Total Cholesterol In Serum Samples Research Articles

Optical detection of total cholesterol based on a dye-displacement method

In the present study, the dye methylene blue (MB) was entrapped in an agarose gel and used as a sensing probe for the detection of total cholesterol. When methylene blue-entrapped agarose cubes were added to the cholesterol solution, methylene blue was displaced by cholesterol and released into the solution. A calibration curve was prepared by plotting the rate of release of methylene blue at 664 nm against varying cholesterol concentrations. A linear response was observed in the concentration range of 1 to 5 mM (40 mg/dL to 200 mg/dL) which covers normal and elevated cholesterol levels in humans. Optical detection of cholesterol using this dye-replacement method is simple, economical, and non-toxic. Characterisation of the system was carried out by FT-IR spectroscopy and cyclic voltammetry. The optical method was validated to determine total cholesterol in serum samples with reasonable accuracy.

Turning built-in electric field of porphyrin on Ti3+ self-doped blue-TiO2 hollow nanospheres boosts peroxidase-like activity for high-performance biosensing

The four well-designed porphyrins are synthesized with different electronegativity groups at the meso-substituted phenyl groups to turn the built-in electric field of the porphyrins. Among them, the meso-tetrakis(meta-hydroxyphenyl) porphyrin (mTHPP) with the largest diploes is revealed to form the strongest built-in electric field on the surface of Ti3+ self-doped blue TiO2 hollow nanospheres (b-TiO2). The b-TiO2 with Fenton-like activity couple mTHPP with a mass ration (the mass of mTHPP to the total sample mass) of 29% by stable hydrogen and electrostatic bonding, resulting in b-TiO2/29mTHPP. A synergistic effect of strong built-in electric field and the well-matching energy levels between mTHPP and b-TiO2 promotes the rapid multi-channel transmission of photo-generated electrons from mTHPP to b-TiO2. Consequently, the outstanding peroxidase (POD) -like activity achieved for b-TiO2/29mTHPP with the fast oxidation of TMB (3,3′,5,5′-tetramethylbenzidine) by H2O2 to form the blue product (oxTMB) only in 10 s under visible light. Furthermore, the b-TiO2/29mTHPP combined with cholesterol oxidase (Chox) can quantitatively determine cholesterol in the range of 0.005–10 mM with a limit of detection (LOD) of 1.2 μM within 1 min. The present sensing performance represents one of the best results for the cholesterol colorimetric sensor reported so far. Furthermore, this colorimetric biosensor has been used for accurately determining the levels of total cholesterol in serum samples, indicating the great potential in clinical medical detection. The present work provides new perspectives to design various POD-like nanozyme for biosensing applications.

5,10,15,20-tetrakis (4-carboxylphenyl) porphyrin functionalized NiCo2S4 yolk-shell nanospheres: Excellent peroxidase-like activity, catalytic mechanism and fast cascade colorimetric biosensor for cholesterol

5,10,15,20-tetrakis (4-carboxylphenyl) porphyrin functionalized NiCo2S4 yolk-shell nanospheres (Por-NiCo2S4) were fabricated by a facile and efficient solvothermal strategy and demonstrated to possess excellent peroxidase-like activity. Compared with pure NiCo2S4 nanospheres, Por-NiCo2S4 exhibits the superior peroxidase activity, which was evaluated by the fast oxidation of 3,3′,5,5′-tetramethylbenzidine (a chromogenic substrate, TMB) by H2O2 to form the blue product (oxTMB) only in 10 s. The systematic studies including radical scavenger, fluorescence probe, and electrochemical experiments confirm that superoxide free radicals (O2−) play a key role in catalytic reactions, in addition to the synergistic effect of holes (h+). Based on the developed peroxidase activity of Por-NiCo2S4, and combined with cholesterol oxidase (Chox), a fast cascade reaction sensing platform has been constructed to quantitatively determination cholesterol in a wide linear range of 0.1−9.0 mM with the detection limit of 19.36 μM. The Por-NiCo2S4 based colorimetric biosensing platform has been used for determining the levels of total cholesterol in serum samples, indicating Por-NiCo2S4 is expected to be used as the promising materials for fast visual colorimetry in clinical medical detection.

Usefulness of magnetically-controlled MNPs-enzymes microreactors for the fluorimetric determination of total cholesterol in serum

A new dynamic method containing a magnetically retained enzyme reactor (MRER) located in the reaction/detection zone of a flow injection (FI) system, has been used for the determination of total cholesterol in serum samples. The MRER was formed by a mixture ratio of 2/1 of immobilized enzymes cholesterol esterase (ChE) and cholesterol oxidase (COx) on magnetic nanoparticles (MNPs). The analytical signal is based on the fluorescence decreasing of the fluorophore naphtofluorescein (NF) due to its oxidation by the H2O2 formed in the enzymatic reactions. The dynamic range of the calibration graph was 1.55–100 mmol L−1 expressed as total cholesterol concentration (r2 = 0.9995, n = 5, r = 3), and the detection limit was 0.65 mmol L−1. The precision expressed as relative standard deviation (RSD %) was in the range of 4.7 and 0.6%. The method showed a sampling frequency of 10 h−1 and this method was applied to the determination of cholesterol in serum samples. The results were compared with those obtained using a previous automated clinical analyzer (ILab 600 analyzer). Also, recovery values ranging between 88.5 and 101.5% were achieved.

Fabrication and Optimization of ChE/ChO/HRP-AuNPs/c-MWCNTs Based Silver Electrode for Determining Total Cholesterol in Serum.

The developed method used three enzymes comprised of cholesterol esterase, cholesterol oxidase, and peroxidase for fabrication of amperometric biosensor in order to determine total cholesterol in serum samples. Gold nanoparticles (AuNPs) and carboxylated multiwall carbon nanotubes (cMWCNTs) were used to design core of working electrode, having covalently immobilized ChO, ChE, and HRP. Polyacrylamide layer was finally coated on working electrode in order to prevent enzyme leaching. Chemically synthesised Au nanoparticles were subjected to transmission electron microscopy (TEM) for analysing the shape and size of the particles. Working electrode was subjected to FTIR and XRD. The combined action of AuNP and c-MWCNT showed enhancement in electrocatalytic activity at a very low potential of 0.27 V. The pH 7, temperature 40°C, and response time of 20 seconds, respectively, were observed. The biosensor shows a broad linear range from 0.5 mg/dL to 250 mg/dL (0.01 mM–5.83 mM) with minimum detection limit being 0.5 mg/dL (0.01 mM). The biosensor showed reusability of more than 45 times and was stable for 60 days. The biosensor was successfully tested for determining total cholesterol in serum samples amperometrically with no significant interference by serum components.

A novel conducting copolymer: Investigation of its matrix properties for cholesterol biosensor applications

2-Heptyl-4,7-di(thiophen-2-yl)-1H-benzo[d]imidazole) (BImTh) was synthesized. Electrochemical copolymerization of this monomer with 2-(((9H-fluoren-9-yl)methoxy)carbonylamino)acetic acid (Fmoc-Gly-OH) was achieved on a graphite electrode and used as a matrix for amperometric cholesterol biosensing studies. In order to prepare a new cholesterol biosensor, cholesterol oxidase (ChOx) was covalently immobilized onto the copolymer coated graphite electrode. Cholesterol was used as the substrate and the decrease in oxygen level as a result of enzymatic reaction was monitored at −0.7V vs Ag reference electrode in a phosphate buffer (50mM, pH 7.0). Kinetic parameters, storage stabilities and surface characteristics were investigated. KMapp, Imax, LOD and sensitivity were calculated as 6.25μM, 9.69μA, 0.17μM and 2.47mA/mM cm2, respectively. This biosensor was applied to the determination of total cholesterol in serum samples. The estimation of cholesterol is an important for the diagnosis and prevention of several heart diseases and arteriosclerosis. Hence, it is important to develop new cholesterol biosensors.

Development of an amperometric enzyme electrode based on poly(o-phenylenediamine) for the determination of total cholesterol in serum

A polymer-based amperometric enzyme electrode (Pt/PoPD/ChOx) was prepared for the amperometric detection of free cholesterol. Firstly, poly(o-phenylenediamine) (PoPD) polymer film was prepared in acetonitrile-water medium containing o-phenylenediamine (oPD) monomer and (±)-10-camphorsulfonic acid (HCSA) on Pt electrode by the use of cyclic voltammetry technique. Cholesterol oxidase (ChOx) enzyme was immobilized onto Pt/PoPD electrode surface. The determination of cholesterol was performed via monitoring of the oxidation current of enzymatically produced H2O2 at +0.7 V vs. Ag/AgCl. Optimum buffer concentration, pH and working temperature were found as 0.05 mol L-1, 7.5 and 40 oC, respectively. The working range of enzyme electrode to cholesterol was 9.8 × 10-3-11 µmol L-1 and response time 150 s. The effects of possible interferences present in serum samples on response of enzyme electrode were examined. The determination of total cholesterol in serum samples was performed by using proposed Pt/PoPD/ChOx enzyme electrode and results were in good agreement with those obtained by spectrophotometric method.

SPE biosensor for cholesterol in serum samples based on electrochemiluminescent luminol copolymer

A poly(luminol–3,3′,5,5′-tetramethylbenzidine) copolymer manufactured by electropolymerization on screen-printed gold electrodes greatly improves the electrochemiluminescence of hydrogen peroxide. Cholesterol oxidase was immobilized on the surface of a poly(luminol–3,3′,5,5′-tetramethylbenzidine) screen-printed cell modified with chitosan to prepare an ECL biosensor for cholesterol. Working under the optimized conditions, the linear dynamic range of cholesterol was 2.4×10−5–1.0×10−3M with a limit of detection of 7.3×10−6M and a precision of 10.3% (5.0×10−4M, n=5) expressed as relative standard deviation. This biosensor was applied to the determination of total cholesterol in serum samples obtaining satisfactory results with respect to the reference procedure. This cholesterol biosensor offers an alternative analytical method with low cost and high speed.

Multiwalled carbon nanotube-based bi-enzyme electrode for total cholesterol estimation in human serum

This study aims at fabricating multiwalled carbon nanotubes (MWCNTs) based enzymatic bioelectrode for total cholesterol estimation in human serum. For this purpose, a gold (Au) electrode was modified with MWCNTs uniformly dispersed in nafion (Nf) matrix. Cholesterol oxidase (ChOx) and cholesterol esterase (ChEt) were immobilised onto this Nf–MWCNTs film-modified Au electrode using layer-by-layer technique to fabricate the final bioelectrodes. The immobilisation of ChOx and ChEt onto the electrodes was demonstrated by scanning electron microscopy. The fabricated bioelectrodes were electrochemically characterised using cyclic voltammetry. The bioelectrodes offer reliable response characteristics towards cholesterol and stable electrochemical properties in terms of extended linear response range of 0.080–0.950 mM, detection limit up to 0.01 mM and optimum storage stability up to three weeks. Experimental results reveal that the fabricated bioelectrode offer optimum repeatability and reproducibility towards the cholesterol estimation and can also efficiently exclude interference by the commonly coexisting ascorbic acid, uric acid, lactic acid, glucose and urea, which is favourable for its efficient use in the highly selective analysis of total cholesterol in serum samples.

Hand-held Photometer Based on Liquid-Core Waveguide Absorption Detection for Nanoliter-scale Samples

This paper reports a fully integrated hand-held photometer based on the liquid-core waveguide (LCW) detection principle for nanoliter-scale samples. All components of the photometer including light-emitting diode (LED) light source, LCW flow cell, photodiode detector, dropper pump, electronic circuit, liquid-crystal display screen, and battery were fully integrated into a small-sized (12 x 4.5 x 2.1 cm) instrument. A bent optical coupler was developed to conduct the detection light into or out of the LCW flow cell through its sidewall. This design allowed the sampling probe, input and output optical couplers, and LCW flow cell to be integrated in a single Teflon AF capillary, which significantly simplified system structure, improved working reliability, and reduced sample consumption. Two UV-LEDs were used as light source in the photometer to achieve dual wavelength detection at 260 and 280 nm, which was applied to assess on-site the quality and quantity of DNA samples. The effective optical path length of the photometer was approximately 15 mm with a sample consumption of only 350 nL. The potential of the photometer applied in point of care testing was also demonstrated in the measurement of total cholesterol in serum samples.

Biosensor for total cholesterol estimation using N-(2-aminoethyl)-3-aminopropyltrimethoxysilane self-assembled monolayer

Cholesterol oxidase (ChOx), cholesterol esterase (ChEt), and horseradish peroxidase (HRP) have been co-immobilized covalently on a self-assembled monolayer (SAM) of N-(2-aminoethyl)-3-aminopropyltrimethoxysilane (AEAPTS) deposited on an indium-tin-oxide (ITO) glass surface. These enzyme-modified (ChOx-ChEt-HRP/AEAPTS/ITO) biosensing electrodes have been used to estimate cholesteryl oleate from 10 to 500 mg dL(-1). The sensitivity, Km value, and shelf-life of these ChEt-ChOx-HRP/AEAPTS/ITO biosensing electrodes have been found to be 124 nA mg(-1) dL, 95.098 mg dL(-1) (1.46 mmol L(-1)), and ten weeks, respectively. The ChEt-ChOx-HRP/AEAPTS/ITO bio-electrodes have been used to estimate total cholesterol in serum samples.

Layer-by-layer self-assembled multilayer films of carbon nanotubes and platinum nanoparticles with polyelectrolyte for the fabrication of biosensors

Platinum nanoparticle-doped chitosan (CHIT) solution can be easily prepared by treating the CHIT solution with aqueous H 2PtCl 6 solution followed by chemical reduction of Pt(IV) with NaBH 4. Multiwalled carbon nanotubes (MWCNT) are then dispersed in the nanoparticle-doped solution. The resulting Pt-CNT-CHIT material brings new capabilities for electrochemical devices by using the synergistic action of Pt nanaoparticles and CNT. Positively charged Pt-CNT-CHIT solution and negatively charged poly(sodium- p-styrenesulfonate) salt (PSS) have been employed to fabricate stable ultrathin multilayer films on gold electrode and quartz glass slides in a layer-by-layer fashion. Cyclic voltammetric and UV–vis adsorption spectroscopy confirms the consecutive growth of the multilayer films. The modified gold electrode allows low-potential detection of hydrogen peroxide with high sensitivity and fast response time. With the immobilization of cholesterol oxidase onto the electrode surface using glutaric dialdehyde, a biosensor that responds sensitively to cholesterol has been constructed. In pH 6.98 phosphate buffer, almost interference free determination of cholesterol has been realized at 0.1 V vs. SCE with a linear range from 0.01 to 3 mM and response time<30 s. With the immobilization of another cholesterol esterase enzyme layer, the biosensor was used to determine total cholesterol in serum samples with satisfactory results.