Tan Ling

Potentiometric Urea Biosensor Based on an Immobilised Fullerene-Urease Bio-Conjugate

A novel method for the rapid modification of fullerene for subsequent enzyme attachment to create a potentiometric biosensor is presented. Urease was immobilized onto the modified fullerene nanomaterial. The modified fullerene-immobilized urease (C60-urease) bioconjugate has been confirmed to catalyze the hydrolysis of urea in solution. The biomaterial was then deposited on a screen-printed electrode containing a non-plasticized poly(n-butyl acrylate) (PnBA) membrane entrapped with a hydrogen ionophore. This pH-selective membrane is intended to function as a potentiometric urea biosensor with the deposition of C60-urease on the PnBA membrane. Various parameters for fullerene modification and urease immobilization were investigated. The optimal pH and concentration of the phosphate buffer for the urea biosensor were 7.0 and 0.5 mM, respectively. The linear response range of the biosensor was from 2.31 × 10−3 M to 8.28 × 10−5 M. The biosensors sensitivity was 59.67 ± 0.91 mV/decade, which is close to the theoretical value. Common cations such as Na+, K+, Ca2+, Mg2+ and NH4+ showed no obvious interference with the urea biosensors response. The use of a fullerene-urease bio-conjugate and an acrylic membrane with good adhesion prevented the leaching of urease enzyme and thus increased the stability of the urea biosensor for up to 140 days.

A novel electrochemical sensor for 17β-estradiol from molecularly imprinted polymeric microspheres and multi-walled carbon nanotubes grafted with gold nanoparticles

Molecularly imprinted polymers (MIPs) are generally a more stable material for sensing application. The high selectivity and sensitivity of MIPs for sensors can be achieved if the template molecule is imprinted in the polymer and this makes them an ideal alternative as a recognition element for sensors. A new electrochemical sensor based on molecularly imprinted polymeric microspheres (MIPs) and multi-walled carbon nanotube/gold nanoparticle (MIP–MWCNT–AuNP) modified carbon screen-printed electrodes (SPEs) for the rapid detection of 17β-estradiol (E) hormone in serum samples has been successfully developed. Hydrophobic MIPs were synthesized using photopolymerization in emulsion form. The multi-walled carbon nanotube grafted with gold nanoparticles was firstly deposited onto a carbon screen-printed electrode for the purpose of accelerating electron transfer to the surface of the electrode. The MIP microspheres specific to the 17β-estradiol hormone, prepared via a facile photopolymerization technique, were coated onto the MWCNT–AuNP modified SPE. The presence of 17β-estradiol in biological samples could be detected with the sensor via absorption of 17β-estradiol into the deposited MIPs and this was monitored by differential pulse voltammetry (DPV) at 0.6 V for the reduction of 17β-estradiol. Under optimal conditions, the sensor could detect the concentrations of 17β-estradiol from 1.0 × 10−15 to 1.0 × 10−6 M (R2 = 0.9921), with a detection limit of 2.5 × 10−16 M. The sensor based on MIP microspheres and the MWCNT–AuNP modified electrode demonstrated a stability of 55 days with good reproducibility (RSD < 5%, n = 5) and regenerability (RSD < 4%, n = 5). Using this sensor, the gender of the arowana fish determined via the level of 17β-estradiol using fish serum samples demonstrated good agreement with a conventional test kit based on the immuno-assay method.

Microencapsulated Aliivibrio fischeri in Alginate Microspheres for Monitoring Heavy Metal Toxicity in Environmental Waters

In this article a luminescence fiber optic biosensor for the microdetection of heavy metal toxicity in waters based on the marine bacterium Aliivibrio fischeri (A. fischeri) encapsulated in alginate microspheres is described. Cu(II), Cd(II), Pb(II), Zn(II), Cr(VI), Co(II), Ni(II), Ag(I) and Fe(II) were selected as sample toxic heavy metal ions for evaluation of the performance of this toxicity microbiosensor. The loss of bioluminescence response from immobilized A. fischeri bacterial cells corresponds to changes in the toxicity levels. The inhibition of the luminescent biosensor response collected at excitation and emission wavelengths of 287 ± 2 nm and 487 ± 2 nm, respectively, was found to be reproducible and repeatable within the relative standard deviation (RSD) range of 2.4–5.7% (n = 8). The toxicity biosensor based on alginate micropsheres exhibited a lower limit of detection (LOD) for Cu(II) (6.40 μg/L), Cd(II) (1.56 μg/L), Pb(II) (47 μg/L), Ag(I) (18 μg/L) than Zn(II) (320 μg/L), Cr(VI) (1,000 μg/L), Co(II) (1700 μg/L), Ni(II) (2800 μg/L), and Fe(III) (3100 μg/L). Such LOD values are lower when compared with other previous reported whole cell toxicity biosensors using agar gel, agarose gel and cellulose membrane biomatrices used for the immobilization of bacterial cells. The A. fischeri bacteria microencapsulated in alginate biopolymer could maintain their metabolic activity for a prolonged period of up to six weeks without any noticeable changes in the bioluminescence response. The bioluminescent biosensor could also be used for the determination of antagonistic toxicity levels for toxicant mixtures. A comparison of the results obtained by atomic absorption spectroscopy (AAS) and using the proposed luminescent A. fischeri-based biosensor suggests that the optical toxicity biosensor can be used for quantitative microdetermination of heavy metal toxicity in environmental water samples.

An optical biosensor from green fluorescent Escherichia coli for the evaluation of single and combined heavy metal toxicities.

A fluorescence-based fiber optic toxicity biosensor based on genetically modified Escherichia coli (E. coli) with green fluorescent protein (GFP) was developed for the evaluation of the toxicity of several hazardous heavy metal ions. The toxic metals include Cu(II), Cd(II), Pb(II), Zn(II), Cr(VI), Co(II), Ni(II), Ag(I) and Fe(III). The optimum fluorescence excitation and emission wavelengths of the optical biosensor were 400 ± 2 nm and 485 ± 2 nm, respectively. Based on the toxicity observed under optimal conditions, the detection limits of Cu(II), Cd(II), Pb(II), Zn(II), Cr(VI), Co(II), Ni(II), Ag(I) and Fe(III) that can be detected using the toxicity biosensor were at 0.04, 0.32, 0.46, 2.80, 100, 250, 400, 720 and 2600 μg/L, respectively. The repeatability and reproducibility of the proposed biosensor were 3.5%–4.8% RSD (relative standard deviation) and 3.6%–5.1% RSD (n = 8), respectively. The biosensor response was stable for at least five weeks, and demonstrated higher sensitivity towards metal toxicity evaluation when compared to a conventional Microtox assay.

An Amperometric Biosensor Based on Alanine Dehydrogenase for the Determination of Low Level of Ammonium Ion in Water

An amperometric electrochemical biosensor has been developed for ammonium ( N H 4 + ) ion detection by immobilising alanine dehydrogenase (AlaDH) enzyme in a photocurable methacrylic membrane made up of poly(2-hydroxyethyl methacrylate) (pHEMA) on a screen-printed carbon paste electrode (SPE). The current detected was based on the electrocatalytic oxidation of nicotinamide adenine dinucleotide reduced (NADH) that is proportional to the consumption of N H 4 + ion whilst enzymatic amination of AlaDH and pyruvate is taking place. The biosensor was operated amperometrically at a potential of +0.6 V and optimum pH 7. The N H 4 + biosensor demonstrated linear response to N H 4 + ion concentration in the range of 0.03–1.02 mg/L with a limit of detection (LOD) of 8.52 μg/L. The proposed method has been successfully applied to the determination of N H 4 + ion in river water samples without any pretreatment. The levels of possible interferents in the waters were negligible to cause any interference on the proposed method. The analytical performance of the biosensor was comparable to the colorimetric method using Nesslerisation but with much lower detection limit and linear response range at ppb level.An amperometric electrochemical biosensor has been developed for ammonium (NH4 ) ion detection by immobilising alanine dehydrogenase (AlaDH) enzyme in a photocurable methacrylic membrane made up of poly(2-hydroxyethyl methacrylate) (pHEMA) on a screen-printed carbon paste electrode (SPE). The current detected was based on the electrocatalytic oxidation of nicotinamide adenine dinucleotide reduced (NADH) that is proportional to the consumption of NH4 + ion whilst enzymatic amination of AlaDH and pyruvate is taking place. The biosensor was operated amperometrically at a potential of +0.6 V and optimum pH 7. The NH4 + biosensor demonstrated linear response to NH4 + ion concentration in the range of 0.03–1.02 mg/L with a limit of detection (LOD) of 8.52 μg/L. The proposed method has been successfully applied to the determination of NH4 + ion in river water samples without any pretreatment. The levels of possible interferents in the waters were negligible to cause any interference on the proposed method. The analytical performance of the biosensor was comparable to the colorimetric method using Nesslerisation but with much lower detection limit and linear response range at ppb level.

Single-step and reagentless analysis of genetically modified soybean DNA with an electrochemical DNA biosensor

In many electrochemical DNA based biosensors, a redox indicator is required to be introduced separately to indicate the DNA hybridization event. In this work, we have developed a simple procedure for voltammetric determination of CaMV 35S gene modified DNA without the need for introduction of a redox indicator. The DNA biosensor contains an immobilized DNA probe and also a methylene blue redox indicator that is able to slow release during the hybridization event. The biosensor was constructed from a screen printed carbon paste electrode (SPE) coated with an acrylic microsphere (AM)–gold nanoparticle (AuNP) composite immobilized with single-stranded DNA probes, whilst methylene blue (MB) was immobilized in the hydrogel poly(2-hydroxyethyl methacrylate) membrane located next to the electrode. Genetically modified (GM) DNA was examined based on the reduction in the MB cathodic peak current (ipc) signal, which was ascribed to the DNA hybridization event, via the differential pulse voltammetry (DPV) method. The ipc current signal of MB after DNA hybridization with target CaMV 35S gene DNA was linearly related to the logarithmic target DNA concentration ranging from 2.0 × 10−12 to 2.0 × 10−7 M (R2 = 0.989) with a limit of detection (LOD) at 1.40 × 10−13 M. The proposed AM–AuNP composite DNA electrode gave satisfactory reproducibility performance with <10% (n = 5) relative standard deviation (RSD). The recoveries between 94.1 ± 2.2% and 103.7 ± 8.2% (n = 5) were obtained when the DNA biosensor was used for GM DNA determination in GM soybean DNA samples. The DNA biosensor based on the AM–AuNP composite deposited SPE and immobilized MB exhibited higher sensitivity by single-step analysis compared with conventional electrochemical sensors.

The Effect of Multilayer Gold Nanoparticles on the Electrochemical Response of Ammonium Ion Biosensor Based on Alanine Dehydrogenase Enzyme

The use of multilayer of gold nanoparticles (AuNPs) attached on gold electrode surface via thiol chemistry to fabricate an ammonium (NH41 School of Chemical Sciences and Food Technology, Faculty of Science and Technology, Universiti Kebangsaan Malaysia (UKM) Selangor, 43600 Bangi, Malaysia 2 Industrial Chemical Technology Program, Faculty of Science and Technology, Universiti Sains Islam Malaysia, Bandar Baru Nilai, Negeri Sembilan, 71800 Nilai, Malaysia 3 Department of Chemistry, Faculty of Science, National University of Singapore, Singapore 117543

Aminated hollow silica spheres for electrochemical DNA biosensor

An electrochemical DNA biosensor for e.coli determination based on aminated hollow silica was successfully developed. Aminated hollow silica spheres were prepared through the reaction of Tween 20 template and silica precursor. The template was removed by the thermal decomposition at 620°C. Hollow silica spheres were modified with (3-Aminopropyl) triethoxysilane (APTS) to form aminated hollow silica spheres.Aminated DNA probe were covalently immobilized on to the amine functionalized hollow silica spheres through glutaradehyde linkers. The formation hollow silica was characterized using FTIR and FESEM. A range of 50-300nm particle size obtained from FESEM micrograph. Meanwhile for the electrochemical study, a quasi-reversible system has been obtain via cyclic voltammetry (CV).