Enhancing metformin-induced tumor metabolism destruction by glucose oxidase for triple-combination therapy

Despite decades of laboratory and clinical trials,breast cancer remains the main cause of cancer-related disease burden in women.Considering the metabolism destruction effect of metformin(Met)and cancer cell starvation induced by glucose oxidase(GOx),after their efficient delivery to tumor sites,GOx and Met may consume a large amount of glucose and produce sufficient hydrogen peroxide in situ.Herein,a pH-responsive epigallocatechin gallate(EGCG)-conjugated low-molecular-weight chitosan(LC-EGCG,LE)nanoparticle(Met–GOx/Fe@LE NPs)was constructed.The coordination between iron ions(Fe3+)and EGCG in this nanoplatform can enhance the efficacy of chemodynamic therapy via the Fenton reaction.Met–GOx/Fe@LE NPs allow GOx to retain its enzymatic activity while simultaneously improving its stability.Moreover,this pH-responsive nanoplatform presents controllable drug release behavior.An in vivo biodistribution study showed that the intracranial accumulation of GOx delivered by this nanoplatform was 3.6-fold higher than that of the free drug.The in vivo anticancer results indicated that this metabolism destruction/starvation/chemodynamic triple-combination therapy could induce increased apoptosis/death of tumor cells and reduce their proliferation.This triple-combination therapy approach is promising for efficient and targeted cancer treatment.

Inhibitory activities of microalgal fucoxanthin againstα-amylase,α-glucosidase, and glucose oxidase in 3T3-L1cells linked to type 2 diabetes

Postprandial hyperglycemia is an early indication of type 2 diabetes and the target of many anti-diabetic and anti-obesity studies.α-Glucosidase and α-amylase are the crucial factors in regulating starch digestion and glucose absorption,making them key targets for many studies to treat postprandial hyperglycemia.We studied the inhibitory activities of microalgal fucoxanthin against rat-intestinalα-glucosidase and pancreaticα-amylase along with the antidiabetic eff ect to induce diff erentiation in 3T3-L1 pre-adipocytes using Oil Red-O staining.Fucoxanthin displayed strong hindrance activities towardα-amylase in a concentration-dependent manner,with an IC50 value of 0.68mmol/L,whereas weak inhibitory activity against α-glucosidase,with an IC 50 value of 4.75 mmol/L.Fucoxanthin also considerably elevated glucose oxidase activity in 3T3-L1 cells by 31.3%at 5μmol/L.During adipocyte differentiation,fucoxanthin showed lipid accumulation in 3T3-L1 cells with no cytotoxicity up to 20μmol/L.However,fucoxanthin had no inhibitory activity on glucose-6-phosphate dehydrogenase.These results suggest that fucoxanthin might be useful for the prevention of obesity or diabetes by inhibiting carbohydrate-hydrolyzing enzymes and lipid accumulation and be utilized as an ingredient for a functional food or dietary supplement.

Changes in the gut microbiota mediate the differential regulatory effects of two glucose oxidases produced by Aspergillus niger and Penicillium amagasakiense on the meat quality and growth performance of broilers

Background: Glucose oxidase(GOD), an aerobic dehydrogenase, has been used as an antibiotic substitute in feed.A study was conducted to evaluate the differential effects of 2 different GODs fermented by Aspergillus niger or Penicillium amagasakiense on caecal microbiota and to further illuminate the potential roles of changes in the gut microbiota in regulating the growth performance and meat quality of broiler chickens.Results: A total of 420 one-day-old healthy Arbor Acres broilers were randomly assigned to 4 treatments: the control group,the antibiotic growth promoter(AGP) supplementation group, and the GOD-A and GOD-P(GODs produced by A. niger and P. amagasakiense, respectively) groups. As a result, supplementation with GOD produced by P. amagasakiense could significantly improve the average daily weight gain and average daily feed intake of broilers before 21 days of age by significantly increasing the enzymatic activities of jejunal amylase and those of ileal amylase, chymotrypsin, and lipase in21-day-old broilers and could increase the enzymatic activities of duodenal amylase, jejunal amylase and lipase, and ileal chymotrypsin and lipase in 42-day-old broilers. Meanwhile, compared with AGP treatment, supplementation with GOD produced by P. amagasakiense significantly decreased the L value of 21-day-old broilers and the Δp H and L* value of 42-day-old broilers, while supplementation with GOD produced by A. niger significantly increased the p H24 hvalue of 21-day-old and 42-day-old broilers by reducing plasma malondialdehyde content. By using 16 S r RNA sequencing, we found that the beneficial bacteria and microbiota in broilers were not disturbed but were improved by GOD supplementation compared with ADP treatment, including the genera Eubacterium and Christensenel a and the species uncultured_Eubacterium_sp,Clostridium_asparagiforme, and uncultured_Christensenel a_sp, which were positively related to the improved intestinal digestive enzymatic activities, growth performance, and meat quality of broilers.Conclusion: The altered gut microbiota induced by supplementation with glucose oxidase produced by P. amagasakiense mediate better regulatory effects on the meat quality and growth performance of broilers than that induced by supplementation with glucose oxidase produced by A. niger.

Predominating stable adsorption and direct electrochemistry of glucose oxidase on carbon nanotubes by oxygen-containing groups

Stable adsorption and direct electrochemistry of glucose oxidase （COx） occurred on nitric acid （HNO3）-treated multi-walled carbon nanotubcs （MWNTs） instead of as-received MWNTs, demonstrating the critical roles of oxygen-containing groups in stable adsorption and direct electrochemistry of GOx on carbon nanotubcs （CNTs）.

New amperometric glucose biosensor by entrapping glucose oxidase into chitosan/nanoporous ZrO_2/multiwalled carbon nanotubes nanocomposite film

A new nanocomposite material for construction of glucose biosensor was prepared. The biosensor was formed by entrapping glucose oxidase(Gox) into chitosan/nanoporous ZrO2/multiwalled carbon nanotubes nanocomposite film. In this biosensing thin film, the multiwalled carbon nanotubes can effectively catalyze hydrogen peroxide and nanoporous ZrO2 can enhance the stability of the immobilized enzyme. The resulting biosensor provides a very effective matrix for the immobilization of glucose oxidase and exhibits a wide linear response range from 8 μmol/L to 3 mmol/L with a correlation coefficient of 0.994 for the detection of glucose. And the response time and detection limit of the biosensor are determined to be 6 s and 3.5 μmol/L, respectively. Another attractive characteristic is that the biosensor is inexpensive, stable and reliable.

Polyaniline-graphite composite film glucose oxidase electrode

A novel polyaniline-graphite composite film glucose oxidase （PGCF GOD） electrode was developed. The PGCF was synthesized by cyclic voitammetry method in 0.5 mol/L H2SO4 solution containing 1 g/L graphite powder and 0.2 mol/L aniline. The PGCF GOD electrode was prepared by doping GOD into the composite film. The morphology of the PGCF and the response property of the PGCF GOD electrode were investigated by scanning electron microscopy and electrochemical measurement, respectively. The results show that the PGCF has a porous and netty structure and the PGCF GOD electrode has excellent response property such as high sensitivity and short response time. Influences of pH value, temperature, glucose concentration and potential on the response current of the electrode were also discussed. The sensor has a maximum steady-state current density of 357.17μA/cm2 and an apparent Michaelis-Menten constant of 16.57 mmol/L. The maximum current response of the enzyme electrode occurs under the condition ofpH 5.5, 0.8 V and 65℃.

Amperometric Glucose Biosensor Based on Integration of Glucose Oxidase with Palladium Nanoparticles/Reduced Graphene Oxide Nanocomposite

We report on a new type of amperometric glucose biosensor that was made by integration of glucose oxidase (GOD) with palladium nanoparticles/reduce graphene oxide (Pd/RGO) nanocomposite. The Pd/RGO was prepared by a one-step reduction method in which the palladium nanoparticles and the reduced graphene oxide (RGO) were simultaneously accomplished from the reduction of dispersed solution of PdCl2 and graphite oxide (GO) with hydrazine. The asprepared nanocomposite exhibits favorable electrocatalytic activities towards the oxidation of H2O2, which makes it a good platform for the construction of the glucose biosensor. The analytical performance of the glucose biosensor is fully evaluated. It shows good analytical properties in terms of a short response time (3 s), high sensitivity (14.1 μA/mM), and low detection limit (0.034 mM). In addition, the effects of pH value, applied potential, electroactive interference and the stability of the biosensor were discussed as well.

Determination of trace mercury in compost extract by inhibition based glucose oxidase biosensor

A novel inhibition based biosensor of glucose oxidase(GOx) for environmental mercury detection was developed. An electropolymerized aniline membrane was prepared on a platinum electrode containing ferrocene as electron transfer mediator, on which GOx was cross-linked by glutaraldehyde. The response of the sensor was based on the current reduction in the electrochemical system by inhibition of mercury against GOx electrode. The detection limit of the inhibition-based sensor for mercury is 0.49 μg/L, and the linear response ranges are 0.49-783.21 μg/L and 783.21 μg/L-25.55 mg/L. The GOx membrane can be completely reactivated after inhibition, and remains 70% of the activity in more than one month. The sensor was used for mercury determination in compost extract with good results.

A Chemiluminescence Optical Fiber Glucose Biosensor Based on Co-immobilizing Glucose Oxidase and Horseradish Peroxidase in a Sol-gel Film

An optical fiber bienzyme sensor based on the luminol chemiluminescent reaction was developed and demonstrated to be sensitive to glucose. Glucose oxidase(GOD) and horseradish peroxidase(HRP) were co-immobilized by microencapsulation in a sol-gel film derived from tetraethyl orthosilicate(TEOS). The calibration plots for glucose were established by the optical fiber glucose sensor fabricated by attaching the bienzyme silica gel onto the glass window of the fiber bundle. The linear range was 0 2-2 mmol/L and the detection limit was approximately 0 12 mmol/L. The relative standard deviation was 5.3% ( n =6). The proposed biosensor was applied to glucose assay in ofloxacin injection successfully.

Immobilization of Glucose Oxidase on Cellulose/Cellulose Acetate Membrane and its Detection by Scanning Electrochemical Microscope (SECM)

s: Cellulose/cellulose acetate membranes were prepared and functionalized by introducing amino group on it, and then immobilized the glucose oxidase (Gox) on the functionalizd membrane. SECM was applied for the detection of enzyme activity immobilized on the membrane. Immobilized biomolecules on such membranes was combined with analysis apparatus and can be used in bioassays.

Direct Electron Communication Between Glucose Oxidase and Carbon Electrode Covered with Polypyrrole

Glucose oxidase can be effectively adsorbed onto the polypyrrole (PPy) thin film electrochemically formed on an anodized galssy carbon electrode (GCEa ). Direct electron communication between the redox of GOD and the modified electrode was successfully achieved, which was detected using cyclic voltammetry. GOD entrapped in PPy film still remained its biological activity and could catalyze the oxidation of glucose. As a third generation biosensor, GOD-PPy/GCEa responded linearly up to 20 mM glucose with a wider linear concentration range.

IMMOBILIZATION OF GLUCOSE OXIDASE AND CELLULASE BY CHITOSAN— POLYACRYLIC ACID COMPLEX

This study is concerned with chitosan-polyacrylic acid complex as a carrier to immobilize glucose oxidase (GOD)and cellulase. The optimum emperature of the immobilized GOD (IG) was determined to be 60℃ which is higher than that of the native GOD about 40℃. The optimum temperature of the immobilized cellulase (IC) was determined to be about 30℃ higher than that of native cellulase. Both of the optimum pH of IG and IC shifted one pH unit to acid. Immobilized enzyme may be used in more wide pH range. Their storage life are much longer compared with their native states. Both of them can be reused at least 12 times.

Direct Electron Transfer Reactions of Glucose Oxidase and D-Amino Acid Oxidase at a Glassy Carbon Electrode in Organic Media

Cyclic voltammetry is employed to demonstrate feasibility of direct electron transfer of glucose oxidase and D amino acid oxidase at a glassy carbon electrode in organic media. The reversible slight conformational change of glucose oxidase is observed by changing 0.1 mol/L phosphate buffer to acetonitrile containing 10% v/v of water and 0.05 mol/L tetrabutyalammonium perchlorate, and vice versa.

STUDIES ON IMMOBILIZED GLUCOSE OXIDASE BY DIETHYLAMINOETHYL CELLULOSE COMPLEXES

The properties of immobilized glucose oxidase (GOD) by the complexes of diethylaminoethyl cellulose(DEAEC) with different polymers, such as polymethylacrylic acid (PMAA), polyacrylic acid (PAA), polystyrene sulfonic acid (PSSA), polyvinylaleohol (PVA), polyethylene oxide (PEO) and styrene-maleic acid copolymer (PSMA) were investigated. The activity of immobilized GOD was obviously influenced by the component of the DEAEC complexes. The relative activity of the immobilized GOD reached to maximum and over 90% of the native GOD. when the DEAEC-PMAA DEAEC-PAA complexes were used as a carrier with the molar ratio of DEAEC and polyacid of about one. Michaelis constants (Km) of the immobilized enzymes of DEAEC-GOD-PMAA and DEAEC-GOD-PAA were determined to be 1.25 and 1.00, respectively. Moreover, the immobilized GOD has a good storage stability and cyclic life.

Effects of Various Factors on Production of Glucose Oxidase in Resting Cell Culture System

n order to demonstrate the effects of various factors on the biosynthesis of glucose oxidase(GOD) without considering the growth status, a resting cell culture system was established. By using this system, the sequence of the effect of carbon sources on the biosynthesis of (GOD) could be revealed as follows: mannose>glycerol>glucose>cellulose>lactose>sucrose>galactose>soluble starch>maltose>fructose. The enzyme activity was promoted by adding tiny amount of αmethylDglucoside. When 0.1% NaNO3 was replaced by 0.05% of amino acid, Tyr, Leu, Orn and Glu were shown to have positive effect, while Phe, Ile, Lys, His and Arg have negative effect on the biosynthesis of GOD, and no net growth was observed in the above experiments. The biosynthesis of GOD was highly susceptible to the concentration of CaCl2 added, 0.01% of CaCl2 was in favor of the enzyme synthesis, whereas 0.02% was unfavorable. The starting time for GOD biosynthesis was estimated through addition of actinomycin D at various time of incubation, it turned out to be between 11～13h after inoculation.

Membrane Microfiltration Fermentation of Glucose Oxidase with Cell Recycling

Membrane microfiltration fermentation (MMF) with cell recycling was successfully applied to the production of glucose oxidase (GOD). A plate microfiltration module was found suitable for such purpose. By feeding whole medium in MMF, the productivity of GOD was much higher than that by feeding glucose alone. With increasing dilution rate the enzyme productivity increased and average enzyme activity decreased. The enzyme productivity of MMF under D = 0.12h-1 and 0.20h-1 were 3871 and 3945U·h-1 respectively, which was about 3 times as that of batch fermentation (BF) and the average enzyme activity was still as high as STU·mL-1 under D = 0.12h-1. The relative efficiency of MMF applied to low yield strain was higher than that applied to high yield strain.

Mathematical Modeling of the Amperometric Response to Glucose of Glucose Oxidase Films Deposited by AC-Electrophoresis

Previous work illustrated that glucose oxidase (GOx) could be deposited on conducting substrates using asymmetrical alternating current electrophoretic deposition (AC-EPD) to form thick enzyme layers suitable for the manufacturing of highly active biosensors. Here, we modeled the amperometric response of GOx layers to glucose as a function of the thickness of the enzyme layer. The model is based on reaction-diffusion equations with irreversible first-order catalytic reactions. The numerical results displayed qualitative and reasonable quantitative agreement with the experimental data obtained for oxidation currents due to glucose, which increase with the enzyme thickness.

Simple Method for Preparing Glucose Biosensor Based on Glucose Oxidase in Nanocomposite Material of Single-Wall Carbon Nanotubes/Ionic Liquid

Based on electric conductivity and wide potential window of ionic liquid (IL) and electric property of single-wall car- bon nanotubes (SWCNTs), composite material of IL-SWCNTs was prepared, glucose sensor was built with this mate-rial for immobilizing glucose oxidase (GOx). It showed good response, sensitivity and stability for long time for glu-cose detection. Linear range for the detection of glucose was from 0.5 × 10–6 M to 12 × 10–6 M while detection limit was 6.26 × 10–8 M (S/N = 3).

Heterologous expression of the glucose oxidase gene in Trichoderma atroviride leads enhanced ability to attack phytopathogenic fungi and induction of plant systemic disease resistance

A transgenic strain of Trichoderma atroviride that expresses the Aspergillus niger glucose oxidase gene goxA under a homologous pathogen-inducible promoter (nag1) has been constructed, with the aim of increasing the ability of this biocontrol agent (BCA) to attack phytopathogenic fungi and enhance plant systemic disease resistance. The sporulation and growth rate of the transgenic progenies were similar to the wild-type strain P1. goxA expression occurred immediately after contact with the plant pathogen, and the glucose oxidase formed was secreted extracellularly. The transformed strain SJ3 4, containing 12-14 copies of the transgene, produced significantly less N-acetyl-glucosaminidase and endochitinase then wild type. However, the ability of its culture filtrate to inhibit the germination of Botrytis cinerea spores was increased by about 3-fold. In comparison to P1, the transgenic strain more quickly overgrew and lysed in vitro the pathogens Rhizoctonia solani and Pythium ultimum. In assays in vivo SJ3 4 showed a highly improved biocontrol ability in soil heavily infested with those pathogens, where the wild type was unable to protect the plant and allow seeds to germinate. The Trichoderma-gox was able to induce a much higher level of systemic resistance against the foliar pathogen B. cinerea, as compared to the parent strain. This work demonstrate that i) heterologous genes driven by pathogen-inducible promoters can improve the biocontrol and Induced Systemic Resistance properties of fungal BCAs such as Trichoderma spp., and ii) these microbes can be used as vectors to provide the plant with useful molecules able, for instance, to increase pathogen

饲粮中复合添加马齿苋、柠檬酸和葡萄糖氧化酶对肉兔生长性能、屠宰性能以及免疫和抗氧化功能的影响

本试验旨在研究饲粮中复合添加马齿苋、柠檬酸和葡萄糖氧化酶对肉兔生长性能、屠宰性能以及免疫和抗氧化功能的影响。试验选取200只健康的35日龄断奶伊拉商品肉兔(公母各占1/2),按照体重和性别随机分为4组,每组50个重复,每个重复1只兔。对照组饲喂基础饲粮,试验1组饲喂在基础饲粮中添加3%马齿苋替换等量麸皮的饲粮,试验2组和试验3组分别饲喂在试验1组饲粮基础上添加1%柠檬酸和1%柠檬酸+200 mg/kg葡萄糖氧化酶的饲粮。预试期7 d,正试期35 d。结果表明:1)与对照组相比,试验2组和试验3组肉兔终末体重显著提高(P<0.05),死亡率显著降低(P<0.05),腹泻频率有降低趋势(0.05<P<0.10);试验2组肉兔平均日增重和平均日采食量显著提高(P<0.05);试验3组肉兔料重比和平均日采食量显著低于试验1组和试验2组(P<0.05)。2)与对照组相比,3个试验组肉兔半净膛屠宰率显著提高(P<0.05),试验1组和试验2组肉兔全净膛屠宰率显著提高(P<0.05)。3)与对照组相比,3个试验组肉兔血清免疫球蛋白G含量显著提高(P<0.05),试验1组和试验2组肉兔血清白细胞介素-1β含量显著降低(P<0.05),试验2组和试验3组肉兔血清白细胞介素-6(IL⁃6)含量显著降低(P<0.05);试验3组肉兔胸腺指数显著高于试验1组(P<0.05),血清IL⁃6含量显著低于试验1组(P<0.05)。4)各组间肉兔肝脏抗氧化指标均无显著差异(P>0.05)。综上所述,饲粮中复合添加马齿苋、柠檬酸和葡萄糖氧化酶能够提高肉兔生长性能和屠宰性能,提高机体免疫力,降低死亡率和腹泻频率。