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Biosensor DNA Elektrokimia untuk Deteksi Makanan Mengandung Babi (Sus scrofa) Menggunakan Screen Printed Carbon Electrode Termodifikasi Emas
Abstract
Metode deteksi berbasis deoxyribonucleic acid (DNA) merupakan metode yang paling akurat, spesifik, dan sensitif untuk mengidentifikasi adanya campuran komponen babi dalam produk pangan. Penelitian ini bertujuan untuk mengembangkan biosensor DNA secara elektrokimia menggunakan Screen Printed Carbon Elektrode termodifikasi emas (SPCE-Au) untuk mendeteksi DNA babi dalam makanan olahan. Permukaan SPCE dimodifikasi dengan emas menggunakan metode adsorbsi pasif, kemudian dikarakterisasi dengan Scanning Electron Microscopy (SEM) dan Differential Pulse Voltammetry (DPV). DNA probe yang spesifik terhadap DNA mitokondria babi diimobilisasi ke permukaan SPCE-Au melalui perantara gugus tiol. Proses hibridisasi DNA probe-DNA komplemen dikarakterisasi menggunakan DPV berdasarkan sinyal oksidasi guanin. Kondisi percobaan dioptimasi menggunakan desain eksperimen Box-Behnken, yaitu konsentrasi DNA probe (0,5; 1,0; 1,5 µg/mL), waktu imobilisasi DNA probe (10, 20, 30 menit), dan waktu hibridisasi DNA probe-DNA komplemen (5, 10, 15 menit). Kondisi optimum digunakan untuk menentukan respons arus oksidasi guanin menggunakan DPV terhadap variasi konsentrasi DNA komplemen. Selanjutnya, biosensor DNA diaplikasikan terhadap sampel bakso yang mengandung campuran daging babi, ayam, dan sapi. Hasil penelitian menunjukkan modifikasi SPCE dengan Au menghasilkan peningkatan arus yang diuji menggunakan sistem redoks K3[Fe(CN)6] secara DPV. Kondisi optimum percobaan adalah: konsentrasi DNA probe 1 µg/mL, waktu imobilisasi DNA probe 20 menit, dan waktu hibridisasi DNA probe-DNA komplemen 10 menit. Biosensor DNA ini memiliki batas deteksi 0,31 µg/mL, batas kuantifikasi 1,06 µg/mL dan recovery 99,2% untuk rentang konsentrasi 0,1 - 2 µg/mL. Deteksi sampel bakso menggunakan biosensor DNA menunjukkan peningkatan respons arus yang signifikan pada sampel yang mengandung 100% daging babi (3,417 µA) dan masih dapat mendeteksi adanya kandungan daging babi sampai 1%. Metoda biosensor DNA babi menggunakan SPCE-Au tanpa indikator dan biokonjugat yang dikembangkan lebih sederhana dan cepat dan mudah untuk diaplikasikan ke sampel nyata.
Electrochemical DNA Biosensor for Detection of Pork (Sus scrofa) Using Gold Modified Screen Printed Carbon Electrode. The DNA-based detection method is the most accurate, specific, and sensitive method for identifying the presence of a mixture of pork components in food products. This study aims to develop an electrochemical DNA biosensor using a gold-modified Screen Printed Carbon Electrode (SPCE-Au) to detect pork DNA in processed food. The surface of the SPCE was modified with gold using a passive adsorption method, then characterized by scanning electron microscopy (SEM) and differential pulse voltammetry (DPV). The probe DNA specific to porcine mitochondrial DNA was immobilized to SPCE-Au surface via an intermediate thiol group. The hybridization of probe DNA-complement DNA was characterized using DPV based on the guanine oxidation signal. The experimental conditions were optimized using the Box-Behnken experimental design by applying probe DNA concentration (0.5; 1.0; 1.5 µg/mL), the immobilization of the probe DNA (10, 20, 30 minutes), and the hybridization of the probe DNA-complement DNA (5, 10, 15 minutes). The optimum conditions were used to determine the DPV current response vs. complementary DNA concentrations. Furthermore, the DNA biosensors were applied to meatball samples containing a mixture of pork, chicken, and beef. The results showed that modification of SPCE with Au produced an increase in current responses tested using K3[Fe(CN)6] redox system by DPV. The optimum conditions of the experiment were: probe DNA concentration was 1 µg/mL, time to immobilize probe DNA was 20 minutes, and time to hybridize probe DNA-complement DNA was 10 minutes. The limit of detection, limit of quantification and percent recovery of DNA biosensor was 0.31 µg/mL, 1.06 µg/mL and 99.2%, respectively. Detection of meatball samples by DNA biosensor showed a significant increase of current response for sample that contains 100% swine DNA (3.417 µA). The biosensor is still able to detect the pork content until 1%. The pork DNA biosensor using SPCE-Au without indicator and bioconjugate developed in this study is simpler and applicable to real samples.
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