, product and biomass reinstated the ongoing metabolic process. The heat release pattern could be a promising tool for optimization of bioprocess and in situ monitoring, with a possibility of interventions during the biotransformation process. At optimized aeration and agitation conditions, a two-fold increase in inulinase activity could be noticed.

Microbial Synthesis and Characterization of Silver Nanoparticles using Alternaria alternata

In this study, extracellular biosynthesis of silver nanoparticles (AgNPs) was carried out using Alternaria alternata. AgNPs were synthesized using vegetative and cell-free filtrate methods. Silver nitrate (AgNO3) solution was reduced with fungal mycelial mass and cell-free filtrate at a ratio of 1:4 (v/v). Further, the effects of biosynthesis parameters (reaction time, reaction temperature, light and dark incubation, and static and agitated conditions) weredetermined. The structural integrity of the synthesized AgNPs was investigated through UV-visible spectrophotometry, X-ray diffraction (XRD) analysis, and transmission electron microscopy (TEM). Parameter optimization revealed that cell-free filtrate AgNP synthesis at 40 °C with constant agitation and incubation in darkness for 120 h using 1 mM AgNO3resulted in the highest absorbance value (at 400 nm). Further, TEM images showed that the synthesized AgNPs were spherical, homogeneous, and well dispersed while the XRD peaks confirmed the purity of the AgNPs obtained. The diameters of the AgNPs were found to range from 7.48 to 12.15 nm. This study identifies Alternaria alternata as a potential candidate for use in the industrial biosynthesis of AgNPs.