Method For Determination Of Glucose Research Articles

A novel method for glucose determination based on electrochemical impedance spectroscopy using glucose oxidase self-assembled biosensor

A method is developed for quantitative determination of glucose using electrochemical impedance spectroscopy (EIS). The method is based on immobilized glucose oxidase (GOx) on the topside of gold mercaptopropionic acid self-assembled monolayers (Au-MPA-GOx SAMs) electrode and mediation of electron transfer by parabenzoquinone (PBQ). The PBQ is reduced to hydroquinone (H 2Q), which in turn is oxidized at Au electrode in diffusion layer. An increase in the glucose concentration results in an increase in the diffusion current density of the H 2Q oxidation, which corresponds to a decrease in the faradaic charge transfer resistance ( R ct) obtained from the EIS measurements. Glucose is quantified from linear variation of the sensor response (1/ R ct) as a function of glucose concentration in solution. The method is straightforward and nondestructive. The dynamic range for determination of glucose is extended to more than two orders of magnitude. A detection limit of 15.6 μM with a sensitivity of 9.66 × 10 − 7 Ω − 1 mM − 1 is obtained.

Rapid enzymatic chemiluminescent assay of glucose by means of a hybrid flow-injection/sequential-injection method

This work reports a hybrid flow-injection analysis (FIA)/sequential-injection analysis (SIA) method for the rapid enzymatic assay of glucose with soluble glucose oxidase (GOD). The method relies on the sequential injection of segments of the sample and of a solution of enzyme by means of a multi-port selection valve in a flowing water stream. As the two zones are swept downstream, they overlap and merge so that the glucose in the sample is enzymatically oxidised. The generated hydrogen peroxide is merged with an alkaline luminol solution and the chemiluminescence (CL) intensity is monitored and related to the glucose concentration in the sample. The linear range of the method for glucose determination is 0.01–1 mmol L −1, the relative standard deviation is 3.9% at the 0.08 mmol L −1 level ( n = 8), the limit of detection at the 2 σ level is 4 μmol L −1 glucose and the injection rate is 80 h −1. The method was applied to the analysis of energy drinks and honey with relative errors in glucose determination in the range 100 ± 4.3%. The advantages of the proposed method are the wide linear range, the simple instrumentation used, the low consumption of sample and reagents, the elimination of catalysts and immobilised enzymes and the high sample throughput.

Fluorometric Method for Quantitative Determination of Glucose and Its Application to Human Serum

By coupling the Fenton reaction with the glucose oxidase (GOD)-catalyzed reaction, we have developed a fluorometric method for the determination of glucose; the linear response range and the detection limit are 0.108-2.59 microg/ml and 0.0432 microg/ml, respectively. This approach is selective and has the advantages of using inexpensive reagents and avoiding interferences. Satisfactory results were obtained for the determination of glucose in human serum samples.

Determination of several sugars in serum by high-performance anion-exchange chromatography with pulsed amperometric detection

In this paper, a sensitive, simple and direct method for simultaneous determination of glucose, ribose, isomaltose and maltose in serum sample by high-performance anion-exchange chromatography coupled with integrated pulsed amperometric detection was developed. The four target analytes were easily and completely separated on an anion-exchange column at a flow-rate of 0.25 mL/min by binary step gradient elution in about 16 min and the two eluents were deionized water and 500 mM sodium hydroxide, respectively. The separated four analytes were detected directly by using a gold electrode and quadruple-potential waveform integrated pulsed amperometry without derivatization. Under the optimized conditions, when the injection volume was 25 μL, the detection limits (signal-to-noise ratio equal to 3) for glucose, ribose, isomaltose and maltose were 0.92, 7.50, 12.9 and 10.3 ng/mL, respectively. The calibration graphs of peak area for the four analytes were linear over two to three orders of magnitude with correlation coefficients greater than 0.998. R.S.D. of peak areas of the four analytes for five determinations were no more than 5.6%. The analytical method had been applied to the determination of glucose, ribose, isomaltose and maltose in real serum samples and good results with low relative standard deviation not more than 5.3% were obtained. The accuracy of the proposed method was tested by recovery measurements on spiked samples and good recovery results (98.1–107.9%) were obtained.

Application of heated electrodes operating in a non-isothermal mode for interference elimination with amperometric biosensors.

Heated electrodes were applied for the non-isothermal operation of amperometric glucose biosensors based on glucose oxidase immobilised on the electrode surface by entrapment within a polymer layer. The localised deposition of the polymer film under simultaneous entrapment of the enzyme was achieved by an electrochemically induced pH-modulation in the diffusion zone in front of the electrode, thus altering the solubility of the polymer chains. This non-manual sensor preparation protocol could be successfully used for the modification of a novel indirectly heated electrode. The non-isothermal operating mode allows working at the optimum temperature of the enzyme sensors without any thermal distortion of the bulk solution. Increased surface temperature of the sensor thus accelerates transport as well as kinetic processes, resulting in an enhanced amperometric signal. In the presence of interfering compounds such as ascorbic acid, the proposed technique allows use of the diverging thermal impact on the sensing process, for different electrochemically active compounds, for a deconvolution of the amperometric signal at different electrode temperatures. A calculation method for determination of glucose in the presence of one interfering compound is presented as a basis for a calculative interference elimination.

Real-time determination of glucose consumption by live cells using a lab-on-valve system with an integrated microbioreactor.

This paper describes a microquantitative method for glucose determination in situ of living cells in real-time. In this novel technique adherent cells are cultured onto microcarrier beads and packed into a renewable microcolumn within a microsequential injection lab-on-valve system (microSI-LOV). Glucose sensing is performed through the use of a two-step, NAD-linked enzymatic process. The course of the assay is monitored in real-time, by absorbance of NADH at 340 nm. The microsequential assay based on plug/nozzle design has a linear dynamic range for glucose of 0.1 to 5.6 mM. The design of the (microSI-LOV) system allows the assay to be carried out using only 40 microL of the enzyme reagent and 3 microL of sample. The technique was tested on a murine hepatocyte cell line (TABX2S) adhered to Cytopore beads. Rapid cellular glucose consumption, in this technique, is facilitated by a high cell density, which allows a large number of cells (10(4)-10(5)) to be retained in a very small volume (3 microL). In turn, this cell density results in the rapid depletion of glucose from the cell medium over short time periods (< 2 min). In conjunction with the assay development, the plug/nozzle design and its ramifications on mixing in general are presented and discussed.

Improvement of the Quantitative Method for Glucose Determination Using Hexokinase and Glucose 6-Phosphate Dehydrogenase

This paper has two aims. The first one is to point out the shortcomings of Food and Drug Administration's (FDA's) reference method for the measurement of glucose. We found that the quantity of enzyme used in the method recommended by the FDA was more than the exact quantity needed for accurate measurement. The use of exact quantity of enzyme is important to minimize the negative effects due to impurity and side reactions of enzymes. The second aim is to simulate the coupling enzyme reaction using computer. We have successfully established the exact quantity of enzyme needed in the assay through the computer simulation. The quantity of the enzyme was lesser than the that recommended by FDA, but the reaction ended at the same time as in the FDA method. In addition, optimum conditions and inhibitory effects of various reagents have also been successfully analyzed using computer. In conclusion, we suggest a reference method using computer simulation to determine the exact quantity of the coupling enzyme needed in the assay.

Comparison of the glucose oxidase method for glucose determination by manual assay and automated analyzer

In experimental models of diabetes, glucose levels in plasma and blood are commonly determined by colorimetric assay and by automated analyzers based on the glucose oxidase conversion of glucose and O 2 to gluconate and H 2O 2. We have compared the glucose levels obtained by these two methods in control Wistar rats, streptozotocin diabetic Wistar rats, Zucker fa/fa fatty rats and Zucker Diabetic Fatty rats. We found that the manual glucose assay and the glucose analyzer produced comparable values up to concentrations of about 25 mM. Above this level, samples should be diluted.

Combined methods for the determination of microbial activity of leaf litter

A sensitive enzymatic test for glucose is introduced as a specific method for the determination of cellulolytic activity. The amount of released glucose serves as a measure for the cellulase activity of leaf litter, when crystalline cellulose is used as substrate. We compare the enzymatic measurement with an established method for the non-specific determination of glucose as a reducing carbohydrate. In comparison to our results on cellulase activity, the results of a modified determination of microbial respiration for small samples of leaf litter did not show the expected positive correlation to cellulolytic activities. This indication for differential effects of leaf litter characteristics on the composition of microbial populations provides arguments for suggesting simultaneous estimations of different measures for microbial activity.

Implantation of a refillable glucose monitoring-telemetry device

This study describes the components and short-term in vivo evaluation of an integrated implantable system consisting of an amperometric glucose biosensor, a miniature potentiostat, a FM signal transmitter, and a power supply. The device (dimensions: 5·0 × 7·0 × 1·5 cm) was implanted subcutaneously in healthy mongrel dogs. The biosensor performance was evaluated in vitro prior to implantation using standard solutions simulating the physiological environment. A linear response to glucose concentration was observed throughout the physiological and pathophysiological range (with an upper limit of 25 mM glucose, and a sensitivity of 0·5 μA/mM). The results of short-term subcutaneous implantation of the integrated system demonstrated good agreement between the glucose concentration measured by the biosensor and that obtained using standard glucose determination methods. The delay-time between the tissue glucose level (measured by the biosensor) and the blood glucose level (obtained by standard methodology) was 3–7 min. These results demonstrated the feasibility of data transmission by a telemetry system through the skin of a dog and allowed the commencement of chronic in vivo testing. During the chronic implantation the biosensor was refilled in vivo. A rejuvenation of the sensor's response after refilling was observed suggesting the potential of such sensors for long-term implantation.

Computerized Enzymatic Kinetic Method for Glucose Determination in Foodstuff Using Immobilized Glucose Oxidase in Gel Beads

Abstract A new reliable semi-automatic procedure for enzyme immobilization in uniform calcium alginate beads is introduced, and a simple, low-cost measuring kinetic technique for enzymatic analysis is presented. Immobilized glucose oxidase, in calcium alginate beads, was used to catalyze the glucose oxidation reaction. The reaction was monitored photometrically at 460nm, by means of an indicator reaction based on the oxidation of o-dianisidine by hydrogen peroxide, in the presence of peroxidase. Under optimum conditions, the enzyme-gel beads show a good behaviour for at least two months and for at least 20 consecutive measurements on the same sample. The method was applied to the determination of glucose in foodstuffs like honey, artificial beverages, ice-cream toppings, and the results were compared with those obtained by two standard methods.

Glucose Biosensor Based on Oxygen Electrode Part IV: In Vivo Evaluation of the Rechargeable Glucose Sensor

Abstract A glucose monitoring system consisting of a pair of amperometric sensors: a glucose biosensor based on oxygen electrode and an oxygen sensor, two miniature potentiostats, an instrumentation amplifier and a data logger has been developed. The glucose sensor has linear response to the glucose concentration in vitro at 37°C up to 26 mM (480 mg/dL) in the phosphate buffer solution (pH 7.4), and linear range up to 21 mM (380 mg/dL) in undiluted bovine plasma. The system was evaluated in vivo with the sensors subcutaneously implanted in healthy mongrel dogs. During the implantation the system output was continuously recorded. The results of short-term subcutaneous implantation of the integrated system demonstrated good agreement between the glucose concentration measured by the biosensor and that obtained using standard glucose determination methods. The delay-time between the tissue glucose level (measured by the biosensor) and the blood glucose level (obtained by standard methodology) was 3 to 10 min...

Development of the Implantable Glucose Sensor: What Are the Prospects and Why Is It Taking So Long?

The development of an implantable glucose sensor for use in diabetes was first suggested in the 1960s (1–3). The sensor would provide an alternative to the present discrete methods of glucose determination that are based on intermittent blood sampling. Continuous glucose sensing would be particularly important in the detection and management of hypoglycemia. It would also allow early detection of hyperglycemia and provide a basis for insulin administration at more appropriate dosages and timing or for automatic insulin delivery from a pump. An implantable glucose sensor could also be used in parallel with other existing or potential forms of insulin replacement, such as transplantation or hybrid islet devices.

Volume and glucose concentration of rat amniotic fluid: Effects on embryo nutrition and axis rotation

In order to pursue our previous studies of the changes in neural tube microvilli produced by glucose, we developed a micro method of measuring glucose concentration in the very small volumes of amniotic fluid during neurulation. The volume of amniotic fluid was found to increase nearly 10-fold during major neurulation (day 10 to day 11 in the rat). This increase in volume and our repeated observations that physical removal of the restraining amnion initiates embryonic rotation leads us to propose that the growth of the amniotic cavity is essential for conversion from the ventral- to dorsi-flexion of the embryonic axis. Amniotic fluid volume continues to increase until day 18 but dropped by day 20. A method for glucose determination was developed using the color reaction on glucose oxidase indicator paper. The intensity of the color was analyzed with a color scanner. Amniotic glucose was 27.1 +/- 1.6 mg/dl on day 10 and continued in this range with some fluctuation until day 20 when it decreased. We isolated days 10, 11, and 16 embryonic sites from their decidua and incubated them at 0 degrees C and 38 degrees C while measuring glucose concentration. The glucose concentration did not show significant decrease at 0 degrees C on day 10 or 11 or on day 16 at 38 degrees C. At 38 degrees C the day 10 embryo amniotic fluid glucose disappeared after 22 minutes and the day 11 amniotic fluid glucose was gone in 34 minutes. These depletion times were statistically different. The magnitude of glucose depletion on day 10 was shown by calculation to be approximately 323 mumoles/gm protein per hour which is a substantial portion of the glucose utilized by the embryo as determined in previous experiments (731 mumoles/gm protein per hour). This model may serve as a way to study glucose utilization by embryos after their exposure to various teratogens.

Kurzmitteilung

The enzymatic method for glucose determination in plasma or serum using hexokinase/G6PDH in end-point was adapted for glucose determination as tha first rate reaction in blood hemolysate on RA-1000 autoanalyser. The method was found to be linear, precise, sensitive and accurate. Recovery of control samples ranged between 97.2% and 100.2%.

Micro-Enzyme Electrode as a High-Performance Detector of Flow Injection Analysis

Abstract The advantages of miniaturized enzyme electrode have been applied to flow injection analysis for the development of a rapid and selective method for the continuous determination of glucose. By assembling a micro-enzyme electrode having a diameter of 10 μm for an amperometric flow-through detector, at least 600 glucose samples can be determined in one hour. And micro-enzyme electrodes are satisfactorily stable for over 4 weeks even after 1000 glucose injections.

A method for determination of glucose 1,6-bisphosphatase.

A sensitive and specific method to measure glucose 1,6-bisphosphatase activity, which allows the identification of the reaction products is described. [U-14 C]Glucose 1,6-P2, synthesized by the glucose 1-P kinase activity of phosphofructokinase, is used as substrate. The reaction products are separated and identified by chromatography on ion-exchange paper.

Sequential determination of glucose and fructose in foods by flow-injection analysis with immobilized enzymes

A sequential method for determination of glucose and fructose involving the use of enzymes (hexokinase and glucose-6-phosphate dehydrogenase) immobilized on controlled-pore glass is proposed. The flow is selected so as to determine glucose or both sugars by fluorimetric determination of the NADH formed. The method is applied to the determination of these compounds in fruit juices, yoghourt and dessert powder with good results.

Electrocatalytic determination of glucose in a Ringer's solution by pulse-voltammetry.

Improvement of the electrocatalytic method of glucose determination was achieved by applying the pulse voltammetry technique. This technique, compared. to previously applied cyclic voltammetry, enhances selectivity of a platinum-black working electrode for glucose measurement. A low catalytic load working electrode has been used which is less susceptible to poisoning and has longer term stability. With cyclic voltammetry previously applied, those electrodes were limited by their narrow concentration range for glucose detection. By applying a pulsing technique, these electrodes exhibit linearity up to a glucose concentration 200 mg/dl with correlation coefficients greater than 0.9. The pulse voltammetry diminishes interference by low molecular weight serum components such as serum urea. Concentrations from 20. to 40 mg/di had an interference of 4 percent compared to 40 percent with cyclic voltammetry. Further investigations are carried on to adopt pulse voltammetry for direct measurements of glucose in body fluids.

New type valve for flow injection analysis; I. Application to stopped-flow spectrophotometric determination of glucose.

A new type valve for a stopped-flow flow injection analysis (FIA) was developed. The valve has two characteristic functions. The first is that a sample solution is injected in a “plug flow” into a flow of eluant, and the second is passage change-over of the eluant flow.At the load position, the eluant is recycled from tank to tank and the sample loop is filled with sample solution when the flow in reaction tubing is stopped. At the injection position a sample solution is injected into the flow of the eluant and is sent into reaction tubing. By rotating the valve from a injection position to a load position the reaction flow is stopped in a reaction tubing and the reaction will proceed. Stopped-flow FIA using the new type valve was one of the useful method for determination of glucose.