Nanobiosensor Design to Detect Cholic Acid Using Multiwalled Carbon Nanotube /TiO2 Nanoparticle for 3α-Hydroxysteroid Dehydrogenase Immobilization

Document Type : Research Article


1 Faculty of Chemistry, Razi University, Kermanshah, I.R. IRAN

2 Nanobiotechnology Department, Faculty of Innovative Science and Technology, Razi University, Kermanshah, I.R. IRAN

3 Faculty of Education-Chemistry, Thi Qar University, IRAQ


Determination of cholic acid concentration is a useful method to monitor liver diseases. We propose a rapid and simple method for measuring cholic acid. The development of a cholic acid electrochemical biosensor is described that is based on the modification of glassy carbon electrode surface using a mixture of carboxylated multiwalled carbon nanotube and titanium dioxide nanoparticles in chitosan solution and immobilization of 3α-hydroxysteroid dehydrogenase. The modification process of the sensing surface was characterized by Fourier transform infrared spectroscopy, Energy Diverse X-ray Spectrometry, Field Emission Scanning Electron Microscopy, and voltammetry techniques. A good correlation was demonstrated between cholic acid concentration and the peak currents in the presence of nicotinamide adenine dinucleotide. Using a carboxylated multiwalled carbon nanotube and titanium dioxide nanoparticles for electrode modification showed more effective area than an unmodified electrode at optimum pH of 6. Two linear ranges were obtained at 7.1 - 42.7, and 70.9-476.2 nM of cholic acid. Also, the detection limit was 6 nM and the sensitivities of the two ranges were obtained 956.9 and 28.7 µA/µM.


Main Subjects

[1] Kim H.J., Jang C.-H., Liquid Crystal-Based Capillary Sensory Platform for the Detection of Bile Acids, Chem. Phys. Lipids., 204: 10-14 (2017).
[2] Rössger K., Charpin-El-Hamri G., Fussenegger M., Bile Acid-Controlled Transgene Expression in Mammalian Cells and Mice, Metab. Eng., 21: 81-90 (2014).
[3] He S., Liang W., Tanner C., Cheng K.-L., Fang J., Wu S.-T., Liquid Crystal Based Sensors for the Detection of Cholic Acid, Anal. Methods, 5: 4126-4130 (2013).
[4] Niu X., Luo D., Chen R., Wang F., Sun X., Dai H., Optical Biosensor Based on Liquid Crystal Droplets For Detection of Cholic Acid, Opt. Commun., 381: 286-291 (2016).
[5] Okuda H., Obata H., Nakanishi T., Hisamitsu T., Matsubara K., Watanabe H., Quantification of Individual Serum Bile Acids in Patients with Liver Diseases Using High-Performance Liquid Chromatography, Hepato-Gastroenterology., 31: 168-171 (1984).
[6] Ghaffarzadegan T., Nyman M., Jönsson J.Å., Sandahl M., Determination of Bile Acids by Hollow Fibre Liquid-Phase Microextraction Coupled with Gas Chromatography, J. Chromatogr. B Biomed. Appl., 944: 69-74 (2014).
[7] Mashige F., Tanaka N., Maki A., Kamei S., Yamanaka M., Direct Spectrophotometry of Total Bile Acids in Serum, Clin. Chem., 27: 1352-1356 (1981).
[9] Van den Berg J., Van Blankenstein M., Bosman-Jacobs E.P., Frenkel M., Hörchner P., Oost-Harwig O.I., Wilson J., Solid Phase Radioimmunoassay for Determination of Conjugated Cholic Acid in Serum, Clin. Chim. Acta, 73: 277-283 (1976).
[10] Sarafian M.H., Lewis M.R., Pechlivanis A., Ralphs S., McPhail M.J., Patel V.C., Dumas M.-E., Holmes E., Nicholson J.K., Bile Acid Profiling and Quantification in Biofluids Using Ultra-Performance Liquid Chromatography Tandem Mass Spectrometry, Clin. Chim. Acta, 87: 9662-9670 (2015).
[12] Rotariu L., Lagarde F., Jaffrezic-Renault N., Bala C., Electrochemical Biosensors for Fast Detection of Food Contaminants–Trends and Perspective, Trends Analyt. Chem., 79: 80-87 (2016).
[13] Nazari M., Kashanian S., Moradipour P., Maleki N., A Novel Fabrication of Sensor Using ZnO-Al2O3 Ceramic Nanofibers to Simultaneously Detect Catechol and Hydroquinone, J. Electroanal. Chem., 812: 122-131 (2018).
[14] Klouda J., Nesměrák K., Kočovský P., Barek J., Schwarzová-Pecková K., A Novel Voltammetric Approach to the Detection of Primary Bile Acids in Serum Samples, Bioelectrochemistry, 107539 (2020).
[15] Lawrance D., Williamson C., Boutelle M., Cass A., Development of a Disposable Bile Acid Biosensor for Use in the Management of Cholestasis, Anal. Methods., 7: 3714-3719 (2015).
[16] Krajewska B., Application of Chitin-And Chitosan-Based Materials for Enzyme Immobilizations: A Review, Enzyme Microb. Technol., 35: 126-139 (2004).
[17] Wang D., Jiang W., Preparation of Chitosan-Based Nanoparticles for Enzyme Immobilization, Int. J. Biol. Macromol., 126: 1125-1132 (2019).
[18] Brena B., González-Pombo P., Batista-Viera F., Immobilization of Enzymes: A Literature Survey,  Immobilization of Enzymes and Cells, Springer; p. 15-31 (2013).
[19] Wu J.C.Y., Hutchings C.H., Lindsay M.J., Werner C.J., Bundy B.C., Enhanced Enzyme Stability Through Site-Directed Covalent Immobilization, J. Biotechnol., 193: 83-90 (2015).
[21] Kaçar C., Erden P.E., Kılıç E., Amperometric l-lysine Biosensor Based on Carboxylated Multiwalled Carbon Nanotubes-SnO2 Nanoparticles-Graphene Composite, Appl. Surf. Sci., 419: 916-923 (2017).
[23] Benjamin S.R., Vilela R.S., Camargo H., Guedes M., Fernandes K.F., Colmati F., Enzymatic Electrochemical Biosensor Based on Multiwall Carbon Nanotubes and Cerium Dioxide Nanoparticles for Rutin Detection, Int. J. Electrochem. Sci., 13: 563-586 (2018).
[24] Migliorini F.L., Sanfelice R.C., Mercante L.A., Andre R.S., Mattoso L.H., Correa D.S., Urea Impedimetric Biosensing Using Electrospun Nanofibers Modified with Zinc Oxide Nanoparticles, Appl. Surf. Sci., 443: 18-23 (2018).
[25] Mogha N.K., Sahu V., Sharma M., Sharma R.K., Masram D.T., Biocompatible ZrO2-Reduced Graphene Oxide Immobilized AChE Biosensor for Chlorpyrifos Detection, Mater. Des., 111: 312-320 (2016).
[26] Romero-Arcos M., Garnica-Romo M., Martinez-Flores H., Vázquez-Marrufo G., Ramírez-Bon R., González-Hernández J., Barbosa-Cánovas G., Enzyme Immobilization by Amperometric Biosensors with TiO2 Nanoparticles Used to Detect Phenol Compounds, Food Eng. Rev., 8: 235-250 (2016).
[30] Fan Y., Yang X., Yang C., Liu J., Au‐TiO2/Graphene Nanocomposite Film for Electrochemical Sensing of Hydrogen Peroxide and NADH, Electroanalysis, 24: 1334-1339 (2012).
[34] Rao H., Liu Y., Zhong J., Zhang Z., Zhao X., Liu X., Jiang Y., Zou P., Wang X., Wang Y., Gold Nanoparticle/Chitosan@ N, S Co-doped Multiwalled Carbon Nanotubes Sensor: fabrication, Characterization, and Electrochemical Detection of Catechol and Nitrite, ACS Sustain. Chem. Eng., 5: 10926-10939 (2017).
[35] Mehdizadeh B., Maleknia L., Amirabadi A., Shabani M., Glucose Sensing by a Glassy Carbon Electrode Modified with Glucose Oxidase/Chitosan/Graphene Oxide Nanofibers, Diam. Relat. Mater., 109: 108073 (2020).
[36] Rubio-Govea R., Hickey D.P., Garcia-Morales R., Rodriguez-Delgado M., Dominguez-Rovira M.A., Minteer S.D., Ornelas-Soto N., Garcia-Garcia A., MoS2 Nanostructured Materials for Electrode Modification in the Development of a Laccase Based Amperometric Biosensor for Non-Invasive Dopamine Detection, Microchem. J., 155: 104792 (2020).
[38] Venkatachalam N., Palanichamy M., Murugesan V., Sol–Gel Preparation and Characterization of Nanosize TiO2: Its Photocatalytic Performance, Mater. Chem. Phys., 104: 454-459 (2007).
[39] do Amaral Montanheiro T.L., Cristóvan F.H., Machado J.P.B., Tada D.B., Durán N., Lemes A.P., Effect of MWCNT Functionalization on Thermal and Electrical Properties of PHBV/MWCNT Nanocomposites, J. Mater. Res., 30 :55-65 (2015).
[41] Pavia D., Lampman G., Kriz G., Vyvyan J., "Introduction to Spectroscopy Cengage Learning", Ainara López Maestresalas (2014).
[42] Zhang H., Wang X., Li N., Xia J., Meng Q., Ding J., Lu J., Synthesis and Characterization of TiO2 /Graphene Oxide Nanocomposites for Photoreduction of Heavy Metal Ions in Reverse Osmosis Concentrate, RSC Adv., 8: 34241-34251 (2018).
[45] Teodorczyk M., Purdyt W.C., An Amperometric Enzyme Electrode for the Determination of 3α-Hydroxysteroids, Talanta, 37: 795-800 (1990).
[46] Mundaca R., Moreno-Guzmán M., Eguílaz M., Yáñez-Sedeño P., Pingarrón J., Enzyme Biosensor for Androsterone Based on 3α-Hydroxysteroid Dehydrogenase Immobilized onto a Carbon Nanotubes/Ionic Liquid/NAD+ Composite Electrode, Talanta, 99: 697-702 (2012).
[47] Jez J.M., Penning T.M., Engineering Steroid 5β-Reductase Activity into Rat Liver 3α-Hydroxysteroid Dehydrogenase, Biochemistry, 37: 9695-9703 (1998).
[48] Zhang M., Yuan R., Chai Y., Li W., Zhong H., Wang C., Glucose biosensor Based on Titanium Dioxide-Multiwall Carbon Nanotubes-Chitosan Composite and Functionalized Gold Nanoparticles, Bioprocess Biosyst. Eng., 34: 1143-1150 (2011).
[49] Pang Y., Zhang Y., Sun X., Ding H., Ma T., Shen X., Synergistical Accumulation for Electrochemical Sensing of 1-Hydroxypyrene on Electroreduced Graphene Oxide Electrode, Talanta, 192: 387-394 (2019).
[51] Tavakolyan Pour F., Waqifhusain S., Rastegar H., Saber Tehrani M., Abroomand Azar P., Electrochemical Oxidation of Flavonoids and Interaction with DNA on the Surface of Supramolecular Ionic Liquid Grafted on Graphene Modified Glassy Carbon Electrode, Iran. J. Chem. Chem. Eng. (IJCCE), 37(3): 117-125 (2018).
[52] Guan Q., Guo H., Xue R., Wang M., Zhao X., Fan T., Yang W., Xu M., Yang W., Electrochemical Sensor Based on Covalent Organic Frameworks/MWCNT-NH2/AuNPs for Simultaneous Detection of Dopamine and Uric Acid, J. Electroanal. Chem., 880: 114932 (2020)..
[53] Baniasadi M., Maaref H., Dorzadeh A., Mohammad Alizadeh P., A Sensitive SiO2@Fe3O4/GO Nanocomposite Modified Ionic Liquid Carbon Paste Electrode for the Determination of Cabergoline, Iran. J. Chem. Chem. Eng. (IJCCE), 39(4): 11-22 (2020).
[54] Ngamchuea K., Eloul S., Tschulik K., Compton R.G., Planar Diffusion to Macro Disc Electrodes—What Electrode Size is Required for the Cottrell and Randles-Sevcik Equations to Apply Quantitatively?, J. Solid State Electrochem., 18: 3251-3257 (2014).
[55] Tamleh Z., Rafipour R., Kashanian S., Protein-Based Nanobiosensor for Electrochemical Determination of Hydrogen Peroxide, Russ. J. Electrochem., 55: 962-969 (2019).
[56] Zhang G.-H., Cong A.-R., Xu G.-B., Li C.-B., Yang R.-F., Xia T.-A., An Enzymatic Cycling Method for the Determination of Serum Total Bile Acids with Recombinant 3α-Hydroxysteroid Dehydrogenase, Biochem. Biophys. Res. Commun., 326: 87-92
[57] Gültekin A., Karanfil G., Sönmezoğlu S., Say R., Development of a Highly Sensitive MIP Based-QCM Nanosensor For Selective Determination Of Cholic Acid Level in Body Fluids, Mater. Sci. Eng. C, 42: 436-442 (2014).