Editorial Feature

Enzyme-Based Electrochemical Biosensors

Electrochemical sensors involve a transduction element encapsulated by a biological recognition layer. The recognition layer interacts with the target analyte and forces a chemical reaction to occur—a process that is interpreted by the transducing element into an electrical wave.

As early as 2005, analytical research has increasingly involved itself in the application of electrochemical sensors in industries ranging from clinical to industrial, environmental, and agricultural analysis. The bio-catalytic sensor (i.e., the use of an enzyme or cell as an immobilized component) is a common type of electrochemical sensor that has recently garnered special attention in research and development.

Enzyme-Based Electrochemical Biosensors

Enzymes help catalyze a chemical reaction in cellular homeostasis. In relation to enzyme-based electrochemical sensors, there is a coupling reaction between an enzyme and an electrode. In a biological cell, enzyme catalysis normally involves thermal energy that forces the enzyme and substrate to bind via van der Walls forces. During the catalytic reaction, the enzyme reduces the activation energy, meaning reactant molecules form a product with the enzymes easier. With enzyme electrodes, however, the enzyme binds to the electrode surface through a number of processes: covalent cross-linking, entrapment with a gel layer on the electrode surface, encapsulation using low thermal energy, or absorption onto a graphite surface. These methods can immobilize an enzyme to an electrode surface.

Methods to bind an enzyme to the surface of an electrode.

Figure 1. Methods to bind an enzyme to the surface of an electrode. Source: Wang, J. (2006). Analytical Electrochemistry. Canada: John Wiley and Sons, Inc.

Electrical Communication

In understanding electrochemical enzyme-based biosensors, an important consideration is the creation of electrical communication between the enzyme active site and the electrode. These enzyme-electrode reactions are facilitated through three primary methods: artificial redox mediators, use of a natural substrate, and the transfer of electrons between the enzyme and the electron. 

Artificial Redox Mediators

During this reaction, the enzyme either generates or consumes a redox-active compound — which is biologically active charge carriers — during the conversion of the target analyte. The active compound is oxidized by the enzyme, generating electrons that charge the transducing element at the electrode surface. This generates an electrical potential that corresponds to the amount of analyte that was oxidized. Application of a mediated system, whereby molecules are used to shift electrons between the redox reaction point of the enzyme and the electrode, requires less operating voltage to power the catalytic reaction. The most common type of redox mediators includes ferrocene and its derivatives, based on their large potential to power a redox reaction.

Elextrical communication via application of artificial redox mediators. Source: Enzyme Electrodes.

Figure 2. Electrical communication via application of artificial redox mediators. Source: Enzyme Electrodes. PowerPoint Presentation. Biosensors and Bioelectronics (2008/2009). Walter Schottky Institut Center for Nanotechnology and Nanomaterials.

Use of a Natural Substrate

One of the most common types of electrochemical bio-sensing reactions involves natural substrates that are oxidized to transfer electrons to molecular oxygen (O2), resulting in the production of hydrogen peroxide (H2O2). Both oxygen and H2O2 are electrochemically active and can transfer electrons into the neighboring electrode to generate an electrical current. Though this method is simple compared to the use of artificial redox mediators, it does have disadvantages, such as a slow response rate, problems with miniaturization, and poor validity. Oxidation of O2 to H2O2 requires a high voltage, which is perhaps one of the biggest drawbacks of the method.

Transfer of Electrons between the Enzyme and the Electron

Recently, researchers have looked into direct electrical communication between the electrode and the enzyme component. One particular enzyme capable of direct electron transfer is the horseradish peroxide enzyme. The enzyme works by catalyzing the reduction of H2O2. This enzyme does not work alone to catalyze the reduction of H2O2; it works as a complex system with an oxidase enzyme. The peroxidase electrode linked to the peroxidase enzyme works at a low voltage, preventing interference of other electroactive species in this catalytic reaction.

Glucose Biosensors

The glucose biosensor is a relevant example of an electrochemical biosensor. The device has a platinum cathode and silver anode electrode, both responsible for carrying an electrical current. At the front end of the biosensor are four layers that enhance the enzyme reaction. The glucose passes through the O-ring layer and the first polycarbonate membrane to the third membrane layer where the glucose is oxidized to H2O2 and gluconic acid. The H2O2 passes through the cellulose acetate membrane to the platinum cathode where it is converted into O2, a hydrogen molecule, and 2 electrons. The third reaction involves a combination of silver nitrate with an electron on the silver anode, a catalytic reaction that generates silver and chlorine. The hydrogen and O2 molecules react on the silver anode at the opposite end of the biosensor, which releases electrons to power an electrical current.

Researches on enzyme-based electrochemical biosensors focused on the development of in-vivo and in-vitro biosensors for investigating a wider range of physiological compounds. Recently, efforts were made to develop sensors that are more sensitive and reliable. New applications of such biosensors were also discovered, including its use in microfluidic platforms to discover pharmaceutical residue in wastewater. 

Sources and Further Reading

  • Newman J.D., Setford S.J. Enzymatic Biosensors. Molecular Biotechnology 2006;32:249.
  • Wang, J. (2006). Analytical Electrochemistry. Canada: John Wiley and Sons, Inc.
  • Wollenberger U. Electrochemical Biosensors – Ways to Improve Sensor Performance. Biotechnology and Genetic Engineering Reviews 1995;13: 237–266.
  • The Enzyme Electrode
  • Wang J. Glucose Biosensors: 40 Years of Advances and Challenges. Electroanalysis 2001;13(12):983–988.

This article was updated on 13th February, 2020.

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