Detection Limit For Glucose Research Articles

Thermal stability of glucose and other sugar aldoses in normal phase high performance liquid chromatography

Analysis of glucose and other simple sugars are often performed by use of normal phase HPLC methods with acetonitrile as major eluent. The present results clearly show that column temperature plays an important role with respect to chromatographic performance and detection limits of glucose when using a specific carbohydrate column. A change in column temperature from 25 to 45 °C reduced the detection of glucose (with ELSD) by more than 41%, whereas the detection of other sugar aldoses (galactose, xylose and rhamnose) were suppressed even more. By increase of column temperature to 70 °C the detector signal of glucose was found to be less than 2% compared to that obtained at 20 °C. Neither fructose nor sucrose showed similar correlation between column temperature and detection. The rate of decreased response is not dependent on sample concentration or the ELSD settings. The results express the importance of accurate temperature control in the analysis of sugar aldoses, and also values low column temperatures for samples with low concentrations of sugar aldoses in order to improve detection.

Study of different carbon materials for amperometric enzyme biosensor development

Diamond-like carbon (DLC) electrodes which constitute a new research area in electrochemistry and glassy carbon (GC) electrodes were used as transducers for fabrication of glucose oxidase (GOD) biosensors. The amperometric signal of the enzyme electrode was due to the electro-oxidation of H 2O 2 generated in the enzyme layer. This work has shown that the detection limit of glucose on GOD/GC electrode is 20 μM while it is 50 μM on GOD/DLC electrode. The sensitivity of GOD/GC electrode decreases 4% after 8 days and we have good repeatability of measurements for GOD/DLC electrode in the same day.

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.

Combination of Gold‐Modified Electrode and α‐Amyloglucosidase for Simultaneous Determination of Starch and Glucose

The electrocatalytic activity of copper‐modified gold electrodes has been investigated for the simultaneous detection of glucose and starch using α‐amyloglucosidase in solution. The amperometric response of glucose at the modified gold electrode was monitored for different copper loadings (0.04, 0.1, 0.2, and 0.3 mg cm−2), that of 0.2 mg cm−2 deposited on the electrode surface being chosen for further investigation. The surface characteristics were confirmed by the electrochemical impedance spectra. The oxidation of glucose and starch was studied by linear sweep voltammetry (LSV) in order to determine the best applied potential for measuring glucose in the presence of starch, a value of +0.4 V vs. SCE being selected. The glucose detection limit at +0.4 V is equal to 0.64 mg L−1 in the absence of starch. The same detection limit was found after adding 200 mg L−1 starch to the cell. The properties of the copper‐deposited films were investigated in 0.15 M NaOH solution for the determination of α‐amyloglucosidase enzyme activity. The enzymatic reaction conditions were optimized. Conversion of starch into glucose was found to be around 89% by using α‐amyloglucosidase (2970 mUI) at 55°C for 30 min.

Metal clad leaky waveguides for chemical and biosensing applications

Novel metal clad leaky waveguide (MCLW) sensor devices have been developed for sensing applications. These chips are designed to confine the light in a low refractive index waveguide that encompasses the chemically-selective layer, maximising the overlap between the optical mode and the chemistry, thus improving the sensitivity. In this work, a thin metal layer was inserted between the substrate and the thick waveguide layer, increasing the reflectivity of the waveguide/metal interface and decreasing the light lost at each of reflection in the leaky mode, which in turn increases the propagation distance. The device has been used for a range of biosensing applications, including the detection of organophosphoros pesticides. The limit of detection for paraoxon, based on absorbance detection, was calculated to be 6 nM. Refractive index detection was demonstrated by monitoring the change in the out-coupled angle resulting from the binding of protein A to anti-protein A immobilized on agarose. The sensor was also used for detecting the quenching of the fluorescence of an acid–base sensitive ruthenium complex immobilized within the sol–gel and with glucose oxidase enzyme. The limit of detection for glucose was 3 μM. The advantage of using the metal layer in the MCLW was that an electrical potential could be applied to accelerate the diffusion of the analyte to the immobilised antibody, which resulted in a shortened analysis time and a reduction in non-specific binding.

Amperometric screening of bacterial food contamination using a composite modified electrode

An indirect determination of bacterial contamination based on catabolic metabolism of glucose is proposed. The determination of glucose was carried out by a flow injection (FI) system with amperometric detection using a wall-jet flow cell. A new composite PVC–graphite electrode modified with Cu 2O was developed to improve the analytical features of glucose determination. A 25 mM saline buffer sodium phosphate in 0.2 M NaOH was used as the carrier for glucose. After optimising the sensing electrode in terms of composite electrode formulation, hydroxide ion concentration and operating potential, the detection limit for glucose was 1.1×10 −3 M for 100 μl injections. The proposed method allows the successful determination of pollution bacteria in food samples in 3 h as compared to the 24–48 h required using the plate count method. The detection limit for bacteria in the samples was 2×10 2 cfu ml −1 and the reproducibility was in the range 2.6–3.8%. The efficiency of this method was tested for eight types of bacteria and five kinds of food. A comprehensive study of 30 spiked samples and 30 blank samples provided 4% of false positives and 0% of false negatives showing the applicability of the proposed method for sample screening.

Glucose quantification in dried-down nanoliter samples using mid-infrared attenuated total reflection spectroscopy.

The aim of this study was to determine the feasibility of minimally invasive glucose concentration measurement of a body fluid within the physiologically important range below 100 nL with a number of samples such as interstitial fluid, plasma, or whole blood using mid-infrared spectroscopy, but starting with preliminary measurements on samples of simple aqueous glucose solutions. The Fourier transform infrared spectrometer was equipped with a Golden Gate single reflection diamond attenuated total reflection (ATR) accessory and a room-temperature pyroelectric detector. As the necessary detection limits can be achieved only for dried samples within the spectrometric conditions realized by a commercial instrument, the work focused on the optimization of such ATR measurements. We achieved quantification of samples with volumes as low as 7 nL between 10 and 600 mg/dL. The standard error of prediction (SEP) for the concentration range 10-100 mg/dL is 3.2 mg/dL with full interval data between 1180 and 940 cm(-1). The performance of the prediction is given by a coefficient of variation of prediction (CV(pred) ) of 6.2%. When all samples within the whole concentration range are included, the SEP increases to 20.2 mg/dL, and hence the CV(pred) to 10.6% due to a nonlinear signal dependence on glucose concentration. A detection limit for glucose of 0.7 ng with a signal-to-noise ratio of 10 was obtained.

Calorimetric biosensors with integrated microfluidic channels

A microfluidic device capable of measuring real-time enthalpy changes of biochemical reactions and thermal properties of biological fluids is presented in this paper. The device consists of a freestanding microthermopile integrated with a glass microfluidic reaction chamber. The p-type polysilicon/gold microthermopiles fabricated on a 2 μm thick thermally isolated membrane showed a sensitivity of 0.94 V/W and a thermal time constant of less than 100 ms. Although the device is not restricted to enzymatic reactions, in this paper measurements of the heat of reaction from the catalytic action of glucose oxidase, catalase, and urease on glucose, hydrogen peroxide, and urea, respectively, are reported. Reactions were performed in open air using liquid batch testing and in enclosed fluidic reaction chamber by continuous flow experiments. A sensitivity of 53.5 μV/M for glucose, 26.5 μV/M for hydrogen peroxide and 17 μV/M for urea was obtained. Detection limit for glucose in the continuous flow mode is ∼2 mM (30 pmol). The aim of this work is to demonstrate the potential of the integrated calorimetric microfluidic device for fundamental thermodynamic studies in biochemical reactions. Using arrays of such devices with immobilized enzymes multi-analyte detection can be accomplished and the effects of interferents from competing substrates can be compensated. This paper presents the design, fabrication and initial testing results from such a microthermopile-based thermal biosensor.

Reversible Optical Sensor Membrane for Hydrogen Peroxide Using an Immobilized Fluorescent Probe, and its Application to a Glucose Biosensor

An optical sensor for hydrogen peroxide (HP) based on the probe europium tetracycline (EuTc) incorporated into a polyacrylonitrile-co-polyacryl- amide polymer matrix is described. Upon optical exci- tation with 400-nm light, the EuTc in the membrane displays fairly strong fluorescence peaking at 616 nm. The fluorescence intensity increases up to 3-fold if the sensor is exposed to solutions containing HP. The effect is reversible and can thus be used for continuous sen- sing. The largest signal changes with HP are observed at pH levels between 6.5 and 7.5, and the range of the response is between 10 and 300 ppm (w=w) of HP, equal to 0.3 to � 10 mmol L � 1 ). At concentrations above 0.3% of HP, decomposition of the EuTc in the membrane is observed. The limit of detection is � 15 ppm (0.45 mmol L � 1 ). The response is fully reversible and rather slow (� 10 min) in both directions, but the reverse response may be accelerated by addition of a reducing agent such as thiosulfate. Alkali ions and most anions remain inert, but phosphate and citrate interfere, as do Cu 2þ ions, which quench fluorescence. In order to image the spatial distribution of HP concentrations, sensor membranes were placed on the bottom of the wells of a microtiter plate, and their fluorescence was imaged using an LED-based device based on the mea- surement of the luminescence decay time of EuTc. If glucose oxidase is immobilized on the sensor layer, a glucose sensor is obtained in which the HP sensor acts as the transducer and which can quantify glucose in concentrations between 0.1 and 5 mmol L � 1 . The limit of detection for glucose is � 0.2 mmol L � 1 .

Ultrafast analysis of oligosaccharides on microchip with light-emitting diode confocal fluorescence detection.

We have developed a new method for the high-speed separation and high-sensitivity detection of complex oligosaccharides based on microchip electrophoresis (nu-CE) with light-emitting diode (LED) confocal fluorescence detection. Oligosaccharides labeled with 8-aminopyrene-1,3,6-trisulfonate (APTS) were found to strongly adsorb to the surface of polymethylmethacrylate (PMMA) microchips. Accordingly, three classes of major dynamic coating additives were systematically investigated, and cellulose derivatives were found to specifically suppress such adsorption and allow high-performance separation on PMMA chips. Additive concentration, buffer pH and applied field strength were found to be key factors in the high-performance separation& of APTS-labeled oligosaccharides on PMMA chips. Under optimal conditions, 15 oligosaccharides in dextrin hydrolysate can be separated within 45 s with an electrophoretic separation efficiency of over 400 000 theoretical plates per meter. The relative standard deviation (RSD) values of migration times of fourteen oligosaccharides were less than 0.50% between six different channels, and the detection limit for APTS-labeled glucose was about 1.98 x 10(-8) mol/L or 8.61 amol with a signal-to-noise ratio (S/N) of 3. The high speed, high efficiency and high sensitivity of this micro-CE-based method indicate that it can be widely applied to analysis of complex oligosaccharides.

導電性中空糸膜電極による糖検出感度の検討と多成分糖の同時検出

The metal coated hollow fiber membrane (MCH) has been applied as a working electrode of an electrochemical detector for high performance liquid chromatography (HPLC). In this paper, the improvements of the sensitivity of the copper coated hollow fiber membrane (Cu-MCH) electrode as a sensor for carbohydrates were described. The MCH electrode had the tubular structure. Therefore, there was a problem of decreasing the output signal by dilution of analysis with the dependence on the inside volume. In order to reduce the inside volume, the plug put inside of the MCH and a thinner hollow fiber membrane were used. The plug was effective to make the MCH electrode sensitive resulting the detection limit (S/N=3) of glucose lowered from 1.7 pmol to 0.95 pmol. The hollow fiber membrane in which inside diameter was thinner was also effective. The detection limit dropped from 1.7 pmol to 0.56 pmol, when it was changed to 0.6 mm from 1.0 mm. In addition, it was effective to decrease the noise current when the hollow fiber membrane of 0.6 mm inside diameter was covered with the copper on the gold (Cu/Au-MCH).The detection limit was improved to 0.24 pmol. The simultaneous detection of sorbitol, glucose, fructose, lactose, saccharose, raffinose and maltose was demonstrated with the Cu/Au-MCH in HPLC. The detection limits were 0.39, 0.24, 0.39, 0.38, 0.58, 0.53 and 0.82 pmol, respectively.

Properties of glucose biosensors based on dendrimer layers. Effect of enzyme immobilization.

The properties of glucose biosensors based on dendrimer layers on a gold support, which depend on the method of immobilization of glucose oxidase (GOX), were studied by amperometry. The kinetic parameters of enzymatic reactions, response time, sensitivity, detection limit, linear range, and enzyme turnover were determined. We showed that a more stable and sensitive sensor was obtained when GOX was immobilized on the dendrimer by crosslinking with glutaraldehyde in vacuum. This biosensor was stable for at least eight weeks. The response time was approximately 1.3 min, the detection limit of glucose was 25 micro M, and the apparent Michaelis-Menten constant was relative low ( K(m)=1.1+/-0.1 mM) in comparison with that for GOX in solution. The reason for these differences is discussed. The example of the application of the developed biosensors for the detection of mercury is also presented. The inhibitory effect of mercury on GOX activity was observed at mercury concentration of 100 nM.

Preparation, characterization and application of alkanethiol self-assembled monolayers modified with tetrathiafulvalene and glucose oxidase at a gold disk electrode

A critical study of the different variables affecting the preparation of enzyme electrochemical biosensors using self-assembled monolayer (SAM)-modified electrodes is reported. Regarding variables affecting the obtention of SAMs on a gold disk electrode (AuE), the type of alkanethiol employed, its concentration, the time and temperature of formation and the composition of the medium were evaluated. Concerning a glucose biosensor based on SAM-modified AuEs, the performances of several redox mediators for the enzymatic oxidation of glucose were compared. Two glucose oxidase (GOx) immobilization methods on the SAM-modified AuE were tested: a covalent binding using 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide and N-hydroxysulfosuccinimide (NHSS), and a cross-linking with glutaraldehyde. The cross-linking method produced slopes of the substrate calibration graphs at a mercaptopropionic acid (MPA)-modified electrode two orders of magnitude higher than those obtained with the covalent binding. Moreover, the mediator tetrathiafulvalene (TTF) was co-immobilized atop the SAM together with GOx. A heterogeneous electron transfer constant between TTF and the MPA–GOx bioelectrode of 1.25 s −1 was calculated. The enzyme loading, the amount of TTF on the electrode, the applied potential and the pH were also optimized. A good repeatability of the measurements with the TTF–GOx–MPA–AuE biosensor was demonstrated, with no need of pretreatment of the modified electrode. No significant changes in the slope value for the glucose calibration graph were found after 5 days when working with the same biosensor. An apparent Michaelis–Menten constant of (13.9±0.5) mM, and a limit of detection for glucose of 3.5×10 −6 M were obtained. Moreover, the TTF–GOx–MPA–AuE also performed well in the flow-injection mode and in the analysis of glucose in real samples.

ANALYSIS OF CARBOHYDRATES BY CAPILLARY ZONE ELECTROPHORESIS WITH ON-CAPILLARY DERIVATIZATION

Carbohydrates were on-capillary derivatized by p-hydrazine-benzenesulfonic acid (BHZ) and separated by capillary zone electrophoresis using 100 mM boric acid (pH 10.2) as running buffer. The derivatization was achieved by keeping the capillary tip introduced with sample in between two zones of labeling reagent at room temperature for 30 min. The detection limit of galactose and glucose were 15.6 and 31.2 μm, respectively, a bit higher than the corresponding values from pre-capillary derivatization (10.5 and 3.6 μM, respectively). Nevertheless the sample consumption was decreased by two to three orders of magnitude compared to the pre-capillary mode. Reproducible quantification of carbohydrates was demonstrated in the concentration range of 0.3–30 mM. The application of this method to determine the glucose in human serum was also present.

Luminol chemiluminescence in unbuffered solutions with a cobalt(II)-ethanolamine complex immobilized on resin as catalyst and its application to analysis.

Using a heterogeneous catalyst, Co(II)-ethanolamine complex sorbed on Dowex-50W resin, the chemiluminescence (CL) of luminol in unbuffered or weakly acidic solution was studied in the presence of H2O2. The maximum luminol CL wavelength at pH 5.7 was 448 nm, 23 nm longer than that in a basic solution (pH 10.5). Three different ligands, mono-, di-, and triethanolamine, and six transition metal ions, Co(II), Cu(II), Ni(II), Mn-(II), Fe(II), and Fe(III) were compared by CL measurements. The CL intensity decreased in the order mono- > di- > triethanolamine and Co(II) > Cu(II) > Ni(II) > Fe-(III) > Mn(II) > Fe(II). This heterogeneous CL system was developed as H2O2 and glucose flow-through sensors. Detection limits (S/N = 3) of H2O2 and glucose using Dowex-50W-X4-Co(II)-monoethanolamine as catalyst are 1 x 10(-7) M and 1 x 10(-6) M, respectively. On the basis of the studies of the CL, fluorescence, UV-vis and ESCA spectra and the effect of dissolved oxygen in luminol solution, a mechanism for CL emission in unbuffered solution was considered as the formation of a superoxide radical ion during the decomposition of H2O2 catalyzed by the Co(II)-ethanolamine immobilized resin. Then the superoxide radical ion acted on luminol and the CL was emitted. The applications of the proposed method to determine H2O2 in rainwater without any special pretreatment and glucose in human urine and orange juice samples give satisfactory results.

Simultaneous monitoring of extracellular glucose, pyruvate, lactate and glutamate in gerbil cortex during focal cerebral ischemia by dual probe microdialysis

The aim of this study was to monitor dynamic changes in energy-related metabolites in the cortex of gerbils subjected to cerebral ischemia by a dual probe microdialysis technique. Focal cerebral ischemia was produced in anesthetized gerbils by occlusion of the right common carotid artery and the right middle cerebral artery for 60 min. Two microdialysis probes were inserted into both sides of the cortex to simultaneously monitor extracellular glucose, lactate, pyruvate and glutamate. Dynamic and comparative changes in these analytes, on the ipsilateral and contralateral sides of the brain, were simultaneously monitored by liquid chromatography and a microdialysis analyzer. The present study demonstrated decreases in glucose and pyruvate, increases in lactate and glutamate on the ipsilateral side whereas all analytes remain constant on the contralateral side of cortex during cerebral ischemia. In vitro recovery of each microdialysis probe was performed to ensure the quality of experiments. The detection limits of pyruvate, glutamate, lactate and glucose were 0.2, 1.0, 2.0 and 20 μ M, respectively. The intra- and inter-assay correlations were less than 5% in standard mixtures and pooled brain dialysates.

Determination of carbohydrates in food samples by capillary electrophoresis with indirect UV detection

An easy and reproducible capillary electrophoretic method with indirect UV detection for the determination of carbohydrates in food samples was described. A highly alkaline pH condition was used in order to charge every carbohydrate negatively. Electroosmotic flow (EOF) was reversed to detect sugar alcohols and to improve the separation of neutral and amino sugars. Under optimized conditions 28 carbohydrates including mono- and disaccharides, uronic acids, sialic acids, amino sugars and sugar alcohols were separated in less than 25 min. The detection limits for fructose, glucose and sucrose were in the range from 12 to 16 mg l −1 with pressure injection of 50 mbar for 6 s (ca. 6 nl) at a signal to noise ratio of 3. The calibration curves were linear over the range from 50 to 10 000 mg l −1 with correlation coefficients better than 0.9999. This method could be readily applied to the analysis of carbohydrates in several foods and beverages. Sample preparation was minimal and consisted either of dilution or dilution with filtration.

Fiberoptic biosensors based on chemiluminescent reactions.

The chemiluminescence of luminol in the presence of H2O2 has been exploited to develop fiberoptic biosensors associated with flow injection analysis systems. A chlorophenol sensor was developed based on the ability of certain halophenols to enhance the peroxidase-catalyzed luminol chemiluminescence. Horseradish peroxidase immobilized on a collagen membrane was used. Ten chlorophenols have been tested with this chemiluminescent-based sensor. The lower detection limit was obtained with 4-chloro-3-methylphenol and was equal to 0.01 microM. Electrochemiluminescent-based fiberoptic biosensors for glucose and lactate were also developed using glucose oxidase or lactate oxidase immobilized on polyamide membranes. In the presence of oxidase-generated H2O2, the light emission was triggered electrochemically by means of a glassy carbon electrode polarized at +425 mV vs a platinum pseudo-reference electrode. The detection limits for glucose and lactate were 150 and 60 pmol, respectively, and the dynamic ranges were linear from 150 pmol to 600 nmol and from 60 pmol to 60 nmol, respectively.

Nickel content dependence of electrochemical behavior of carbohydrates on a titanium–nickel alloy electrode and its application to a liquid chromatography detector

A highly sensitive amperometric detector for the liquid chromatographic analysis of carbohydrates has been developed based on the optimization of the Ni content in a Ni–Ti alloy electrode. An optimum nickel content of 30 at. % was obtained from a signal-to-noise analysis in a flow injection experiment on glucose in 0.1 M NaOH. An optimum working potential of 0.5 V vs. Ag/AgCl was obtained from a hydrodynamic voltammogram. The glucose detection limit in the flow injection analysis was 11 fmol. The electrode was applied to an LC detector with a microbore column and the detection limit was 50 fmol.

Luminol electrochemiluminescence-based fibre optic biosensors for flow injection analysis of glucose and lactate in natural samples

A fibre optic biosensor based on the electrochemiluminescence of luminol has been developed for glucose and lactate flow injection analysis. The electrochemiluminescence of luminol was generated using a glassy carbon electrode polarised at +425 mV vs. a platinum pseudo-reference electrode. After optimisation of the reaction conditions and physicochemical parameters influencing the sensor response, the measurement of hydrogen peroxide could be performed in the range 1.5 pmol–30 nmol. Glucose oxidase or lactate oxidase, were immobilised on polyamide and collagen membranes. With collagen as the enzymatic support, the detection limits for glucose and lactate were 60 pmol and 30 pmol, respectively, whereas with the enzymatic polyamide membranes, the corresponding values were 150 pmol and 60 pmol. In each case, glucose and lactate measurements could be performed over four decades. The luminol electrochemiluminescence-based biosensors were used for glucose and lactate measurements in sera and also for lactate measurements in whey solutions. Good agreements were obtained between the present method and reference methods. For glucose analysis in serum, the coefficient of variation for 53 repeated measurements performed over a 10 h period was 4.8% while for lactate analysis, 80 assays performed over a 15 h period gave a coefficient of variation of 6.7%.