AJAC  Vol.4 No.6 , June 2013
Enzyme from Banana (Musa sp.) Extraction Procedures for Sensitive Adrenaline Biosensor Construction
Abstract: For building a biosensor, at least two enzyme sources can be employed. The pure enzyme has features with better selectivity and low stability. Crude extract presents better stability and wrong selectivity. Thus, one intermediate condition can be feasible joining both benefits of crude extract and pure enzymes. For that result, several procedures of extraction and semi purification of polyphenol oxidase (PPO) enzyme from banana (Musa sp.) were studied. The results showed that cleaned enzymatic extracts presented higher specific activities than crude extracts (2.8 up to 5.3 fold), despite the total protein concentration diminishing from 27% up to 72%, indicating that polyphenol oxidase (PPO) enzyme was preserved in them. The biosensors with 125 AU mL-1 containing cleaned enzymatic extracts performed better by grinding or grinding plus sonication for 30 s. They were linear over the ranges of 5.9 × 10-6 mol·L-1 to 1.4 × 10-3 mol·L-1 and 7.9 × 10-8 mol·L-1 to 4.0 × 10-3 mol·L-1, respectively. The limit of Detection (LOD) was 5.9 × 10-6 mol·L-1 and 7.9 × 10-8 mol·L-1, respectively. The LOD obtained is adequate to adrenaline determination on blood and medicinal samples, and were applied in medicinal samples with satisfactory results.
Cite this paper: S. Silva, C. Wisniewski, P. Luccas and C. Magalhães, "Enzyme from Banana (Musa sp.) Extraction Procedures for Sensitive Adrenaline Biosensor Construction," American Journal of Analytical Chemistry, Vol. 4 No. 6, 2013, pp. 293-300. doi: 10.4236/ajac.2013.46037.

[1]   O. D. Leite, O. Fatibello-Filho and A. M. Barbosa, “Determination of Catecholamines in Pharmaceutical Formulations Using a Biosensor Modified with a Crude Extract of Fungi Laccase (Pleurotus ostreatus),” Journal of the Brazilian Chemical Society, Vol. 14, No. 2, 2003, pp. 297-303. doi:10.1590/S0103-50532003000200018.

[2]   C. S. Magalhães, J. S. Garcia, A. S. Lopes, E. C. Figueiredo and M. A. Z. Arruda, “Strategies for Sample Preparation Focusing on Biomolecules Determination/Caracterization,” In: M. A. Z. Arruda, Ed., Trends in Sample Preparation, Nova Science Publishers, New York, 2006, pp. 245-288.

[3]   C. A. Signori, I. C. Vieira and O. Fatibello-Filho, “Quim. Nova,” 1994, p. 38.

[4]   A. W. Lima, V. B. Nascimento, J. J. Pedrotti and L. Angnes, “Coconut-Based Plant Tissue Reactor for Biosensing of Catechol in Flow Injection Analysis,” Analytica Chimica Acta, Vol. 354, No. 1-3, 1997, pp. 325-331. doi:10.1016/S0003-2670(97)00428-5

[5]   A. W. Lima, E. K. Vidsiunas, V. B. Nascimento and L. Angnes, “Vegetable Tissue from Latania sp.: An Extraordinary Source of Naturally Immobilized Enzymes for the Detection of Phenolic Compounds,” Analyst, Vol. 123, No. 11, 1998, pp. 2377-2382.

[6]   O. Fatibello-Filho and I. C. Vieira, “Construction and Analytical Application of a Biosensor Based on Stearic Acid- Graphite Powder Modified with Sweet Potato Tissue in Organic Solvents,” Fresenius Journal of Analytical Chemistry, Vol. 368, No. 4, 2000, pp. 338-343.

[7]   O. Fatibello-Filho and I. C. Vieira, “Uso Analítico de Tecidos e de Extratos Brutos Vegetais Como Fonte Enzimática,” Química Nova, Vol. 25, No. 3, 2002, pp. 455-464. doi:10.1590/S0100-40422002000300019

[8]   P. R. B. O. Marques and H. Yamanaka, “Biosensors Based on the Enzymatic Inhibition Process,” Química Nova, Vol. 31, No. 7, 2008, pp. 1791-1799. doi:10.1590/S0100-40422008000700034

[9]   J. W. Anderson, “Extraction of Enzymes and Subcellular Organelles from Plant Tissues,” Phytochemistry, Vol. 7, No. 11, 1968, pp. 1973-1988.

[10]   A. M. Mayer and E. Harel, “Polyphenol Oxidases in Plants,” Phytochemistry, Vol. 18, No. 2, 1979, pp. 193-215. doi:10.1016/0031-9422(79)80057-6

[11]   A. M. Mayer, “Polyphenol Oxidases in Plants-Recent Progress,” Phytochemistry, Vol. 26, No. 1, 1986, pp. 11-20. doi:10.1016/S0031-9422(00)81472-7

[12]   L. R. V. Mataveli, N. J. Antunes, M. R. P. L. Brigagão, C. S. Magalhães, C. Wisniewski and P. O. Luccas, “Evaluation of a Simple and Low Cost Potentiometric Biosensor for Pharmaceutical and in Vivo Adrenaline Determination,” Biosensors and Bioelectronics, Vol. 26, No. 2, 2010, pp. 798-802. doi:10.1016/j.bios.2010.06.021

[13]   I. C. Vieira and O. Fatibello-Filho, “Biosensor Based on Paraffin/Graphite Modified with Sweet Potato Tissue for the Determination of Hydroquinone in Cosmetic Cream in Organic Phase,” Talanta, Vol. 52, No. 4, 2000, pp. 681-689. doi:10.1016/S0039-9140(00)00420-3

[14]   C. R. Dawson and W. B. Tarpley, “Copper Oxidases,” In: J. B. Summer and K. Myrback, Eds., The Enzymes—Chemistry and Mechanism of Action, Academic Press, New York, 1951, pp. 455-491.

[15]   L. N. Moreira, C. S. Magalhães and P. O. Luccas, “Emprego de Sistema e Análise em Fluxo Contínuo com Biossensor Potenciométrico para Determinação de Adrenalina em Medicamentos,” Infarma, Vol. 16, No. 13-14, 2005, p. 59.

[16]   L. S. Goodman and A. Gilman, “As Bases Farmacológicas da Terapêutica,” 11th Edition, McGraw-Hill Interamericana do Brasil, Rio de Janeiro, 2006.

[17]   S. S. Rosatto, R. S. Freire, N. Duran and L. T. Kubota, “Biossensores Amperométricos para Determinação de Compostos Fenólicos em Amostras de Interesse Ambiental,” Química Nova, Vol. 24, No. 1, 2001, pp. 77-86. doi:10.1590/S0100-40422001000100014

[18]   F. S. Felix, M. Yamashita and L. Angnes, “Epinephrine Quantification in Pharmaceutical Formulations Utilizing Plant Tissue Biosensors,” Biosensors and Bioelectronics, Vol. 21, No. 12, 2006, pp. 2283-2289. doi:10.1016/j.bios.2005.10.025

[19]   Q. L. Yang, P. Atasanov and E. Wilkins, “Needle-Type Lactate Biosensor,” Biosensors and Bioelectronics, Vol. 14, No. 2, 1999, pp. 203-210. doi:10.1016/S0956-5663(98)00109-2

[20]   I. Abdel-Hamid, P. Atanasov and E. Wilkins, “Development of a Needle-Type Biosensor for Intravascular Glucose Monitoring,” Analytica Chimica Acta, Vol. 313, No. 1-2, 1995, pp. 45-54. doi:10.1016/0003-2670(95)00172-V

[21]   E. C. Figueiredo, L. R. Souza, C. S. Magalhães, C. Wisniewski and P. O. Luccas, “A Homemade Autosampler/ Injector Commutator for Flow Injection Analysis,” Journal of Automated Methods & Management in Chemistry, Vol. 2006, 2006, Article ID: 42987.

[22]   G. Li, B. J. Polk, L. A. Meazell and D. W. Hatchett, “ISE Analysis of Hydrogen Sulfide in Cigarette Smoke,” Journal of Chemical Education, Vol. 77, No. 8, 2000, pp. 1049- 1052.

[23]   C. S. Caruzo, I. C. Vieira and O. Fatibello-Filho, “Determination of Epinephrine and Dopamine in Pharmaceutical Formulations Using a Biosensor Based on Carbon Paste Modified with Crude Extract of Cara Root (Dioscorea bulbifera),” Analytical Letters, Vol. 32, No. 1, 1999, pp. 39-50. doi:10.1080/00032719908542597

[24]   C. S. Magalhães and M. A. Z. Arruda, “Sample Preparation for Metalloproteins Analysis: A Case Study Using Horse Chestnuts,” Talanta, Vol. 71, No. 5, 2007, pp. 1958- 1963. doi:10.1016/j.talanta.2006.08.039

[25]   M. Becerra, E. Rodríguez-Belmonte, M. A. Cerdán and M. I. G. Siso, “Extraction of Intracellular Proteins from Kluyveromyces Lactis,” Food Technology and Biotechnology, Vol. 39, No. 2, 2001, pp. 135-139.

[26]   C. C. Nascentes, M. Korn, C. S. Sousa and M. A. Z. Arruda, “Use of Ultrasonic Baths for Analytical Applications: A New Approach for Optimisation Conditions,” Journal of the Brazilian Chemical Society, Vol. 12, No. 2, 200, pp. 157-163.

[27]   K. Kabaha, A. Taralp, I. Cakmak and L. Ozturk, “Accelerated Hydrolysis Method to Estimate the Amino Acid Content of Wheat (Triticum Durum Desf.) Flour Using Microwave Irradiation,” Journal of Agricultural and Food Chemistry, Vol. 59, No. 7, 2011, pp. 2958-2965.

[28]   L. Long, P. Wu, X. Guan, J. Zhang, Y. Jin, Z. Zhang, Y. Wang, Y. Li, J. Chen and W. Wang, “Determination of Protein-Bound Methionine Oxidation in the Hippocampus of Adult and Old Rats by LC-ESI-ITMS Method after Microwave-Assisted Proteolysis,” Applied Biochemistry and Biotechnology, Vol. 399, No. 6, 2011, pp. 2267- 2274.

[29]   S. Shin, A. Lee, S. Lee, K. Lee, J. Kwon, M. Y. Yoon, J. Hong, D. Lee, G. Lee and J. Kim, “A Label-Free Biosensor Assay for Botulinum Neurotoxin B in Food and Human Serum,” Analytical Biochemistry, Vol. 407, No. 2, 2010, pp. 281-288. doi:10.1016/j.ab.2010.08.021

[30]   H. Zheng, J. Yin, Z. Gao, H. Huang, X. Ji and C. Dou, “Disruption of Chlorella Vulgaris Cells for the Release of Biodiesel-Producing Lipids: A Comparison of Grinding, Ultrasonication, Bead Milling, Enzymatic Lysis, and Microwaves,” Applied Biochemistry and Biotechnology, Vol. 164, No. 7, 2011, pp. 1215-1224. doi:10.1007/s12010-011-9207-1

[31]   H. P. Harper, “Manual de Química Fisiológica,” 3rd. Edition, Atheneu, São Paulo, 1973.

[32]   M. M. Bradford, “A Rapid and Sensitive Method for the Quantitation of Microgram Quantities of Protein Utilizing the Principle of Protein-Dye Binding,” Analytical Biochemistry, Vol. 72, No. 1-2, 1976, pp. 248-254. doi:10.1016/0003-2697(76)90527-3

[33]   D. A. M. Zaia, C. T. B. V. Zaia and J. Lichtig, “Determinação de Proteínas Totais via Espectrofometria: Vantagens e Desvantagens dos Métodos Existentes,” Química Nova, Vol. 21, No. 6, 1998, pp. 787-793. doi:10.1590/S0100-40421998000600020

[34]   S. J. Compton and C. G. Jones, “Mechanism of Dye Response and Interference in the Bradford Protein Assay,” Analytical Biochemistry, Vol. 151, No. 2, 1985, pp. 369- 374. doi:10.1016/0003-2697(85)90190-3

[35]   L. R. V. Mataveli, “Medida de Atividade Enzimática de Extrato Bruto de Banana (Musa sp.): Comparação Entre Diferentes Metodologias,” Infarma (Brasília), Vol. 19, No. 9-10, 2007, pp. 33-35.

[36]   J. C. S. Barboza and A. A. Serra, “Ultra-Som I: Influencia do Ultra-Som Na Quimica,” Química Nova, Vol. 15, No. 4, 1992, pp. 302-316.

[37]   R. P. Buck and E. Lindner, “Recommendations for Nomenclature of Ionselective Electrodes (IUPAC Recommendations 1994),” Pure and Applied Chemistry, Vol. 66, No. 12, 1994, pp. 2527-2536. doi:10.1351/pac199466122527

[38]   E. S. Louzada, P. O. Luccas and C. S. Magalhães, “Construção e Caracterização de um Biossensor Potenciométrico Para Determinação de Pirogalol,” Revista Analytica, Vol. 11, 2004, pp. 52-57.

[39]   C. A. S. Perone, M. P. Capobianco and S. P. Junior, “Kinetic Characterization of Enzyme Polyphenol Oxidase, Utilizing Crude Extract of Pell of Stunded Banana (Musa Acuminata),” Revista do Instituto de Ciências da Saúde, Vol. 26, No. 2, 2008, pp. 201-206.

[40]   A. A. M. Chasin, M. Chasin and M. C. Salvadori, “Validação de Métodos Cromatográficos em Análises Toxicológicas,”Revista Brasileira de Toxicologia, Vol. 11, 1998, pp. 1-6.

[41]   “Farmacopéia Brasileira,” 3rd Edition, São Paulo, 1977.

[42]   D. A. Skoog, F. J. Holler and T. A. Nieman, “Principles of Instrumental Analysis,” 5th Edition, Bookman Ed., São Paulo, No. 1, 2006.