pH-Responsive Cellulose/Silk/Fe3O4 Hydrogel Microbeads Designed for Biomedical Applications.

In this study, cellulose/Fe3O4 hydrogel microbeads were prepared through the sol-gel transition of a solvent-in-oil emulsion using various cellulose-dissolving solvents and soybean oil without surfactants. Particularly, 40% tetrabutylammonium hydroxide (TBAH) and 40% tetrabutylphosphonium hydroxide (TBPH) dissolved cellulose at room temperature and effectively dispersed Fe3O4, forming cellulose/Fe3O4 microbeads with an average diameter of ~15 µm. Additionally, these solvents co-dissolved cellulose and silk, allowing for the manufacture of cellulose/silk/Fe3O4 hydrogel microbeads with altered surface characteristics. Owing to the negatively charged surface characteristics, the adsorption capacity of the cellulose/silk/Fe3O4 microbeads for the cationic dye crystal violet was >10 times higher than that of the cellulose/Fe3O4 microbeads. When prepared with TBAH, the initial adsorption rate of bovine serum albumin (BSA) on the cellulose/silk/Fe3O4 microbeads was 18.1 times higher than that on the cellulose/Fe3O4 microbeads. When preparing TBPH, the equilibrium adsorption capacity of the cellulose/silk/Fe3O4 microbeads for BSA (1.6 g/g) was 8.5 times higher than that of the cellulose/Fe3O4 microbeads. The pH-dependent BSA release from the cellulose/silk/Fe3O4 microbeads prepared with TBPH revealed 6.1-fold slower initial desorption rates and 5.2-fold lower desorption amounts at pH 2.2 than those at pH 7.4. Cytotoxicity tests on the cellulose and cellulose/silk composites regenerated with TBAH and TBPH yielded nontoxic results. Therefore, cellulose/silk/Fe3O4 microbeads are considered suitable pH-responsive supports for orally administered protein pharmaceuticals.

Determinants of microbial colonization on microplastics through wastewater treatment processes: The role of polymer type and sequential treatment.

This study examines the microbial colonization characteristics of microplastics (MPs) in wastewater treatment plants (WWTPs), focusing on polymer types (High-Density Polyethylene (HDPE) and Polyethylene Terephthalate (PET)) and various stages of wastewater treatments. Through individual and sequential deployment approaches, the research aimed to identify the determinants of bacterial colonization on MPs, whether they were introduced at each stage of treatment individually or in sequence from primary to tertiary stages. The study revealed that the stage of wastewater treatment profoundly influenced bacterial colonization on the polymer types MPs, with bacterial attachment being largely niche-specific. HDPE showed increased sensitivity to wastewater composition, leading to selective biofilm formation. For instance, in HDPE, Firmicutes accounted for 25.1 ± 0.04 % during primary treatment, while Alphaproteobacteria increased significantly in the tertiary treatment to 19.8 ± 0.1 %. Conversely, PET exhibited a stochastic pattern of bacterial colonization due to differences in surface hydrophilicity. Additionally, in sequential deployments, a notable shift towards stochastic bacterial attachment on MPs, particularly with HDPE was observed. The Shannon diversity values for MP biofilms were consistently higher than those for wastewater across all stages, with PET showing an increase in diversity in sequential deployments (Shannon diversity: 5.01 ± 0.03 for tertiary stage). These findings highlight the critical role of MPs as carriers of diverse bacteria, emphasizing the necessity for strategies to mitigate their impact in WWTPs. This study presents a significant advancement in our understanding of the interactions between MPs and microbial populations in WWTP environments.

Key factors for the survival of Lactiplantibacillus plantarum IDCC 3501 in manufacturing and storage.

Functional microbiome development has steadily increased; with this, the viability of microbial strains must be maintained not only after the manufacturing process but also at the time of consumption. Survival is threatened by various unavoidable factors during freeze-drying and shelf storage. Here, the aim was to optimize the manufacturing process of the functional strain Lactiplantibacillus plantarum IDCC 3501 after freeze-drying and storage. Explosive growth was achieved using a medium composition with two nitrogen sources and a mineral, and growth was drastically improved by neutralizing the medium pH during the culture of L. plantarum IDCC 3501. Culture optimization involved a smaller cell size, leading to less intracellular free water. Moreover, when maltodextrin (MD) powder was directly added to the harvested cells, some intracellular free water was extracted from the bacterial cells, resulting in a dramatic increase in the viability of L. plantarum IDCC 3501 after freeze-drying and subsequent storage. Furthermore, MD enhanced survival in a dose-dependent manner. Bacterial survival was correlated with lysozyme tolerance; therefore, the positive result might have been caused by the osmotic dehydration of intracellular free water, which would potentially damage the bacterial cells via ice crystallization and/or a phase transition during freeze-drying. These critical factors of L. plantarum IDCC 3501 processing provide perspectives on survival issues for manufacturing microbiome strains. KEY POINTS: • Culture conditions for probiotic bacteria were optimized for high growth yield. • Osmotic dehydration improved bacterial survival after manufacturing and shelf storage. • Reduction in intracellular free water content is crucial for intact survival.

Development of Blended Biopolymer-Based Photocatalytic Hydrogel Beads for Adsorption and Photodegradation of Dyes.

Blended biopolymer-based photocatalytic hydrogel beads were synthesized by dissolving the biopolymers in 1-ethyl-3-methylimidazolium acetate ([Emim][Ac]), adding TiO2, and reconstituting the beads with ethanol. The incorporation of modifying biopolymer significantly enhanced the adsorption capacity of the cellulose/TiO2 beads. Cellulose/carrageenan/TiO2 beads exhibited a 7.0-fold increase in adsorption capacity for methylene blue (MB). In contrast, cellulose/chitosan/TiO2 beads showed a 4.8-fold increase in adsorption capacity for methyl orange (MO) compared with cellulose/TiO2 beads. In addition, cellulose/TiO2 microbeads were prepared through the sol-gel transition of the [Emim][Ac]-in-oil emulsion to enhance photodegradation activity. These microbeads displayed a 4.6-fold higher adsorption capacity and 2.8-fold higher photodegradation activity for MB than the millimeter-sized beads. Furthermore, they exhibited superior dye removal efficiencies for various dyes such as Congo red, MO, MB, crystal violet, and rhodamine B, surpassing the performance of larger beads. To expand the industrial applicability of the microbeads, biopolymer/TiO2 magnetic microbeads were developed by incorporating Fe2O3. These magnetic microbeads outperformed millimeter-sized beads regarding the efficiency and time required for MB removal from aqueous solutions. Furthermore, the physicochemical properties of magnetic microbeads can be easily controlled by adjusting the type of biopolymer modifier, the TiO2 and magnetic particle content, and the ratio of each component based on the target molecule. Therefore, biopolymer-based photocatalytic magnetic microbeads have great potential not only in environmental fields but also in biomedical fields.

Reswellable alginate/activated carbon/carboxymethyl cellulose hydrogel beads for ibuprofen adsorption from aqueous solutions.

In this study, alginate (Alg) composite beads were prepared by blending with activated carbon (AC) to enhance adsorption capacity for ibuprofen and carboxymethyl cellulose (CMC) to create a reswellable hydrogel. The dried Alg/AC/CMC composite beads could be recovered to sizes and morphologies similar to the initial hydrogel states via a simple reswelling process; however, the dried Alg/AC composite beads without CMC could not be recovered to the initial hydrogel state. Following the reswelling process, the dried Alg/AC/CMC beads demonstrated an 86 % recovery (qe = 34.0 mg/g) in the adsorption capacity for ibuprofen compared to the initial hydrogel beads (qe = 39.6). In contrast, the reswelled Alg/AC beads exhibited only 18 % (qe = 8.6) of the initial adsorption capacity (qe = 48.1). We elucidated the effects of the substitution degree of CMC, AC content, and solution pH on the reswelling property and ibuprofen adsorption capacity of the Alg/AC/CMC composite beads. The adsorption kinetics and isotherms of the prepared composite beads in the hydrogel and reswelled states fit the pseudo-second-order and Langmuir models, respectively. Furthermore, the reswelled Alg composite beads exhibited high adsorption capacity (>93 %) after 10 cycles. Taken together, our findings indicate that the Alg/AC/CMC composite beads can be used as adsorbents without a considerable decrease in adsorption performance by reswelling the beads with distilled water after long-term storage in a dry state.

Exploring the vertical transport of microplastics in subsurface environments: Lab-scale experiments and field evidence.

Microplastics (MPs) defined as smaller 5 mm plastic particles have received increasing attention due to their global occurrence and potential toxicity. This study investigated the effects of environmental factors (rainfall intensity, 13 and 29 mm/h) and MP characteristics (morphology (fiber, flake, and film), polymer type (polypropylene (PP), polyethylene terephthalate (PET), and polystyrene (PS)) and size (100-300, 300-500, and 500-1000 µm)) on the vertical transport of MP in unsaturated soil conditions using lab-scale column experiments. Additionally, the occurrence and characteristics of MP detected in soil/sediment (total 13 samples) and groundwater samples (total 6 samples) were explored in the field study. Laboratory-scale column experiments revealed that heavy rainfall intensity (29 mm/h) increased the degree of MP vertical transport in unsaturated soil conditions and MP fibers showed the greatest vertical mobility among the various morphologies of MPs assessed. For the polymer type and size, the lighter PP polymer or the larger size of MP (500-1000 µm) showed higher mobility. In the field study, a statistical difference in MP abundance was observed depending on the population density and degree of urban development in both soil and groundwater samples. Comparing to the two different types of environmental media samples obtained from the same site, there was a significant difference in the composition of polymer types present while statistically no difference in MP abundance was observed between the two media samples (i.e., soil or sediment and groundwater). In addition, MP fibers and polyethylene (PE) were predominantly detected in our two study areas. These results suggest that various types of MP can pass through the unsaturated zone by water infiltration, even if it takes a long time to reach groundwater. Overall, we found that the degree of vertical transport of the MPs was highly sensitive to environmental conditions and MP characteristics.

Denitrification dynamics in unsaturated soils with different porous structures and water saturation degrees: A focus on the shift in microbial community structures.

Despite its environmental significance, little is known about denitrification in vadose zones owing to the complexity of such environments. Here, we investigated denitrification in unsaturated soils with different pore distributions. To this end, we performed batch-type denitrification experiments and analyzed microbial community shifts before and after possible reactions with nitrates to clarify the relevant denitrifying mechanism in the microcosms. For quantitative comparison, pore distribution in the test soil samples was characterized based on the uniformity coefficient (Cu) and water saturation degree (SD). Micro-CT analysis of the soil pore distribution confirmed that the proportion of bigger-sized pores increased with decreasing Cu. However, oxygen diffusion into the system was controlled by SD rather than Cu. Within a certain SD range (51-67%), the pore condition changed abruptly from an oxic to an anoxic state. Consequently, denitrification occurred even under unsaturated soil conditions when the SD increased beyond 51-67%. High throughput sequencing revealed that the same microbial species were potentially responsible for denitrification under both partially (SD 67%), and fully saturated (SD of 100%) conditions, implying that the mechanism of denitrification in a vadose zone, if it exists, might be possibly similar under varying conditions.

Long-Term Examination of Water Chemistry Changes Following Treatment of Cyanobacterial Bloom with Coagulants and Minerals.

The abundant growth in cyanobacterial blooms poses severe ecological threats with a high risk to aquatic organisms and global public health. Control of cyanobacterial blooms involves spraying cyanobacteria removal materials, including coagulants. However, little is known about the fate of the coagulated-cyanobacteria-laden water. Here, we examined long-term changes in water quality following treatment with various coagulants and minerals for cyanobacterial removal when the coagulated cyanobacterial cells were not removed from the water. An experiment in a controlled water system tested the effects of six different compounds, one conventional coagulant, two natural inorganic coagulants, and three minerals. All tested coagulants and minerals exhibited >75% of cyanobacterial removal efficiency. However, compared to the control, higher concentrations of nitrogen were observed from some samples treated during the experimental period. After 20 months, the final total phosphorus concentration of the raw water increased 20-fold compared to the initial concentration to 11.82 mg/L, indicating significant nutrient release over time. Moreover, we observed that the decomposition of sedimented cyanobacterial cells caused the release of intracellular contents into the supernatant, increasing phosphorous concentration over time. Therefore, cyanobacterial cells should be removed from water after treatment to prevent eutrophication and maintain water quality.

Adsorptive Behavior of Cu2+ and Benzene in Single and Binary Solutions onto Alginate Composite Hydrogel Beads Containing Pitch Pine-Based Biochar.

In this study, we prepared alginate composite hydrogel beads containing various compositions of biochar produced from pitch pine (Pinus rigida) for the removal of Cu2+ and benzene from model pollutant solutions. The properties of the alginate/biochar hydrogel beads were evaluated using scanning electron microscopy, Fourier transform infrared spectroscopy, and Brunauer−Emmet−Teller analyses. Adsorption behavior of alginate/biochar hydrogel beads indicated that the adsorption capacities for Cu2+ (28.6−72.7 mg/g) were enhanced with increasing alginate content, whereas the adsorption capacities for benzene (20.0−52.8 mg/g) were improved with increasing biochar content. The alginate/biochar hydrogel beads exhibited similar adsorption capacities for Cu2+ and benzene in the concurrent system with Cu2+ and benzene compared to those in a single pollutant system. Adsorption kinetics and isotherm studies of the alginate/biochar hydrogel beads followed the pseudo-second-order model (r2 = 0.999 for Cu2+, and r2 = 0.999 for benzene), and Langmuir model (r2 = 0.999 for Cu2+, and r2 = 0.995 for benzene). In addition, alginate/biochar hydrogel beads (containing 1 and 4% biochar) exhibited high reusability (>80%). Therefore, alginate/biochar hydrogel beads can be applied as adsorbents for the removal of multiple pollutants with different properties from wastewater.

Laccase-Mediator System Using a Natural Mediator as a Whitening Agent for the Decolorization of Melanin.

In this study, a laccase-mediator system (LMS) using a natural mediator was developed as a whitening agent for melanin decolorization. Seven natural mediators were used to replace synthetic mediators and successfully overcome the low redox potential of laccase and limited access of melanin to the active site of laccase. The melanin decolorization activity of laccases from Trametes versicolor (lacT) and Myceliophthora thermophila (lacM) was significantly enhanced using natural mediators including acetosyringone, syringaldehyde, and acetovanillone, which showed low cytotoxicity. The methoxy and ketone groups of natural mediators play an important role in melanin decolorization. The specificity constants of lacT and lacM for melanin decolorization were enhanced by 247 and 334, respectively, when acetosyringone was used as a mediator. LMS using lacM and acetosyringone could also decolorize the melanin present in the cellulose hydrogel film, which mimics the skin condition. Furthermore, LMS could decolorize not only synthetic eumelanin analogs prepared by the oxidation of tyrosine but also natural melanin produced by melanoma cells.

Biogeochemical Alteration of an Aquifer Soil during In Situ Chemical Oxidation by Hydrogen Peroxide and Peroxymonosulfate.

In this study, the effects of in situ chemical oxidation (ISCO) on the biogeochemical properties of an aquifer soil were evaluated. Microcosms packed with an aquifer soil were investigated for 4 months in two phases including oxidant exposure (phase I) and biostimulation involving acetate addition (phase II). The geochemical and microbial alterations from different concentrations (0.2 and 50 mM) of hydrogen peroxide (HP) and peroxymonosulfate (PMS) were assessed. The 50 mM PMS-treated sample exhibited the most significant geochemical changes, characterized by the decrease in pH and the presence of more crystalline phases. Microbial activity decreased for all ISCO-treated microcosms compared to the controls; particularly, the activity was severely inhibited at high PMS concentration exposure. The soil microbial community structures were shifted after the ISCO treatment, with the high PMS causing the most distinct changes. Microbes such as the Azotobacter chroococcum and Gerobacter spp. increased during phase II of the ISCO treatment, indicating these bacterial communities can promote organic degradation despite the oxidants exposure. The HP (low and high concentrations) and low concentration PMS exposure temporarily impacted the microbial activity, with recovery after some duration, whereas the microbial activity was less recovered after the high concentration PMS exposure. These results suggest that the use of HP and low concentration PMS are suitable ISCO strategies for aquifer soil bioattenuation.

In vivo determination of the Infrared-A protection factor on human skin.

BACKGROUND: Chronic exposure to infrared A (IR-A) irradiation causes photoaging. However, daily or acute exposure to IR-A rarely induces erythema or pigmentation. Thus, evaluation of the physiological changes taking place on the skin surface is insufficient for clinical investigations. MATERIALS AND METHODS: We fabricated a novel device to obtain the IR-A protection factor (IPF) on human skin. This device consists of an artificial light source that mimics the actual IR-A intensity of sunlight, and a spectrophotometer to measure the spectral reflectance on the skin surface. The IPF can be determined by measuring the difference in spectral reflectance on the skin before and after the use of products and can be verified by the statistical criterion. A validation study was performed using different light intensities and two experimenters. Finally, we monitored the IPF on 12 commercial cosmetics. RESULTS: After considering the IPF and L*-values, we selected the optimal sample and performed a validation study. Neither the intensity of IR-A irradiation or the experimenters significantly affected the IPF. 12 commercial products exhibited their own IPF values and were verified by statistical criteria, with one exception. CONCLUSION: The present IPF evaluation method was concluded to be robust and reliable. This method is simple and safe for the subjects, and could be helpful for the development of IR-A protection products and the confirmation of product performances.

Microfluidic pore model study of precipitates induced by the pore-scale mixing of an iron sulfate solution with simulated groundwater.

Precipitates induced by the pore-scale mixing of iron sulfate solutions with simulated groundwater were investigated using a microfluidic pore model to assess the environmental impacts of the infiltration of acid mine drainage into a shallow aquifer. This model was employed to visualize the formation of precipitates in a porous network and to evaluate their physicochemical influences on pore flow. Four types of groundwater (Na-HCO3, Na-SO4, Na-Cl, and Ca-Cl) were evaluated, and precipitation rates were calculated by processing images of precipitates in the pores captured via microscopy. The results showed that all groundwater types formed a yellow-brownish precipitate at the interface of the iron solution and simulated groundwater flow. Microscopic X-ray analyses demonstrated that precipitate morphology varied with groundwater type. Faster precipitation was observed in the following order by groundwater type: Na-HCO3 > Na-Cl > Na-SO4 > Ca-Cl, which was attributed to the different stability constants of the major anions in each simulated groundwater with Fe ions. Chemical equilibrium models suggested that precipitates were Fe minerals, with FeOOH as the predominant form consistent with the results of X-ray photoelectron spectrometry. The presence of FeOOH implies that precipitates may serve as an effective sorption barrier against some nutrients and heavy metals for the underlying groundwater. However, dye-flow experiments suggested that the precipitates may clog aquifer pores, thereby altering hydrogeological properties in the aquifer.

Identification of iron and sulfate release processes during riverbank filtration using chemical mass balance modeling.

Various hydrogeochemical processes can modify the quality of river water during riverbank filtration (RBF). Identifying the subsurface processes responsible for the bank-filtered water quality is challenging, but essential for predicting water quality changes and determining the necessity of post-treatment. However, no systematic approach for this has been proposed yet. In this study, the subsurface hydrogeochemical processes that caused the high concentrations of total iron (Fe) and sulfate (SO42-) in the bank-filtered water were investigated at a pilot-scale RBF site in South Korea. For this purpose, water quality variations were monitored in both the extraction well and the adjacent river over five months. The volumetric mixing ratio between the river water and the native groundwater in the RBF well was calculated to understand the effect of mixing on the quality of water from the well and to assess the potential contribution of subsurface reactions to water quality changes. To identify the subsurface processes responsible for the evolution of Fe and SO42- during RBF, an inverse modeling based on the chemical mass balance was conducted using the water quality data and the calculated volumetric mixing ratio. The modeling results suggest that pyrite oxidation by abundant O2 present in an unsaturated zone could be a primary process explaining the evolution of total Fe and SO42- during RBF at the study site. The presence of pyrite in the aquifer was indirectly supported by iron sulfate hydroxide (Fe(SO4)(OH)) detected in oxidized aquifer sediments.

Immobilization of laccase via cross-linked enzyme aggregates prepared using genipin as a natural cross-linker.

Genipin is a nontoxic natural cross-linker that was successfully used to prepare cross-linked enzyme aggregates (CLEAs) of Trametes versicolor laccase. The recovered activity of CLEAs was influenced by the co-solvent type, genipin concentration, cross-linking time, preparation pH, and bovine serum albumin (BSA; amino group feeder) concentration. The characteristics of CLEAs prepared using genipin under optimal conditions (genipin-BSA-CLEAs) were compared with those of typical CLEAs prepared using glutaraldehyde or dextran polyaldehyde. Genipin-BSA-CLEAs were nano-sized (average diameter, approximately 700 nm), had a ball-like shape, showed a narrow size distribution, and exhibited the highest substrate affinity among the prepared CLEAs. The thermal stability of genipin-BSA-CLEAs was 6.8-fold higher than that of free laccase, and their pH stability was also much higher than that of free laccase in the tested range. Additionally, genipin-BSA-CLEAs retained 85% of their initial activity after 10 cycles of reuse. Particularly, genipin-BSA-CLEAs showed higher thermal and pH stability than CLEAs that were cross-linked using glutaraldehyde. Therefore, genipin represents an alternative to toxic compounds such as glutaraldehyde during cross-linking to prepare CLEAs.

Effect of hydrogen bond donor on the choline chloride-based deep eutectic solvent-mediated extraction of lignin from pine wood.

In this work, twenty-five kinds of choline chloride (ChCl)-based deep eutectic solvents (DESs) containing acid, hydroxyl, amide, and binary hydrogen bond donors (HBDs) were prepared and successfully used to pretreat pine wood powder. As a result of the pretreatment, the glucan content in the pretreated biomass was increased, whereas the contents of hemicellulose and lignin were significantly decreased. The biomass pretreatment efficiency of the DESs had improved with increasing the polarity and hydrogen bond acidity (α) of the DESs. Among the studied DESs, ChCl:lactic acid:formic acid (1:1:1) with the highest α value was the most efficient DES in extracting lignin from biomass. The pretreated biomass also showed an enhanced enzymatic saccharification yield owing to the decreased particle size of the biomass and reduced content of hemicellulose and lignin. During the pretreatment process of biomass using DESs, the extracted lignin could be recovered successfully, with a yield of up to 60% and purity of over 90%. The molecular weight of the extracted lignin was much lower than that of the native cellulolytic enzyme lignin. The DES used for pretreatment process could be also successfully reused with high recovery yield of DES and high retention of delignification capacity.

Effect of Cellulose Solvents on the Characteristics of Cellulose/Fe2O3 Hydrogel Microspheres as Enzyme Supports.

Cellulose hydrogels are considered useful biocompatible and biodegradable materials. However, as few cellulose-dissolving solvents can be used to prepare cellulose hydrogel microspheres, the use of unmodified cellulose-based hydrogel microspheres for enzyme immobilization remains limited. Here, we prepared cellulose/Fe2O3 hydrogel microspheres as enzyme supports through sol-gel transition using a solvent-in-oil emulsion. Cellulose-dissolving solvents including 1-ethyl-3-methylimidazolium ([Emim][Ac]), an aqueous mixture of NaOH and thiourea, tetrabutylammonium hydroxide, and tetrabutylphosphonium hydroxide were used to prepare regular shaped cellulose/Fe2O3 microspheres. The solvent affected microsphere characteristics like crystallinity, hydrophobicity, surface morphology, size distribution, and swelling properties. The immobilization efficiency of the microspheres for lipase was also significantly influenced by the type of cellulose solvent used. In particular, the lipase immobilized on cellulose/Fe2O3 microspheres prepared using [Emim][Ac] showed the highest protein loading, and its specific activity was 3.1-fold higher than that of free lipase. The immobilized lipase could be simply recovered by a magnet and continuously reused.

Stereoselective Synthesis of Tropanes via a 6π-Electrocyclic Ring-Opening/ Huisgen [3+2]-Cycloaddition Cascade of Monocyclopropanated Heterocycles.

The synthesis of tropanes via a microwave-assisted, stereoselective 6π-electrocyclic ring-opening/ Huisgen [3+2]-cycloaddition cascade of cyclopropanated pyrrole and furan derivatives with electron-deficient dipolarophiles is demonstrated. Starting from furans or pyrroles, 8-aza- and 8-oxabicyclo[3.2.1]octanes are accessible in two steps in dia- and enantioselective pure form, being versatile building blocks for the synthesis of pharmaceutically relevant targets, especially for new cocaine analogues bearing various substituents at the C-6/C-7 positions of the tropane ring system. Moreover, the 2-azabicyclo[2.2.2]octane core (isoquinuclidines), being prominently represented in many natural and pharmaceutical products, is accessible via this approach.

Production of L-Theanine Using Escherichia coli Whole-Cell Overexpressing γ-Glutamylmethylamide Synthetase with Bakers Yeast.

L-Theanine, found in green tea leaves has been shown to positively affect immunity and relaxation in humans. There have been many attempts to produce L-theanine through enzymatic synthesis to overcome the limitations of traditional methods. Among the many genes coding for enzymes in the L-theanine biosynthesis, glutamylmethylamide synthetase (GMAS) exhibits the greatest possibility of producing large amounts of production. Thus, GMAS from Methylovorus mays No. 9 was overexpressed in several strains including vectors with different copy numbers. BW25113(DE3) cells containing the pET24ma::gmas was selected for strains. The optimal temperature, pH, and metal ion concentration were 50°C, 7, and 5 mM MnCl2, respectively. Additionally, ATP was found to be an important factor for producing high concentration of L-theanine so several strains were tested during the reaction for ATP regeneration. Bakers yeast was found to decrease the demand for ATP most effectively. Addition of potassium phosphate source was demonstrated by producing 4-fold higher L-theanine. To enhance the conversion yield, GMAS was additionally overexpressed in the system. A maximum of 198 mM L-theanine was produced with 16.5 mmol/l/h productivity. The whole-cell reaction involving GMAS has greatest potential for scale-up production of L-theanine.

Evaluating perfluorooctanesulfonate oxidation in permanganate systems.

Permanganate (PM) has shown to be able to oxidize a range of organic contaminants including perfluorooctane sulfonate (PFOS). However, mechanisms of PFOS removal by PM have been questioned. To provide clarity to what may be happening to PFOS in PM systems, here we evaluated the ability of PM on PFOS destruction by conducting studies similar to previous studies that reported PFOS destruction which included PM solutions and PM combined with persulfate (PS). We also evaluated if addition of various soluble catalysts could enhance PM's potential to breakdown PFOS. We observed no PFOS destruction by PM. We also show that the F- and SO42- generation reported in a published study as evidence that PM was breaking bonds in PFOS were found below or not significantly higher than reported limits of quantitation and that SO42- impurities in technical PM approach the reported SO42- levels. For PM-PS systems, heterogeneous PFOS distribution was observed when subsampling reaction vessels at different depths and "salting-out" of PFOS was evident. In addition, subsequent sonication and filtering of the samples led to the apparent disappearance of most of the PFOS, which was an artifact arising from the behavior of PFOS aggregates or potential hemi-micelle formation. Given these findings, addition of salts may have application for collecting or concentrating PFOS and other PFAAs in a remediation or water treatment strategy.