Micro-band Boron-doped Diamond Electrode in Capillary Electrophoresis for Simultaneous Detection of AMP, ADP, and ATP
At a Glance
Section titled âAt a Glanceâ| Metadata | Details |
|---|---|
| Publication Date | 2021-04-13 |
| Journal | International Journal of Technology |
| Authors | Putu Udiyani Prayikaputri, Prastika Krisma Jiwanti, Mochammad Arfin Fardiansyah Nasution, Jarnuzi Gunlazuardi, Endang Saepudin |
| Institutions | Keio University, Airlangga University |
| Citations | 5 |
| Analysis | Full AI Review Included |
Executive Summary
Section titled âExecutive SummaryâThis research details the fabrication and application of a micro-band Boron-doped Diamond (BDD) electrode coupled with Capillary Zone Electrophoresis (CZE) for the highly sensitive, simultaneous detection of adenosine monophosphate (AMP), diphosphate (ADP), and triphosphate (ATP).
- Core Achievement: Successful fabrication of a micro-band BDD electrode (effective area 1.11x10-7 m2) using a simple, cost-effective lamination technique (Teflon/silicone sandwich).
- Performance Enhancement: The micro-band geometry utilizes radial diffusion, providing superior sensitivity compared to traditional macro BDD electrodes (e.g., AMP sensitivity gradient 8.80x vs. 7.29x).
- Optimal Conditions: Electrochemical oxidation of APs was optimized at a potential of +1.0 V (vs. Ag/AgCl) and separation achieved using Britton-Robinson buffer at pH 2.0 and 10 kV separation voltage.
- Ultra-low Detection Limits: The system achieved excellent Limits of Detection (LODs) for the adenosine phosphates: 0.004 ”M (AMP), 0.006 ”M (ADP), and 0.011 ”M (ATP).
- Real-World Validation: The method was successfully applied to spiked human urine samples, demonstrating high recovery rates (99.2% to 107.7%) and effective separation of all three APs, plus adenine and guanine.
- Material Quality: Raman and XPS confirmed the high quality of the BDD film, showing high sp3 hybridization and minimal sp2 impurities.
Technical Specifications
Section titled âTechnical Specificationsâ| Parameter | Value | Unit | Context |
|---|---|---|---|
| B/C Doping Ratio | 1% | - | Boron concentration during MPCVD synthesis |
| BDD Film Thickness (Average) | 5 | ”m | Polycrystalline film thickness |
| Micro-band Thickness | ~10 | ”m | Longitudinal section of fabricated electrode |
| BDD Grain Size | ±4-5 | ”m | Homogeneous particle grains on BDD surface |
| Effective Surface Area (Aeff) | 1.11 x 10-7 | m2 | Calculated using Cottrell equation |
| Optimal Detection Potential | +1.0 | V | Amperometry (vs. Ag/AgCl) |
| Optimal Separation Voltage | 10 | kV | Capillary Electrophoresis (CE) |
| Optimal Buffer pH | 2.0 | - | Britton-Robinson buffer for separation |
| Capillary Length | 0.3 | m | Fused silica capillary |
| ATP Diffusion Coefficient (D) | 0.0874 | m2/s | Calculated via amperometric technique |
| Linear Concentration Range | 0.1 to 2.0 | mM | For AMP, ADP, and ATP detection |
| LOD (ATP, Micro-band) | 0.011 | ”M | Limit of Detection (Abstract value) |
| LOD (AMP, Micro-band) | 0.004 | ”M | Limit of Detection (Abstract value) |
| AMP Sensitivity (Micro-band) | 8.80x ± 0.09 | ”A/mM | Gradient of calibration curve (R2 = 0.999) |
| AMP Sensitivity (Macro-electrode) | 7.29x ± 0.05 | ”A/mM | Comparison sensitivity (R2 = 0.999) |
Key Methodologies
Section titled âKey MethodologiesâThe micro-band BDD electrode was fabricated and integrated into a Capillary Electrophoresis (CE) system using the following steps:
- BDD Film Deposition: Polycrystalline BDD film was grown on a Si (100) plate using Microwave Plasma-Assisted Chemical Vapor Deposition (MPCVD). The precursor was methanol solution doped with boron at a B/C ratio of 1%.
- Substrate Etching: The Si substrate was removed by immersing the BDD film in a 1:1 mixture of HF (48%) and HNO3 (60%) for 12 hours.
- Micro-band Fabrication (Lamination): The BDD film and silicon rubber were cut into 1 cm x 1 cm squares. A sandwich structure was created using Teflon support plates in the order: Teflon-silicone-BDD-silicone-Teflon. A copper wire was inserted for electrical contact.
- Electrochemical Characterization: Cyclic Voltammetry (CV) was performed in Phosphate Buffer Solution (PBS) at pH 6.8 to determine the oxidation potential of APs (+0.9 V) and confirm diffusion-controlled kinetics.
- CE Buffer Optimization: Britton-Robinson buffers were tested across pH 2.0 to 10.0. Optimal oxidation current and potential stability were achieved at pH 2.0.
- CE Separation Optimization: Separation voltages ranging from 5 kV to 25 kV were tested. An optimal separation voltage of 10 kV was selected to achieve complete separation of AMP, ADP, and ATP within 30 minutes.
- Amperometric Detection: The micro-band BDD electrode was used as the detector, applying a constant potential of +1.0 V (vs. Ag/AgCl) to measure the oxidation currents of the separated analytes.
Commercial Applications
Section titled âCommercial ApplicationsâThe developed micro-band BDD electrode technology, leveraging the stability and wide potential window of diamond combined with the high sensitivity of microelectrodes, is highly relevant for several high-value analytical and biomedical fields:
- Clinical Diagnostics and Disease Monitoring:
- Detection of APs (AMP, ADP, ATP) in biological fluids (urine, blood plasma) as biomarkers for metabolic disorders, liver disease, epilepsy, Alzheimerâs disease, and Parkinsonâs disease.
- High-throughput screening for drug abuse metabolites.
- Pharmaceutical and Biomedical Research:
- Monitoring extracellular signaling agents (APs) and their metabolites in cellular studies.
- Integration into microdialysis systems for continuous, real-time monitoring of energy metabolites in vivo.
- Microfluidics and Lab-on-a-Chip Devices:
- The small size and robust nature of the micro-band BDD make it ideal for integration into miniaturized analytical systems requiring high-performance electrochemical detection.
- Environmental and Industrial Sensing:
- While focused on biomolecules here, BDD electrodes are generally used for robust electrochemical sensing in harsh environments due to their chemical stability and resistance to fouling.
View Original Abstract
A micro-band boron-doped diamond (BDD) electrode was prepared by sealing a piece of BDD film with an area of 1.11ÂŽ10-7 m2 between two insulating plates, one Teflon and one silicon rubber, to form sandwich-like layers, so the surface area could be investigated. The micro-band BDD was combined with capillary zone electrophoresis as an electrode for the simultaneous detection of adenosine monophosphate (AMP), adenosine diphosphate (ADP), and adenosine triphosphate (ATP) in a solution. These adenosine phosphates can be separated with a 0.3 m-long fused silica capillary using Britton-Robinson buffers at pH 2.0. Current in the concentration range of 0.1 to 2.0 mM were linear with the limits of detection of 0.004 ?M, 0.006 ?M, and 0.011 ?M for AMP, ADP, and ATP, respectively. A comparison with an unmodified BDD as the detector in the same electrophoresis system showed that the micro-band generated better limits of detection (LODs) than the macroelectrode. This method was successfully applied to human urine samples injected with three adenosine phosphates, as well as adenine and guanine, which can be well-separated with recovery percentages of adenine, guanine, AMP, ADP, and ATP of 99.2%, 102.5%, 107.4%, 107.7%, and 105.4%, respectively.