Formation Of Gluconic Acid Research Articles

Stimuli-Responsive DNA-Based Hydrogels on Surfaces for Switchable Bioelectrocatalysis and Controlled Release of Loads.

The assembly of enzyme [glucose oxidase (GOx)]-loaded stimuli-responsive DNA-based hydrogels on electrode surfaces, and the triggered control over the stiffness of the hydrogels, provides a means to switch the bioelectrocatalytic functions of the hydrogels. One system includes the assembly of GOx-loaded, pH-responsive, hydrogel matrices cross-linked by two cooperative nucleic acid motives comprising permanent duplex nucleic acids and "caged" i-motif pH-responsive duplexes. Bioelectrocatalyzed oxidation of glucose leads to the formation of gluconic acid that acidifies the hydrogel resulting in the separation of the i-motif constituents and lowering the hydrogel stiffness. Loading of the hydrogel matrices with insulin results in the potential-triggered, glucose concentration-controlled, switchable release of insulin from the hydrogel-modified electrodes. The switchable bioelectrocatalyzed release of insulin is demonstrated in the presence of ferrocenemethanol as a diffusional electron mediator or by applying an electrically wired integrated matrix that includes ferrocenyl-modified GOx embedded in the hydrogel. The second GOx-loaded, stimuli-responsive, DNA-based hydrogel matrix associated with the electrode includes a polyacrylamide hydrogel cooperatively cross-linked by duplex nucleic acids and "caged" G-quadruplex-responsive duplexes. The hydrogel matrix undergoes K+-ions/crown ether-triggered stiffness changes by the cyclic K+-ion-stimulated formation of G-quadruplexes (lower stiffness) and the crown ether-induced separation of the G-quadruplexes (higher stiffness). The hydrogel matrices demonstrate switchable bioelectrocatalytic functions guided by the stiffness properties of the hydrogels.

Optical and microstructural characteristics of CuO thin films by sol gel process and introducing in non-enzymatic glucose biosensor applications

In this work, we reported the synthesis of copper oxide (CuO) thin films by sol-gel process assisted by spin coating technique at room temperature. Chemical characterization of CuO precursor solution was determined by Fourier Transformed Infrared Spectroscopy (FTIR) and thermal stability by Thermogravimetric/ Differential Scanning Calorimetry Analysis (TGA/DSC). The influence of variation parameters for the thin films were studied: speed deposition, number of layers and annealing effect on the deposited films. The optical properties of CuO thin films were analyzed using a spectrophotometer (UV–vis-NIR) in a range of 300−1100 nm. The calculated band gap for CuO this films was between 3.35 eV–3.89 eV for as deposited thin films and 2.4 eV–3.6 eV for annealed films. Homogeneous deposited films with monoclinic structure were determined using scanning electron microscopy (SEM) and X-Ray Diffraction respectively. The compositions and homogeneous elemental distributions were analyzed by energy dispersive spectroscopy (EDAX). From the profilometer measurement, the thicknesses of the CuO thin films prepared were found to be ∼1/2−1 μm, depending of speed deposition. Finally, CuO solution precursor and thin films were implemented in a glucose test which showed the formation of gluconic acid by FTIR, indicating the glucose oxidation due to CuO interaction.

Synthesis-property-performance relationships of multifunctional bacterial cellulose composites fermented in situ alkali lignin medium

This work demonstrates a unique approach of utilizing alkali lignin (AL), as smart additive to in situ BC fermentation in which it concurrently acts as promoter to microbial growth as well as reinforcing filler for fabrication of multifunctional composites. Traditionally, BC fermentation is accompanied by inhibitor formation with sudden drop in pH leading to low yield and biomass growth. AL due to its antioxidant nature prevents formation of gluconic acid as byproduct, at ∼0.25 wt.% AL based on inhibitory byproduct kinetics. Interestingly, AL self-assembles to form primary and secondary structures in BC pores, resulting in simultaneous improvement in thermal stability as well as toughness. The BC/AL films show strong UV-blocking capacity with prolonged radical scavenging activity and preventing browning of freshly cut apples making it suitable as food packaging. Therefore, present work opens up new avenues for fabrication of high-performance BC-based composites through selection of smart materials which can simultaneously improve BC bioprocessing.

Insulin Delivery from Glucose-Responsive, Self-Assembled, Polyamine Nanoparticles: Smart "Sense-and-Treat" Nanocarriers Made Easy.

Polyamine-salt aggregates (PSA) are biomimetic soft materials that have attracted great attention due to their straightforward fabrication methods, high drug-loading efficiencies, and attractive properties for pH-triggered release. Herein, a simple and fast multicomponent self-assembly process was used to construct cross-linked poly(allylamine hydrochloride)/phosphate PSAs (hydrodynamic diameter of 360 nm) containing glucose oxidase enzyme, as a glucose-responsive element, and human recombinant insulin, as a therapeutic agent for the treatment of diabetes mellitus (GI-PSA). The addition of increasing glucose concentrations promotes the release of insulin due to the disassembly of the GI-PSAs triggered by the catalytic in situ formation of gluconic acid. Under normoglycemia, the GI-PSA integrity remained intact for at least 24 h, whereas hyperglycemic conditions resulted in 100 % cargo release after 4 h of glucose addition. This entirely supramolecular strategy presents great potential for the construction of smart glucose-responsive delivery nanocarriers.

Exploring K2G30 Genome: A High Bacterial Cellulose Producing Strain in Glucose and Mannitol Based Media.

Demands for renewable and sustainable biopolymers have rapidly increased in the last decades along with environmental issues. In this context, bacterial cellulose, as renewable and biodegradable biopolymer has received considerable attention. Particularly, acetic acid bacteria of the Komagataeibacter xylinus species can produce bacterial cellulose from several carbon sources. To fully exploit metabolic potential of cellulose producing acetic acid bacteria, an understanding of the ability of producing bacterial cellulose from different carbon sources and the characterization of the genes involved in the synthesis is required. Here, K2G30 (UMCC 2756) was studied with respect to bacterial cellulose production in mannitol, xylitol and glucose media. Moreover, the draft genome sequence with a focus on cellulose related genes was produced. A pH reduction and gluconic acid formation was observed in glucose medium which allowed to produce 6.14 ± 0.02 g/L of bacterial cellulose; the highest bacterial cellulose production obtained was in 1.5% (w/v) mannitol medium (8.77 ± 0.04 g/L), while xylitol provided the lowest (1.35 ± 0.05 g/L) yield. Genomic analysis of K2G30 revealed a peculiar gene sets of cellulose synthase; three bcs operons and a fourth copy of bcsAB gene, that encodes the catalytic core of cellulose synthase. These features can explain the high amount of bacterial cellulose produced by K2G30 strain. Results of this study provide valuable information to industrially exploit acetic acid bacteria in producing bacterial cellulose from different carbon sources including vegetable waste feedstocks containing mannitol.

IR-Compatible PDMS microfluidic devices for monitoring of enzyme kinetics

Coupling infrared (IR) spectroscopy to microfluidic devices provides a powerful tool for characterizing complex chemical and biochemical reactions. Examples of microfluidic devices coupled with infrared spectroscopy have been limited, however, largely due to the difficulties associated with fabricating systems in common infrared transparent materials like CaF2. Recent reports have shown that polydimethylsiloxane (PDMS) can be used as an IR transparent substrate when fabricated with thin layers. The use of soft lithography with PDMS expands the library of possible designs that can be achieved for IR measurements in microfluidics. In initial reports with thin PDMS, the target analytes were small molecules; however, IR spectroscopy offers a powerful tool to study protein structure and reactions. Here, a PDMS microfluidic device compatible with IR spectroscopy was fabricated by means of spin-coating of PDMS pre-polymer to obtain thin PDMS microfluidic features. The device was comprised of only PDMS and IR absorption of PDMS was significantly minimized due to the thickness (∼40 μm) of the PDMS layer. The use of thin PDMS allowed for measuring the amide I and II vibrational bands of proteins that have been difficult to measure in other microfluidic devices. To demonstrate the power of the system, the microfluidic device was successfully used to measure the enzyme kinetics as one class of important biochemical reactions with broad use in a variety of fields from medicine to biotechnology. As a model, the reaction of glucose oxidase with glucose was tracked by following the formation of gluconic acid. Michaelis-Menten kinetics from the device were compared with bulk solution measurements and found to be in good agreement.

A Novel Colorimetric Immunoassay of Ultrasensitive Alpha-Fetoprotein Sensing in Magnetic Bead-Based Mimic Enzyme-Chromogenic Substrate System

This work described a simple and feasible colorimetric immunoassay. Using hemin (a horseradish peroxidase (HRP) mimic enzyme) and the mimic enzyme-chromogenic substrate system, it can qualitatively and quantitatively detect a-fetoprotein (AFP) at an ultralow concentration. The glucose oxidase (GOx) catalyzed oxidation of glucose leads to the formation of gluconic acid and hydrogen peroxide (H2O2). The latter can oxidize 4-aminoatipyrine (4-AAP) to chromogenic products, and the reaction was catalyzed by hemin. With the increase of H2O2, the absorbance increased, and the color of the solution changed from colorless to pink. On the basis of the system, monitored by recording the color or absorbance (λ = 505 nm), a new immunoassay protocol with GOx-labeled anti-AFP detection antibody was designed. A wide linear dependence was obtained in the range from 0.075 to 280 ng mL-1 with a low detection limit (LOD) of 0.0247 ng mL-1 (S/N = 3).

Porous structured CuO-CeO2 nanospheres for the direct oxidation of cellobiose and glucose to gluconic acid

Porous-structured CuO-CeO2 nanospheres were synthesized using a hydrothermal method and were tested as catalysts for the direct oxidation of cellobiose to gluconic acid. Catalytic reaction along with catalyst characterization results and 18O-oxygen isotope labeled experiments revealed that the surface lattice oxygen of CuO in CuO-CeO2 nanospheres was consumed during the oxidation of cellobiose. This provides a direct evidence of our previous work (Amaniampong et al., Angew. Chem. Int. Ed. 54 (2015) 8928–8933). Characterization results further suggested that the lattice oxygen in CeO2 did not participate in the oxidation; nonetheless, the addition of CeO2 to CuO enhanced the surface area of the catalyst composite which was crucial for the reaction. The spent catalyst upon re-oxidation regained its activity. In addition, isotope labeled deuterium oxide (D2O) experiments suggested that hydrogen exchange between the solvent and the substrate (glucose) are not involved in the mechanistic formation of gluconic acid and confirmed the solvent had no direct influence in the formation of gluconic acid.

Peroxidase-like activity of gum kondagogu reduced/stabilized palladium nanoparticles and its analytical application for colorimetric detection of glucose in biological samples

The peroxidase like activity of gum kondagogu (Cochlospermum gossypium) reduced/stabilized palladium nanoparticles (GK-Pd NPs) was investigated using 3,3′,5,5′-tetramethylbenzidine (TMB) as a chromogen in the presence of H2O2. GK-Pd NPs could catalyze oxidization of TMB by H2O2 to produce a blue coloured oxidation product which is quantified using UV–vis spectrophotometer (UV–vis) at λmax of 652nm. Effect of various reaction conditions, such as temperature, pH and nanoparticle concentration on the catalytic activity of GK-Pd NPs was studied. The peroxidase like catalysis of GK-Pd NPs was found to follow Michaelis–Menten kinetics. The calculated kinetic parameters of the GK-Pd NPs catalysis showed strong affinity towards both, the substrate (H2O2) and the chromogen (TMB). It is known that the oxidation of glucose by glucose oxidase (GOx) enzyme leads to the formation of gluconic acid and H2O2 is generated as a by product. The H2O2 released in this reaction was consequently quantified using GK-Pd NPs as peroxides mimics and TMB as chromogen. Thus, a combination of above two reactions was exploited to establish glucose concentrations. Under the optimum conditions, the linear range of this method was from 10μM to 1000μM with the detection limit down to 6.0μM. Moreover, the developed method was applied to detect glucose in serum samples and the results were in good agreement with standard GOD-POD method.

High-temperature reduction improves the activity of rutile TiO2 nanowires-supported gold-copper bimetallic nanoparticles for cellobiose to gluconic acid conversion

Titania nanowires (NW) supported gold–copper (Au–Cu) bimetallic nanoparticles were synthesized and pretreated in hydrogen and air at 300, 500 and 700°C, for the one-pot conversion of cellobiose to gluconic acid. Catalyst samples were characterized by temperature-programmed desorption of NH3, Fourier transform infrared spectroscopy (FT-IR), Energy-dispersive X-ray spectroscopy, Field emission scanning electron microscopy (FE-SEM), X-ray powder diffraction (XRD), Transmission electron microscopy (TEM), and X-ray photoelectron spectroscopy (XPS). The structure and activity of Au–Cu/TiO2 NW were highly affected by the pretreatment conditions. Catalyst samples reduced in H2 and at higher temperatures resulted better catalytic performance as compared with those calcinated in air at the same temperature. The influence of support, calcination temperature and atmosphere as well as gold content on the catalytic performance of Au–Cu/TiO2 NWs are investigated. The characterization results suggested high hydrogen reduction temperature created oxygen vacant sites on the titania NW support. This is consequently associated with the stabilization of highly reactive oxygen species at the periphery of the metal–support interface. Interactions between the metals and the titania NWs support and between the promoter and the active metal enhanced the formation of gluconic acid.

Development of novel sourdoughs with in situ formed exopolysaccharides from acetic acid bacteria

Acetic acid bacterial strains were investigated for their ability to grow in aerobic and anaerobic sourdoughs (dough yield = 500) produced from wheat, whole wheat, spelt and rye flour and to produce effectual levels of exopolysaccharides (EPS) within 48 h of fermentation. The strains Kozakia (K.) baliensis DSM 14400 and Neoasaia (N.) chiangmaiensis NBRC 101099 were chosen. Both strains remained in aerobic doughs produced with any of the tested flours. For EPS optimization, 50, 75 and 100 g/l sucrose (sucrose) were added to the doughs and the parameters EPS, pH value, sucrose, acetic acid and gluconic acid were analysed. Results were subjected to statistical data analysis. In sucrose-enriched spelt dough, K. baliensis reached up to 49 g/kg EPS, followed by whole wheat (33 g/kg), wheat and rye dough (32–36 g/kg). N. chiangmaiensis produced 24–29 g/kg EPS in spelt and whole wheat doughs. EPS formation was positively correlated with start sucrose dosage. In contrast to this, a rather negative relation between acetic acid formation and start sucrose was found for both strains, and a positive relation with fermentation time. In spelt doughs, K. baliensis and N. chiangmaiensis produced up to 100 and 21–38 mM/kg acetic acid, respectively. Formation of gluconic acid was positively correlated with start sucrose dosage. K. baliensis produced at maximum 165 mM/kg flour and N. chiangmaiensis at maximum 119 mM/kg flour. Remaining sucrose contents were always positively correlated with start sucrose dosage, whereby both tested strains reached remaining sucrose amounts below 50 g/kg. A subsequent experiment with K. baliensis in backslopped spelt dough showed that this strain can remain in the dough over at least 8 days.

Magnetic bead-based enzyme-chromogenic substrate system for ultrasensitive colorimetric immunoassay accompanying cascade reaction for enzymatic formation of squaric acid-iron(III) chelate.

This work reports on a simple and feasible colorimetric immunoassay with signal amplification for sensitive determination of prostate-specific antigen (PSA, used as a model) at an ultralow concentration by using a new enzyme-chromogenic substrate system. We discovered that glucose oxidase (GOx), the enzyme broadly used in enzyme-linked immunosorbent assay (ELISA), has the ability to stimulate in situ formation of squaric acid (SQA)-iron(III) chelate. GOx-catalyzed oxidization of glucose leads to the formation of gluconic acid and hydrogen peroxide (H2O2). The latter can catalytically oxidize iron(II) to iron(III), which can rapidly (<1 min) coordinate with the SQA. Formation of the iron-squarate complex causes the color of the solution to change from bluish purple to bluish red accompanying the increasing absorbance with the increment of iron(III) concentration. On the basis of the SQA-iron(III) system, a new immunoassay protocol with GOx-labeled anti-PSA detection antibody can be designed for the detection of target PSA on capture antibody-functionalized magnetic immunosensing probe, monitored by recording the color or absorbance (λ = 468 nm) of the generated SQA-iron(III) chelate. The absorbance intensity shows to be dependent on the concentration of target PSA. A linear dependence between the absorbance and target PSA concentration is obtained under optimal conditions in the range from 1.0 pg mL(-1) to 30 ng mL(-1) with a detection limit (LOD) of 0.5 pg mL(-1) (0.5 ppt) estimated at the 3Sblank level. The sensitivity displays to be 3-5 orders of magnitude better than those of most commercialized human PSA ELISA kits. In addition, the developed colorimetric immunoassay was validated by assaying 12 human serum samples, receiving in good accordance with those obtained by the commercialized PSA ELISA kit. Importantly, the SQA-based immunosensing system can be further extended for the detection of other low-abundance proteins or biomarkers by controlling the target antibody.

Amine-stabilized small gold nanoparticles supported on AlSBA-15 as effective catalysts for aerobic glucose oxidation

The preparation of gold nanoparticles (AuNPs) supported on the alumina- and amine-modified silica SBA-15 and systematic studies of their structural properties and behaviour in glucose oxidation are presented. The AuNPs were deposited using either direct reduction or colloidal deposition method. Catalysts were characterized by nitrogen adsorption, XRD, XPS, STEM, HRTEM, 27Al MAS NMR, 1H NMR, DLS, HPLC, ICP MS and thermogravimetry to determine structure/activity relationship in the aerobic glucose oxidation. The highest catalytic activity ca. 800mmolGA/gAumin was achieved for AuNPs prepared by a direct reduction method using SBA-15 first modified with alumina (1wt.%) and then with aminopropyl groups serving as strong adsorption sites. It was notably larger than shown by commercial Au/Al2O3 and Au/TiO2 catalysts (380 and 340mmolGA/gAumin, respectively). Owing to excellent and stable dispersion of AuNPs and their small sizes <6nm, the catalysts showed very good stability, demonstrated in 7 reaction cycles. The impact of specific functionalities on the size, dispersion and stability of AuNPs was discussed and high selectivity towards gluconic acid formation was demonstrated.

Do new cellulolytic enzyme preparations affect the industrial strategies for high solids lignocellulosic ethanol production?

Production of ethanol from lignocellulosic materials has a promising market potential, but the process is still only at pilot/demonstration scale due to the technical and economical difficulties of the process. Operating the process at very high solids concentrations (above 20% dry matter-DM) has proven essential for economic feasibility at industrial scale. Historically, simultaneous saccharification and fermentation (SSF) was found to give better ethanol yields compared to separate hydrolysis and fermentation (SHF), but data in literature are typically based on operating the process at low dry matter conditions. In this work the impact of selected enzyme preparation and processing strategy (SHF, presaccharification and simultaneous saccharification and fermentation-PSSF, and SSF) on final ethanol yield and overall performance was investigated with pretreated wheat straw up to 30% DM. The experiments revealed that an SSF strategy was indeed better than SHF when applying an older generation enzyme cocktail (Celluclast-Novozym 188). In case of the newer product Cellic CTec 2, SHF resulted in 20% higher final ethanol yield compared to SSF. It was possible to close the mass balance around cellulose to around 94%, revealing that the most relevant products could be accounted for. One observation was the presence of oxidized sugar (gluconic acid) upon enzymatic hydrolysis with the latest enzyme preparation. Experiments showed gluconic acid formation by recently discovered enzymatic class of lytic polysaccharides monoxygenases (LPMO's) to be depending on the processing strategy. The lowest concentration was achieved in SSF, which could be correlated with less available oxygen due to simultaneous oxygen consumption by the yeast. Quantity of glycerol and cell mass was also depending on the selected processing strategy.

Heterologous expression of Pycnoporus cinnabarinus cellobiose dehydrogenase in Pichia pastoris and involvement in saccharification processes

BackgroundCellobiose dehydrogenase (CDH) is an extracellular hemoflavoenzyme produced by lignocellulose-degrading fungi including Pycnoporus cinnabarinus. We investigated the cellulolytic system of P. cinnabarinus, focusing on the involvement of CDH in the deconstruction of lignocellulosic biomass.ResultsFirst, P. cinnabarinus growth conditions were optimized for CDH production. Following growth under cellulolytic conditions, the main components secreted were cellulases, xylanases and CDH. To investigate the contribution of P. cinnabarinus secretome in saccharification processes, the Trichoderma reesei enzymatic cocktail was supplemented with the P. cinnabarinus secretome. A significant enhancement of the degradation of wheat straw was observed with (i) the production of a large amount of gluconic acid, (ii) increased hemicellulose degradation, and (iii) increased overall degradation of the lignocellulosic material. P. cinnabarinus CDH was heterologously expressed in Pichia pastoris to obtain large amounts of pure enzyme. In a bioreactor, the recombinant CDH (rCDH) expression level reached 7800 U/L. rCDH exhibited values of biochemical parameters similar to those of the natural enzyme, and was able to bind cellulose despite the absence of a carbohydrate-binding module (CBM). Following supplementation of purified rCDH to T. reesei enzymatic cocktail, formation of gluconic acid and increased hemicellulose degradation were observed, thus confirming the previous results observed with P. cinnabarinus secretome.ConclusionsWe demonstrate that CDH offers an attractive tool for saccharification process enhancement due to gluconic acid production from raw lignocellulosic material.

Biotransformation of pineapple juice sugars into dietetic derivatives by using a cell free oxidoreductase from Zymomonas mobilis together with commercial invertase

An easy procedure for cell free biotransformation of pineapple juice sugars into dietetic derivatives was accomplished using a commercial invertase and an oxidoreductase from Zymomonas mobilis. First, pineapple juice sucrose was quantitatively converted into glucose and fructose by invertase, thus increasing the concentration of each monosaccharide in the original juice to almost twice. In a second step, glucose–fructose oxidoreductase (GFOR) transformed glucose into gluconolactone, and fructose into the low calorie sweetener sorbitol. The advantage of using GFOR is simultaneous reduction of fructose and oxidation of glucose, allowing the continuous regeneration of the essential coenzyme NADP(H), that is tightly bound to the enzyme. The yield of GFOR catalyzed sugar conversion depends on initial pH and control of pH during the reaction. At optimal conditions (pH control at 6.2) a maximum of 80% (w/v) sugar conversion was obtained. Without pH control, GFOR is inactivated rapidly due to gluconic acid formation. Therefore, conversion yields are relatively low at the natural pH of pineapple juice. The application of this process might be more advantageous on juices of other tropical fruits (papaya, jackfruit, mango) due to their naturally given higher pH.

Electrospun Nanofibrous PLGA/Fullerene-C60 Coated Quartz Crystal Microbalance for Real-Time Gluconic Acid Monitoring

In this paper, a piezoelectric (PZ) sensor based on quartz crystal microbalance (QCM) for the determination of gluconic acid is described. PZ quartz crystals were coated with a 550-700 nm-thick layer of nanoflbers comprising of poly(DL-lactide-co-glycolide) (PLGA) and fullerene-C60 by electrospinning. Glucose oxidase (GOx) was immobilized on coverslips electrospun with nanofibrous PLGA for the interaction of the PZ sensor with gluconic acid, the oxidation product of β-D-glucose. Immobilization of GOx was confirmed by the activity assay and scanning electron microscopy analyses. The interaction of fullerene-C60 with electron-withdrawing gluconic acid molecules was followed by the change in the resonance frequency of the coated crystals in a flow system. The effects of glucose (gluconic acid) concentration, interfering species and temperature on the frequency response of the PLGA/C60-QCM sensor was investigated. The resonance frequency decreased owing to the adsorption of gluconic acid on the sensor. A linear relationship between the frequency response and gluconic acid concentration was observed in the range of 1.4-14.0 mM at room temperature. The optimal temperature of 37°C for gluconic acid formation was also shown by frequencies recorded with the PZ sensor. A good selectivity for gluconic acid, in the presence of some potential interferents, namely urea, ascorbic acid, cysteine, ammonia, Na <sup xmlns:mml="http://www.w3.org/1998/Math/MathML" xmlns:xlink="http://www.w3.org/1999/xlink">+</sup> , K <sup xmlns:mml="http://www.w3.org/1998/Math/MathML" xmlns:xlink="http://www.w3.org/1999/xlink">+</sup> , creatine, acetone, and ethanol was also determined.

Enzymatically synthesized polyaniline layer for extension of linear detection region of amperometric glucose biosensor

In this article a new method for fabrication of enzymatic electrodes suitable for design of amperometric glucose biosensor and/or anode of biofuel cell powered by glucose is presented. Glucose oxidase (GOx) E.C. 1.1.3.4. from Penicillium vitale was immobilized on the carbon rod electrode by cross-linking it with glutaraldehyde (GOx-electrode). Catalytic activity of immobilized GOx was exploited for polymerisation of aniline by taking a high concentration of hydrogen peroxide produced during the catalytic action of immobilized GOx and locally lowered pH due to the formation of gluconic acid; it created optimal conditions for the polymerisation of aniline. The GOx layer was self-encapsulated within formed polyaniline (PANI) matrix (GOx/PANI-electrode). Properties of the GOx/PANI-electrode have been studied and results were compared with GOx-electrode. The results show that the upper detection limit of glucose using GOx-electrode was dramatically changed by the formation of PANI layer. An increase in the upper detection limit, optimal pH region for operation and stability of GOx based electrode modified by PANI was detected when comparing that of an unmodified GOx-electrode.

Capillary electrophoresis for the monitoring of carboxylic acid production by Gluconobacter oxydans

Determination of carboxylic acids in Gluconobacter oxydans fermentations of wheat straw hydrolyzate was carried out. This matrix is of complex composition containing carbohydrates, organic compounds (e.g., amino acids, toxins), and inorganic salts making the analysis challenging even with separation techniques. A method based on capillary electrophoresis with indirect UV detection was developed for the simultaneous quantification of 18 carboxylic acids. The background electrolyte solution of ammonia, 2,3-pyridinedicarboxylic acid, and Ca 2+ and Mg 2+ salts, containing myristyltrimethylammonium hydroxide as a dynamic capillary coating reagent, was validated for the robust and repeatable separation of the carboxylic acids. Intraday relative standard deviations in the optimized method were less than 1.6% for migration times and between 1.0% and 5.9% for peak area. Interday relative standard deviations were less than 5.0% for migration times and between 5.7% and 9.3% for peak area. With 11 nl injected, detection limits for the analytes were between 10 and 43 μmol/l. Detection limits ranged from 0.1 to 0.5 pmol at signal-to-noise ratio of 3. The results demonstrated that wheat straw hydrolyzate was a suitable substrate for G. oxydans with a product yield of 45% for the formation of xylonic acid from xylose and 96% for the formation of gluconic acid from glucose.

Role of Gluconic Acid Production in the Regulation of Biocontrol Traits of Pseudomonas fluorescens CHA0

The rhizobacterium Pseudomonas fluorescens CHA0 promotes the growth of various crop plants and protects them against root diseases caused by pathogenic fungi. The main mechanism of disease suppression by this strain is the production of the antifungal compounds 2,4-diacetylphloroglucinol (DAPG) and pyoluteorin (PLT). Direct plant growth promotion can be achieved through solubilization of inorganic phosphates by the production of organic acids, mainly gluconic acid, which is one of the principal acids produced by Pseudomonas spp. The aim of this study was to elucidate the role of gluconic acid production in CHA0. Therefore, mutants were created with deletions in the genes encoding glucose dehydrogenase (gcd) and gluconate dehydrogenase (gad), required for the conversion of glucose to gluconic acid and gluconic acid to 2-ketogluconate, respectively. These enzymes should be of predominant importance for rhizosphere-colonizing biocontrol bacteria, as major carbon sources provided by plant root exudates are made up of glucose. Our results show that the ability of strain CHA0 to acidify its environment and to solubilize mineral phosphate is strongly dependent on its ability to produce gluconic acid. Moreover, we provide evidence that the formation of gluconic acid by CHA0 completely inhibits the production of PLT and partially inhibits that of DAPG. In the Deltagcd mutant, which does not produce gluconic acid, the enhanced production of antifungal compounds was associated with improved biocontrol activity against take-all disease of wheat, caused by Gaeumannomyces graminis var. tritici. This study provides new evidence for a close association of gluconic acid metabolism with antifungal compound production and biocontrol activity in P. fluorescens CHA0.