Optimization of soy protein isolate, microbial transglutaminase and glucono-δ-lactone in gluten-free rice noodles.

BACKGROUND: Rice flour does not contain gluten and lacks cohesion and extensibility, which is responsible for the poor texture of rice noodles. Different technologies have been used to mitigate this challenge, including hydrothermal treatments of rice flour, direct addition of protein in noodles, use of additives such as hydrocolloids and alginates, and microbial transglutaminase (MTG). Recently, the inclusion of soy protein isolate (SPI), MTG, and glucono-δ-lactone (GDL) in the rice noodles system yielded rice noodles with improved texture and more compact microstructure, hence the need to optimize the addition of SPI, MTG, and GDL to make quality rice noodles. RESULTS: Numerical optimization showed that rice noodles prepared with SPI, 68.32 (g kg-1 of rice flour), MTG, 5.06 (g kg-1 of rice flour) and GDL, 5.0 (g kg-1 of rice flour) gave the best response variables; hardness (53.19 N), springiness (0.76), chewiness (20.28 J), tensile strength (60.35 kPa), and cooking time (5.15 min). The pH, sensory, and microstructure results showed that the optimized rice noodles had a more compact microstructure with fewer hollows, optimum pH for MTG action, and overall sensory panelists also showed the highest preference for the optimized formulation, compared to other samples selected from the numerical optimization and desirability tests. CONCLUSION: Optimization of the levels of SPI, MTG, and GDL yielded quality noodles with improved textural, mechanical, sensory, and microstructural properties. This was partly due to the favourable pH value of the optimized noodles that provided the most suitable conditions for MTG crosslinking and balanced electrostatic interaction of proteins. © 2020 Society of Chemical Industry.

Robust detection of oncometabolic aberrations by 1H-13C heteronuclear single quantum correlation in intact biological specimens.

Magnetic resonance (MR) spectroscopy has potential to non-invasively detect metabolites of diagnostic significance for precision oncology. Yet, many metabolites have similar chemical shifts, yielding highly convoluted 1H spectra of intact biological material and limiting diagnostic utility. Here, we show that hydrogen-carbon heteronuclear single quantum correlation (1H-13C HSQC) offers dramatic improvements in sensitivity compared to one-dimensional (1D) 13C NMR and significant signal deconvolution compared to 1D 1H spectra in intact biological settings. Using a standard NMR spectroscope with a cryoprobe but without specialized signal enhancing features such as magic angle spinning, metabolite extractions or 13C-isotopic enrichment, we obtain well-resolved 2D 1H-13C HSQC spectra in live cancer cells, in ex vivo freshly dissected xenografted tumors and resected primary tumors. This method can identify tumors with specific oncometabolite alterations such as IDH mutations by 2-hydroxyglutarate and PGD-deleted tumors by gluconate. Results suggest potential of 1H-13C HSQC as a non-invasive diagnostic in precision oncology.

Characterization of chemical profiles of pH-sensitive cleavable D-gluconhydroximo-1, 5-lactam hydrolysates by LC-MS: A potential agent for promoting tumor-targeted drug delivery.

Currently, controllable linker cleavage at the target site will facilitate the clinical treatment of cancer. Dual-functional prodrugs in combination of carbohydrate as targeting group and pH-sensitive cleavable linker are desired in clinical development. Here, a qualified structure of N-phenylcarbamate-d-gluconhydroximo-1,5-lactam was employed and proved to be a potential candidate prodrug in the drug design. To proof this concept, the possible mechanism of Beckmann rearrangement and the degraded products were confirmed by HPLC and LC-MS under the acid condition mimic lysosome. Hence, the strategy of d-gluconhydroximo-1,5-lactam as a prodrug carrier fabricated with interested drugs will provide a great potential approach for chemotherapy.

Distinct biotite-weathering activities of Arthrobacter pascens F74 under different contact conditions.

Bacteria play important roles in mineral weathering and soil formation. However, little is known regarding the interactions between biotite and Arthrobacter strains. In this study, the mineral-mineral activities of the Arthrobacter pascens F74 isolated from a weathered rock surface were evaluated for its weathering behavior under direct contact and no contact with biotite. No contact was obtained by using dialysis bags. When directly in contact with biotite, Al and Fe concentrations increased by 9- to 47-fold compared with the controls in the presence of strain F74. Furthermore, strain F74 increased mobilized Al by 106% to 175% and Fe by 29% to 123% under direct contact than under no contact conditions. During biotite dissolution, significantly higher cell numbers and lower pH in the culture medium were observed in the presence of strain F74 under direct contact conditions than under no contact conditions. Significantly higher gluconic acid concentration and glucose dehydrogenase activity were found under direct contact conditions than under no contact and no biotite conditions. Scanning electron microscopy analysis showed cell adhesion on the biotite surface. These results demonstrated that strain F74 behaved differently with respect to biotite-weathering effectiveness and mechanisms under different contact conditions. The results also suggested that direct contact between biotite and strain F74 was important for the production of gluconic acid, cell adhesion on the mineral surface, and the mineral dissolution of the strain.

A simple enzymatic assay for the quantification of C1-specific cellulose oxidation by lytic polysaccharide monooxygenases.

OBJECTIVE: The development of an enzymatic assay for the specific quantification of the C1-oxidation product, i.e. gluconic acid of cellulose active lytic polysaccharide monooxygenases (LPMOs). RESULTS: In combination with a ß-glucosidase, the spectrophotometrical assay can reliably quantify the specific C1- oxidation product of LPMOs acting on cellulose. It is applicable for a pure cellulose model substrate as well as lignocellulosic biomass. The enzymatic assay compares well with the quantification performed by HPAEC-PAD. In addition, we show that simple boiling is not sufficient to inactivate LPMOs and we suggest to apply a metal chelator in addition to boiling or to drastically increase pH for proper inactivation. CONCLUSIONS: We conclude that the versatility of this simple enzymatic assay makes it useful in a wide range of experiments in basic and applied LPMO research and without the need for expensive instrumentation, e.g. HPAEC-PAD.

A new method for biological synthesis of agriculturally relevant nanohydroxyapatite with elucidated effects on soil bacteria.

The study describes a novel and environment friendly route of biosynthesis of nanohydroxyapatite (nHAP). Bacillus licheniformis mediated synthesis of nHAP has been carried out with different phosphate concentrations (2%, 5%, 10% and 20% w/v) of potassium dihydrogen orthophosphate monobasic (K2HPO4). The synthesis is supported by a two-step mechanism - (i) solubilization of P by organic acids extracellularly secreted by the bacterial strain and (ii) gelation of P and Ca. The nHAP particles were characterized using electron microscopy and XRD analysis. Powdered crystalline particles with a size range of 30 ± 5 nm were obtained with shape and size dependent on phosphate concentrations. The particles showed no adverse effect on plant growth-promoting bacteria. Evaluation of nHAP prepared by this route with 2% P source provides scope for a wide range of applications, especially as a nano-fertilizer.

Heart-cutting two-dimensional liquid chromatography combined with isotope ratio mass spectrometry for the determination of stable carbon isotope ratios of gluconic acid in honey.

Liquid chromatography/isotope ratio mass spectrometry (LC/IRMS) is used to analyze various types of samples, including foodstuffs, to determine their authenticity and trace their origin on the basis of their stable carbon isotope ratios (δ13C). However, multicomponent samples are difficult to analyze. For example, determining the δ13C values of the organic acids in honey is complicated by the presence of large amounts of carbohydrates. Herein, we present a heart-cutting two-dimensional LC/IRMS method for analysis of honey samples. In this method, the organic acids in the samples were first separated from the carbohydrates by a size-exclusion column, and then the organic acids were separated from each other by a reverse-phase column connected to the first column via a switching valve. By means of this method, the δ13C values for three organic acids in high-carbohydrate-content simulated honey samples could be determined with high accuracy and precision (≤0.3‰ and ≤0.1‰, respectively). In addition, the gluconic acid δ13C values for 25 honey samples were determined with high precision and found to range from -31.7 to -28.5‰ (mean: -30.0 ±â€¯0.7‰). These values shed some light on the mechanism of gluconic acid production. Taken together, our results suggest that this two-dimensional LC method has the potential to be more effective than one-dimensional LC for use in isotopic research.

Modified semi-continuous fermentation for resuscitating nongrowing cells during high-temperature gluconic acid production by Acetobacter senegalensis.

AIMS: The formation of metabolically inactive and nongrowing cells is an inevitable by-product of intensive fermentation. This study investigated whether co-feeding can be used to resuscitate nongrowing Acetobacter senegalensis cells to enable them to produce gluconic acid in successive fermentation runs at 38°C. METHODS AND RESULTS: In the first fermentation cycle, 75 g l-1 of glucose were converted to gluconic acid. Subsequently, however, stationary-phase cells were unable to initiate a new fermentation cycle. The majority of stationary-phase cells (97%) were nonculturable on glucose at 38°C. In addition, 54 and 41% of cells contained non-active cellular dehydrogenases and a compromised cell envelope respectively. Co-feeding stationary-phase cells with a mixture of ethanol, glucose and acetic acid for 7 h enabled these cells to grow on 75 g l-1 of glucose and produce gluconic acid. Additionally, 74% of cells contained active forms of cellular dehydrogenases after 7 h of co-feeding. However, co-feeding did not improve cell envelope integrity. Quantification of cellular NAD content showed that stationary-phase cells contained moderately reduced levels of total NAD (NADt) as compared with exponential-phase cells. Interestingly, the analysis of stationary-phase cells showed that co-feeding resulted in higher levels of NADt and NADH, suggesting that the regeneration of NADH is one of the limiting factors of glucose consumption. Expression of catalase and superoxide dismutase was increased in stationary-phase cells, but analysis of protein carbonylation and lipid peroxidation did not confirm an extensive oxidative stress. CONCLUSIONS: Co-feeding with favourable nutrients may enable resuscitation of cells and utilization of less-favourable carbon sources in successive cycles. SIGNIFICANCE AND IMPACT OF THE STUDY: This study proposed a unique method for resuscitation of nongrowing cells during high-temperature fermentation. By applying this method, cells can be used for consecutive fermentation cycles.

Stable carbon isotope ratios for organic acids in commercial honey samples.

Stable carbon isotope ratios (δ13C) for glucose, fructose, disaccharides, trisaccharides, and organic acids in 116 commercial honey samples were measured by LC/IRMS. On the basis of EA/IRMS and LC/IRMS authenticity criteria, 39 of the samples were judged to have been adulterated. The δ13C values for organic acids from pure honey, reported here for the first time, ranged from -33.6 to -26.5‰. The mean Δδ13C (glucose-organic acids) value was +3.7 ±â€¯0.9‰. Glucose and organic acid δ13C values were strongly correlated (R = 0.71, P < 0.001). Gluconic acid, the predominant organic acid in honey, has been reported to be produced via decomposition of glucose by bee glucose-oxidase and certain Gluconobacter spp. This fact was confirmed by isotope analysis.

Different temperatures select distinctive acetic acid bacteria species and promotes organic acids production during Kombucha tea fermentation.

Kombucha is a traditional beverage produced by tea fermentation, carried out by a symbiotic consortium of bacteria and yeasts. Acetic Acid Bacteria (AAB) usually dominate the bacterial community of Kombucha, driving the fermentative process. The consumption of this beverage was often associated to beneficial effects for the health, due to its antioxidant and detoxifying properties. We characterized bacterial populations of Kombucha tea fermented at 20 or 30 °C by using culture-dependent and -independent methods and monitored the concentration of gluconic and glucuronic acids, as well as of total polyphenols. We found significant differences in the microbiota at the two temperatures. Moreover, different species of Gluconacetobacter were selected, leading to a differential abundance of gluconic and glucuronic acids.

Determination of calcium and zinc in gluconates oral solution and blood samples by liquid cathode glow discharge-atomic emission spectrometry.

A novel flowing liquid cathode glow discharge (LCGD) was developed as an excitation source of the atomic emission spectrometry (AES) for the determination of Ca and Zn in digested calcium and zinc gluconates oral solution and blood samples, in which the glow discharge is produced between the electrolyte (as cathode) overflowing from a quartz capillary and the needle-like Pt anode. The electron temperature and electron density of LCGD were calculated at different discharge voltages. The discharge stability and parameters affecting the LCGD were investigated in detail. In addition, the measured results of real samples using LCGD-AES were verified by ICP-AES. The results showed that the optimized analytical conditions are pH = 1 HNO3 as supporting electrolyte, 4.5mLmin-1 solution flow rate. The power consumption of LCGD is 43.5-66.0W. The R2 and the RSD ranged from 630 to 680V are 0.9942-0.9995 and 0.49%-2.43%, respectively. The limits of detections (LODs) for Zn and Ca are 0.014-0.033 and 0.011-0.097mgL-1, respectively, which are in good agreement with the closed-type electrolyte cathode atmospheric glow discharge (ELCAD). The obtained results of Ca and Zn in real samples by LCGD-AES are basically consistent with the ICP-AES and reference value. The results suggested that LCGD-AES can provide an alternative analytical method for the detection of metal elements in biological and medical samples.

The Molting Biomarker Molecule Exists as 2-Acetamido-2-deoxy-gluconic Acid in Urine of Blue Crabs and Helmet Crabs.

N-Acetyl-d-glucosamino-1,5-lactone 1 has been reported as a candidate component of the sex pheromone mixture of female blue crabs, Callinectes sapidus, since it is present in the urine of reproductive females and males detect it. Theoretically, 1 can convert to a 1,4-lactone isomer 2 or to the corresponding carboxylic acid, 2-acetamido-2-deoxygluconic acid 3 by hydrolysis in aqueous solution. In this study, we examined the biologically relevant state of equilibrium mixture of 1, 2, and 3 in crab urine using ESI-MS and NMR analyses. The ESI-MS analysis showed that the dominant form of solubilized synthetic 1 is lactone 1 and/or 2, immediately after solubilization in deuterated water, seawater, and phosphate buffer and gradually changing to carboxylic acid 3 which becomes most predominant in phosphate buffer. The NMR analysis showed that synthetic 1 converts to other forms in deuterated water and seawater, and reaches an equilibrium mixture of at least three forms within 24 h. In contrast, 1 converts to a single state of another form in deuterated water with 35 mm phosphate buffer pH 7.6 within 24 h, which is identical to the state in urine with or without phosphate buffer. Thus, we conclude that the molting biomarker sensed by male crabs is 3.

The use of glucose oxidase and catalase for the enzymatic reduction of the potential ethanol content in wine.

Due to the increase of sugar levels in wine grapes as one of the impacts of climate change, alcohol reduction in wines becomes a major focus of interest. This study combines the use of glucose oxidase and catalase activities with the aim of rapid conversion of glucose into non-fermentable gluconic acid. The H2O2 hydrolysing activity of purified catalase is necessary in order to stabilize glucose oxidase activity. After establishing the adequate enzyme ratio, the procedure was applied in large-scale trials (16L- and 220L-scale) of which one was conducted in a winery under industrial wine making conditions. Both enzyme activity and wine flavour were clearly influenced by the obligatory aeration in the different trials. With the enzyme treatment an alcohol reduction of 2%vol. was achieved after 30h of aeration. However the enzyme treated wines were significantly more acidic and less typical.

Comparison of copper and zinc in vitro bioaccessibility from cyanobacteria rich in proteins and a synthetic supplement containing gluconate complexes: LC-MS mapping of bioaccessible copper complexes.

An analytical procedure was proposed to estimate bioaccessibility of copper and zinc in Spirulina Pacifica tablets with respect to that of copper and zinc in gluconate complexes. Spirulina is the common name for diet supplements produced primarily from two species of cyanobacteria, namely Arthrospira platensis and Arthrospira maxima. Spirulina tablets are an excellent source of proteins, vitamins and minerals. To obtain information about the bioavailability of these elements, an in vitro bioaccessibility test was performed by application of a two-step protocol which simulated the gastric (pepsin) and intestinal (pancreatin) digestion. The species obtained were investigated by size exclusion chromatography on a chromatograph coupled to a mass spectrometer with inductively coupled plasma (SEC-ICP-MS) and an on-capillary liquid chromatograph coupled to an electrospray mass spectrometer (µ-HPLC-ESI-MS). Both copper and zinc were found to be highly bioaccessible in Spirulina tablets (90-111%) and those containing gluconate complexes (103% for Cu and 62% for Zn). In Spirulina tablets, copper was found to form two types of complex: (1) polar ones with glycine and aspartic acid and (2) more hydrophobic ones containing amino acids with cyclic hydrocarbons (phenylalanine, histidine, proline and tyrosine). Zinc and copper were also proved to form complexes during the digestion process with products of pepsin digestion, but the stability of these complexes is lower than that of the complexes formed in Spirulina. The results proving the involvement of proteins in the enhancement of copper and zinc bioaccessibility will be useful for the design of new copper and zinc supplements.

Development of a rapid and direct method for the determination of organic acids in peach fruit using LC-ESI-MS.

An accurate, simple and rapid liquid chromatography mass spectrometry method for the determination of organic acids in peach fruit has been developed. Direct injection and sample clean-up with a mixed-mode sorbent was compared. The best results for the determination of gluconic, oxalic, malic, citric and fumaric acids were obtained with only a simple dilution and filtration step, and nylon filters should be avoided since some organic acids are retained by them. It is the first time that gluconic acid has been determined in peach fruit. Different parameters involved in the separation and detection process have been optimized. Since matrix effects were observed in the peach commodity, organic acids were quantified by the standard addition method. All validation parameters of the method were found acceptable of all organic acids. Finally, the method was successfully applied to the analysis of samples of peach from two cultivars.

Spectroscopic approaches to resolving ambiguities of hyper-polarized NMR signals from different reaction cascades.

The influx of exogenous substrates into cellular reaction cascades on the seconds time scale is directly observable by NMR spectroscopy when using nuclear spin polarization enhancement. Conventional NMR assignment spectra for the identification of reaction intermediates are not applicable in these experiments due to the non-equilibrium nature of the nuclear spin polarization enhancement. We show that ambiguities in the intracellular identification of transient reaction intermediates can be resolved by experimental schemes using site-specific isotope labelling, optimised referencing and response to external perturbations.

Terahertz time-domain spectroscopy and quantitative analysis of metal gluconates.

A series of metal gluconates (Na(+), K(+), Mg(2+), Ca(2+), Fe(2+), Cu(2+), and Zn(2+)) were investigated by terahertz (THz) time-domain spectroscopy. The absorption coefficients and refractive indices of the samples were obtained in the frequency range of 0.5-2.6 THz. The gluconates showed distinct THz characteristic fingerprints, and the dissimilarities reflect their different structures, hydrogen-bond networks, and molecular interactions. In addition, some common features were observed among these gluconates, and the similarities probably come from the similar carbohydrate anion group. The X-ray powder diffraction measurements of these metal gluconates were performed, and the copper(II) gluconate was found to be amorphous, corresponding to the monotonic increase feature in the THz absorption spectrum. The results suggest that THz spectroscopy is sensitive to molecular structure and physical form. Binary and ternary mixtures of different gluconates were quantitatively analyzed based on the Beer-Lambert law. A chemical map of a tablet containing calcium D-gluconate monohydrate and α-lactose in the polyethylene host was obtained by THz imaging. The study shows that THz technology is a useful tool in pharmaceutical research and quality control applications.

A doped polyaniline modified electrode amperometric biosensor for gluconic acid determination in grapes.

In winemaking gluconic acid is an important marker for quantitative evaluation of grape infection by Botrytis cinerea. A screen-printed amperometric bienzymatic sensor for the determination of gluconic acid based on gluconate kinase (GK) and 6-phospho-D-gluconate dehydrogenase (6PGDH) coimmobilized onto polyaniline/poly (2-acrylamido-2-methyl-1-propanesulfonic acid; PANI-PAAMPSA) is reported in this study. The conductive polymer electrodeposed on the working electrode surface allowed the detection of NADH at low potential (0.1 V) with a linear range from 4 × 10(-3) to 1 mM (R2 = 0.99) and a sensitivity of 419.44 nA∙mM(-1). The bienzymatic sensor has been optimized with regard to GK/6PGDH enzymatic unit ratio and ATP/NADP+ molar ratio which resulted equal to 0.33 and 1.2, respectively. Under these conditions a sensitivity of 255.2 nA∙mM(-1), a limit of detection of 5 µM and a Relative Standard Deviation (RSD) of 4.2% (n = 5) have been observed. Finally, the biosensor has been applied for gluconic acid measurements in must grape samples and the matrix effect has been taken into consideration. The results have been compared with those obtained on the same samples with a commercial kit based on a spectrophotometric enzyme assay and were in good agreement, showing the capability of the bienzymatic PANI-PAAMPSA biosensor for gluconic acid measurements and thus for the evaluation of Botrytis cinerea infection in grapes.

Effects of binders combined with glucono-δ-lactone on the quality characteristics of pressure-induced cold-set restructured pork.

This study investigated the effects of binders and glucono-δ-lactone (GdL) on characteristics of pressure-induced (450MPA for 3min) cold-set restructured pork. Isolated soy protein (SP), wheat flour (WF), and κ-carrageenan (CG) were adopted as binders. The addition of binders improved water-binding properties of restructured pork, and the binders diminished the decrease in water binding properties caused by GdL-induced acidification. Pressure-induced restructured pork prepared with binders showed less harder and more cohesive texture than those of the thermal-treated control (TC). The results indicate that pressure-induced cold-set meat restructuring could be achieved when binders and GdL were used in the formulation.

A gold@silica core-shell nanoparticle-based surface-enhanced Raman scattering biosensor for label-free glucose detection.

The gold nanostar@silica core-shell nanoparticles conjugated with glucose oxidase (GOx) enzyme molecules have been developed as the surface-enhanced Raman scattering (SERS) biosensor for label-free detection of glucose. The surface-immobilized GOx enzyme catalyzes the oxidation of glucose, producing hydrogen peroxide. Under laser excitation, the produced H2O2 molecules near the Au nanostar@silica nanoparticles generate a strong SERS signal, which is used to measure the glucose concentration. The SERS signal of nanostar@silica∼GOx nanoparticle-based sensing assay shows the dynamic response to the glucose concentration range from 25 µM to 25 mM in the aqueous solution with the limit of detection of 16 µM. The sensing assay does not show any interference when glucose co-exists with both ascorbic acid and uric acid. The sensor can be applied to a saliva sample.