The Malaysian Journal of Analytical Sciences Vol 12 No 2 (2008): 473 – 479

 

 

 

 

BIOPENDERIA ELEKTROKIMIA BERASASKAN ENZIM ALKALINE FOSFATASE TERPEGUN UNTUK PENGESANAN KETOKSIKAN ASID

2,4-DIKLOROFENOKSIASETIK

 

(An Electrochemical Biosensor Based on the Immobilisation of Alkaline Phosphatase for the Determination of Toxicity of 2,4-Dichlorophenoxylacetic Acid)

 

Loh Kee Shyuan¹, Lee Yook Heng¹*, Musa Ahmad¹, Salmah Abdul Aziz² & Zamri Ishak²

 

¹Pusat Pengajian Sains Kimia & Teknologi Makanan, Fakulti Sains & Teknologi,

Universiti Kebangsaan Malaysia (UKM), 43600 Bangi, Selangor

² Pusat Penyedilikan Bioteknologi, Institut Penyelidikan & Kemajuan Pertanian Malaysia (MARDI),

P.O. BOX 12301, GPO Kuala Lumpur

 

*Corresponding author: yhl1000@ukm.my

 

Abstract

An electrochemical biosensor based on the immobilisation of alkaline phosphatase was developed for the determination of 2,4-dichlorophenoxylacetic acid (2,4-D). The biosensor was constructed from the immobilization of alkaline phosphatase enzyme onto a screen-printed electrode (SPE). The ascorbic acid 2-phosphate (AA2P) was used as substrate for the enzymic reaction. The enzyme was entrapped in a hybrid sol-gel/chitosan material with certain fixed composition. The determination of toxicity of 2,4-D pesticides quantitatively and qualitatively could be carried out by the inhibition of the alkaline phosphatase. A potential of +600 mV was suitable to be used for the oxidation of the products from the enzyme-substrate reaction, where the reaction pH was at 8.5. The linear response range of the biosensor to the AA2P substrate was 10 mM-80 mM. The inhibition of the alkaline phosphatase enzyme of the 2,4-D biosensor was maximum at 80 ppm 2,4-D (50% inhibition).

 

Keywords: Electrochemical biosensor, alkaline phosphatase, 2,4-dichlorophenoxylacetic acid, sol-gel, chitosan

 

References

1.       Balague, C.E., Ruiz, C.S., Rey, R., Evangelista de Duffard, A.M, Elena, M. & Nader-Macias. 2002. Effect of herbisid 2,4-D dichlorophenoxyacetic acid on uropathogenic Escherichia coli virulence factors. Toxicology.177: 143-155

2.       Sanchez, F.G., Diaz, A.N., Peinado, M.C.R. &  Belledone, C. 2003. Free and sol-gel immobilized alkaline phosphatase-based biosensor for the determination of pesticides and inorganic compounds. Analytical. Chimica. Acta. 484: 45-51

3.       Kaur, J. Singh, K.V., Schmid, A.H., Varshney, G.C., Suri, C.R. & Raje, M. 2004. Atomic force spectroscopy-based study of antibody pesticide interactions for characterization of immunosensor surface. Biosensors & Bioelectronics. 20 : 284-293

4.       Nunes, S. G., Jeanty, G. & Marty, Jean-Louis. 2004. Enzyme immobilization procedures on screen-printed electrodes used for the detection of anticholinesterase pesticides comparative study. Analytica Chimica Acta. 523: 107-115

5.       Mazzei, F., Botre, F., Montilla, S., Pilloton, R., Podesta, E. & Botre, C. 2004. Alkaline phosphatase inhibition based electrochemical sensors for the detection pesticides. Journal of Electroanalytical Chemistry. 574:95-100

6.       Wang, J. & Chen, L. 1999. Amperometric thick-film strip electrodes for monitoring organophosphate nerve agents based on immobilized organophophorus hydrolase. Analytical Chemistry.71: 2246-2249

7.       Satoh, I. & Iijima, Y. 1995. Multi-ion biosensor with use of a hybrid-enzyme membrane. Sensors & Actuators B. 24-25: 103-106

8.       Diaz, A. N., Sanchez, F. G., Ramos, M.C. & Torijas, M. C. 2002. Horseradish peroxidase sol-gel immobilized for chemiluminescence measurements of alkaline-phosphatase activity. Sensors & Actuators B. 82: 176-179.

9.       Guo, Y. & Gudalupe, A. R. 1998. Screen-printable surfactant-induce sol-gel graphite composites for electrochemical sensors. Sensor & Actuators B. 46: 213-219.

10.    Yang, Y.M., Wang, J.W., Tan, R.X. 2004. Immobilization of glucose oxidase on chitosan-SiO2 gel. Enzyme and Microbial Technology. 34: 126-131.

11.    Miao, Y & Tan, S.N. 2001. Amperometric hydrogen peroxide biosensor with silica sol-gel/chitosan film as immobolization matrix. Analytica Chimica Acta. 437: 87-93

12.    Mazzei, F., Botre, F., Montilla, S., Pilloton, R., Podesta, E. & Botre, C. 2004. Alkaline phosphatase inhibition based electrochemical sensors for the detection pesticides. Journal of Electroanalytical Chemistry. 574:95-100.

13.    Wang, J., Pamidi, P.V.A. & Park, D.S. 1996. Screen-printable sol-gel enzyme-containing carbon inks. Analytical Chemistry. 68: 2705-2708

14.    Kokado, A., Arakawa, H. & Maeda, M. 2000. New electrochemical assay of alkaline phosphatase using ascorbic acid 2-phosphate and its application to enzyme immunoassay. Analytica Chimica Acta. 407: 119-125A